| 1. |
Chie Kikutake, Mikita Suyama, Pan-cancer analysis of whole-genome doubling and its association with patient prognosis., BMC cancer, 10.1186/s12885-023-11132-6, 23, 1, 619-619, 2023.07, BACKGROUND: Whole-genome doubling (WGD) is a common mutation in cancer. Various studies have suggested that WGD is associated with a poor prognosis in cancer. However, the detailed association between WGD occurrence and prognosis remains unclear. In this study, we aimed to elucidate the mechanism by which WGD affects prognosis using sequencing data from the Pan-Cancer Analysis of Whole Genomes (PCAWG) and The Cancer Genome Atlas. METHODS: Whole-genome sequencing data of 23 cancer types were downloaded from PCAWG project. We defined the WGD event in each sample using the WGD status annotated using PCAWG. We used MutationTimeR to predict the relative timings of mutations and loss of heterozygosity (LOH) in WGD, thus evaluating their association with WGD. We also analyzed the association between WGD-associated factors and patient prognosis. RESULTS: WGD was associated with several factors, e.g., length of LOH regions. Survival analysis using WGD-associated factors revealed that longer LOH regions and LOH in chr17 were associated with poor prognosis in samples with WGD (WGD samples) and samples without WGD (nWGD samples). In addition to these two factors, nWGD samples showed that the number of mutations in tumor suppressor genes was associated with prognosis. Moreover, we explored the genes associated with prognosis in both samples separately. CONCLUSION: The prognosis-related factors in WGD samples differed significantly compared with those in nWGD samples. This study emphasizes the need for different treatment strategies for WGD and nWGD samples.. |
| 2. |
Chie Kikutake, Mikita Suyama, Possible involvement of silent mutations in cancer pathogenesis and evolution., Scientific reports, 10.1038/s41598-023-34452-w, 13, 1, 7593-7593, 2023.05, Recent studies have shown that some silent mutations can be harmful to various processes. In this study, we performed a comprehensive in silico analysis to elucidate the effects of silent mutations on cancer pathogenesis using exome sequencing data derived from the Cancer Genome Atlas. We focused on the codon optimality scores of silent mutations, which were defined as the difference between the optimality of synonymous codons, calculated using the codon usage table. The relationship between cancer evolution and silent mutations showed that the codon optimality score of the mutations that occurred later in carcinogenesis was significantly higher than of those that occurred earlier. In addition, mutations with higher scores were enriched in genes involved in the cell cycle and cell division, while those with lower scores were enriched in genes involved in apoptosis and cellular senescence. Our results demonstrate that some silent mutations can be involved in cancer pathogenesis.. |
| 3. |
Hyeonjeong Kim, Mikita Suyama, Genome-wide identification of copy neutral loss of heterozygosity reveals its possible association with spatial positioning of chromosomes., Human Molecular Genetics, 10.1093/hmg/ddac278, 32, 7, 1175-1183, 2023.03, Loss of heterozygosity (LOH) is a genetic alteration that results from the loss of one allele at a heterozygous locus. In particular, copy neutral LOH (CN-LOH) events are generated, for example, by mitotic homologous recombination after monoallelic defection or gene conversion, resulting in novel homozygous locus having two copies of the normal counterpart allele. This phenomenon can serve as a source of genome diversity and is associated with various diseases. To clarify the nature of the CN-LOH such as the frequency, genomic distribution and inheritance pattern, we made use of whole-genome sequencing data of the three-generation CEPH/Utah family cohort, with the pedigree consisting of grandparents, parents and offspring. We identified an average of 40.7 CN-LOH events per individual taking advantage of 285 healthy individuals from 33 families in the cohort. On average 65% of them were classified as gonosomal-mosaicism-associated CN-LOH, which exists in both germline and somatic cells. We also confirmed that the incidence of the CN-LOH has little to do with the parents' age and sex. Furthermore, through the analysis of the genomic region including the CN-LOH, we found that the chance of the occurrence of the CN-LOH tends to increase at the GC-rich locus and/or on the chromosome having a relatively close inter-homolog distance. We expect that these results provide significant insights into the association between genetic alteration and spatial position of chromosomes as well as the intrinsic genetic property of the CN-LOH.. |
| 4. |
Ryusei Kaneko, Ako Matsui, Mahiro Watanabe, Yoshihiro Harada, Mitsuhiro Kanamori, Natsumi Awata, Mio Kawazoe, Tomoaki Takao, Yutaro Kobayashi, Chie Kikutake, Mikita Suyama, Takashi Saito, Takaomi C Saido, Minako Ito, Increased neutrophils in inflammatory bowel disease accelerate the accumulation of amyloid plaques in the mouse model of Alzheimer's disease., Inflammation and regeneration, 10.1186/s41232-023-00257-7, 43, 1, 20-20, 2023.03, BACKGROUND: Alzheimer's disease (AD) is one of the neurodegenerative diseases and characterized by the appearance and accumulation of amyloid-β (Aβ) aggregates and phosphorylated tau with aging. The aggregation of Aβ, which is the main component of senile plaques, is closely associated with disease progression. AppNL-G-F mice, a mouse model of AD, have three familial AD mutations in the amyloid-β precursor gene and exhibit age-dependent AD-like symptoms and pathology. Gut-brain interactions have attracted considerable attention and inflammatory bowel disease (IBD) has been associated with a higher risk of dementia, especially AD, in humans. However, the underlying mechanisms and the effects of intestinal inflammation on the brain in AD remain largely unknown. Therefore, we aimed to investigate the effects of intestinal inflammation on AD pathogenesis. METHODS: Wild-type and AppNL-G-F mice at three months of age were fed with water containing 2% dextran sulfate sodium (DSS) to induce colitis. Immune cells in the brain were analyzed using single-cell RNA sequencing (scRNA-seq) analysis, and the aggregation of Aβ protein in the brain was analyzed via immunohistochemistry. RESULTS: An increase in aggregated Aβ was observed in the brains of AppNL-G-F mice with acute intestinal inflammation. Detailed scRNA-seq analysis of immune cells in the brain showed that neutrophils in the brain increased after acute enteritis. Eliminating neutrophils by antibodies suppressed the accumulation of Aβ, which increased because of intestinal inflammation. CONCLUSION: These results suggest that neutrophils infiltrate the AD brain parenchyma when acute colitis occurs, and this infiltration is significantly related to disease progression. Therefore, we propose that neutrophil-targeted therapies could reduce Aβ accumulation observed in early AD and prevent the increased risk of AD due to colitis.. |
| 5. |
Yasuhide Yoshioka, Keisuke Anzai, Ryosuke Kowada, Ken Hiratsuka, Teppei Hirayabu, Masashi Yasuda, Yasuyuki Ohkawa, Tetsuya Sato, Mikita Suyama, Hideki Yoshida, Masamitsu Yamaguchi, Drosophila transcription factor NF-Y suppresses transcription of the lipase 4 gene, a key gene for lipid storage., Experimental cell research, 10.1016/j.yexcr.2022.113307, 420, 1, 113307-113307, 2022.11, The CCAAT motif-binding factor NF-Y consists of three different subunits, NF-YA, NF-YB, and NF-YC. Although it is suggested that NF-Y activity is essential for normal tissue homeostasis, survival, and metabolic function, its precise role in lipid metabolism is not clarified yet. In Drosophila, eye disc specific knockdown of Drosophila NF-YA (dNF-YA) induced aberrant morphology of the compound eye, the rough eye phenotype in adults and mutation of the lipase 4 (lip4) gene suppressed the rough eye phenotype. RNA-seq analyses with dNF-YA knockdown third instar larvae identified the lip4 gene as one of the genes that are up-regulated by the dNF-YA knockdown. We identified three dNF-Y-binding consensuses in the 5'flanking region of the lip4 gene, and a chromatin immunoprecipitation assay with the specific anti-dNF-YA IgG demonstrated dNF-Y binding to this genomic region. The luciferase transient expression assay with cultured Drosophila S2 cells and the lip4 promoter-luciferase fusion genes with and without mutations in the dNF-Y-binding consensuses showed that each of the three dNF-Y consensus sequences negatively regulated lip4 gene promoter activity. Consistent with these results, qRT-PCR analysis with the dNF-YA knockdown third instar larvae revealed that endogenous lip4 mRNA levels were increased by the knockdown of dNF-YA in vivo. The specific knockdown of dNF-YA in the fat body with the collagen-GAL4 driver resulted in smaller oil droplets in the fat body cells. Collectively, these results suggest that dNF-Y is involved in lipid storage through its negative regulation of lip4 gene transcription.. |
| 6. |
Thomas Fleming, Yukiko Kikuchi, Mikoto Nakajo, Masaya Tachizawa, Tomoaki Inazumi, Soken Tsuchiya, Yukihiko Sugimoto, Daisuke Saito, Mikita Suyama, Yasuyuki Ohkawa, Takashi Baba, Ken-Ichirou Morohashi, Kataaki Okubo, Prostaglandin E2 receptor Ptger4b regulates female-specific peptidergic neurons and female sexual receptivity in medaka., Communications biology, 10.1038/s42003-022-04195-x, 5, 1, 1215-1215, 2022.11, In vertebrates, female receptivity to male courtship is highly dependent on ovarian secretion of estrogens and prostaglandins. We recently identified female-specific neurons in the medaka (Oryzias latipes) preoptic area that express Npba, a neuropeptide mediating female sexual receptivity, in response to ovarian estrogens. Here we show by transcriptomic analysis that these neurons express a multitude of neuropeptides, in addition to Npba, in an ovarian-dependent manner, and we thus termed them female-specific, sex steroid-responsive peptidergic (FeSP) neurons. Our results further revealed that FeSP neurons express a prostaglandin E2 receptor gene, ptger4b, in an ovarian estrogen-dependent manner. Behavioral and physiological examination of ptger4b-deficient female medaka found that they exhibit increased sexual receptivity while retaining normal ovarian function and that their FeSP neurons have reduced firing activity and impaired neuropeptide release. Collectively, this work provides evidence that prostaglandin E2/Ptger4b signaling mediates the estrogenic regulation of FeSP neuron activity and female sexual receptivity.. |
| 7. |
Chie Kikutake, Mikita Suyama, Pan-cancer analysis of mutations in open chromatin regions and their possible association with cancer pathogenesis., Cancer medicine, 10.1002/cam4.4749, 11, 20, 3902-3916, 2022.10, BACKGROUND: Open chromatin is associated with gene transcription. Previous studies have shown that the density of mutations in open chromatin regions is lower than that in flanking regions because of the higher accessibility of DNA repair machinery. However, in several cancer types, open chromatin regions show an increased local density of mutations in activated regulatory regions. Although the mutation distribution within open chromatin regions in cancer cells has been investigated, only few studies have focused on their functional implications in cancer. To reveal the impact of highly mutated open chromatin regions on cancer, we investigated the association between mutations in open chromatin regions and their possible functions. METHODS: Whole-genome sequencing data of 18 cancer types were downloaded from the PanCancer Analysis of Whole Genomes and Catalog of Somatic Mutations in Cancer. We quantified the mutations located in open chromatin regions defined by The Cancer Genome Atlas and classified open chromatin regions into three categories based on the number of mutations. Then, we investigated the chromatin state, amplification, and possible target genes of the open chromatin regions with a high number of mutations. We also analyzed the association between the number of mutations in open chromatin regions and patient prognosis. RESULTS: In some cancer types, the proportion of promoter or enhancer chromatin state in open chromatin regions with a high number of mutations was significantly higher than that in the regions with a low number of mutations. The possible target genes of open chromatin regions with a high number of mutations were more strongly associated with cancer than those of other open chromatin regions. Moreover, a high number of mutations in open chromatin regions was significantly associated with a poor prognosis in some cancer types. CONCLUSIONS: These results suggest that highly mutated open chromatin regions play an important role in cancer pathogenesis and can be effectively used to predict patient prognosis.. |
| 8. |
Miki Inoue, Takashi Baba, Fumiya Takahashi, Miho Terao, Shogo Yanai, Yuichi Shima, Daisuke Saito, Kei Sugihara, Takashi Miura, Shuji Takada, Mikita Suyama, Yasuyuki Ohkawa, Ken-Ichirou Morohashi, Tmsb10 triggers fetal Leydig differentiation by suppressing the RAS/ERK pathway., Communications biology, 10.1038/s42003-022-03941-5, 5, 1, 974-974, 2022.09, Leydig cells in fetal testes play crucial roles in masculinizing fetuses through androgen production. Gene knockout studies have revealed that growth factors are implicated in fetal Leydig cell (FLC) differentiation, but little is known about the mechanisms regulating this process. We investigate this issue by characterizing FLC progenitor cells using single-cell RNA sequencing. The sequence datasets suggest that thymosin β10 (Tmsb10) is transiently upregulated in the progenitors. While studying the function of Tmsb10, we reveal that platelet-derived growth factor (PDGF) regulates ciliogenesis through the RAS/ERK and PI3K/AKT pathways, and thereby promotes desert hedgehog (DHH)-dependent FLC differentiation. Tmsb10 expressed in the progenitor cells induces their differentiation into FLCs by suppressing the RAS/ERK pathway. Through characterizing the transiently expressed Tmsb10 in the FLC progenitors, this study unveils the molecular process of FLC differentiation and shows that it is cooperatively induced by DHH and PDGF.. |
| 9. |
Toru Masuda, Shojiro Haji, Yasuhiro Nakashima, Mariko Tsuda, Daisaku Kimura, Akiko Takamatsu, Norifusa Iwahashi, Hironobu Umakoshi, Motoaki Shiratsuchi, Chie Kikutake, Mikita Suyama, Yasuyuki Ohkawa, Yoshihiro Ogawa, Identification of a drug-response gene in multiple myeloma through longitudinal single-cell transcriptome sequencing., iScience, 10.1016/j.isci.2022.104781, 25, 8, 104781-104781, 2022.08, Despite recent therapeutic advances for multiple myeloma (MM), relapse is very common. Here, we conducted longitudinal single-cell transcriptome sequencing (scRNA-seq) of MM cells from a patient with relapsed MM, treated with multiple anti-myeloma drugs. We observed five subclusters of MM cells, which appeared and/or disappeared in response to the therapeutic pressure, and identified cluster 3 which emerged during lenalidomide treatment and disappeared after proteasome inhibitor (PI) treatment. Among the differentially expressed genes in cluster 3, we found a candidate drug-response gene; pellino E3 ubiquitin-protein ligase family member 2 (PELI2), which is responsible for PI-induced cell death in in vitro assay. Kaplan-Meier survival analysis of database revealed that higher expression of PELI2 is associated with a better prognosis. Our integrated strategy combining longitudinal scRNA-seq analysis, in vitro functional assay, and database analysis would facilitate the understanding of clonal dynamics of MM in response to anti-myeloma drugs and identification of drug-response genes.. |
| 10. |
Naoto Kubota, Mikita Suyama, Mapping of promoter usage QTL using RNA-seq data reveals their contributions to complex traits., PLoS computational biology, 10.1371/journal.pcbi.1010436, 18, 8, e1010436, 2022.08, Genomic variations are associated with gene expression levels, which are called expression quantitative trait loci (eQTL). Most eQTL may affect the total gene expression levels by regulating transcriptional activities of a specific promoter. However, the direct exploration of genomic loci associated with promoter activities using RNA-seq data has been challenging because eQTL analyses treat the total expression levels estimated by summing those of all isoforms transcribed from distinct promoters. Here we propose a new method for identifying genomic loci associated with promoter activities, called promoter usage quantitative trait loci (puQTL), using conventional RNA-seq data. By leveraging public RNA-seq datasets from the lymphoblastoid cell lines of 438 individuals from the GEUVADIS project, we obtained promoter activity estimates and mapped 2,592 puQTL at the 10% FDR level. The results of puQTL mapping enabled us to interpret the manner in which genomic variations regulate gene expression. We found that 310 puQTL genes (16.1%) were not detected by eQTL analysis, suggesting that our pipeline can identify novel variant-gene associations. Furthermore, we identified genomic loci associated with the activity of "hidden" promoters, which the standard eQTL studies have ignored. We found that most puQTL signals were concordant with at least one genome-wide association study (GWAS) signal, enabling novel interpretations of the molecular mechanisms of complex traits. Our results emphasize the importance of the re-analysis of public RNA-seq datasets to obtain novel insights into gene regulation by genomic variations and their contributions to complex traits.. |
| 11. |
Shinichi Yamamoto, Ako Matsui, Masaki Ohyagi, Chie Kikutake, Yoshihiro Harada, Mana Iizuka-Koga, Mikita Suyama, Akihiko Yoshimura, Minako Ito, In Vitro Generation of Brain Regulatory T Cells by Co-culturing With Astrocytes., Frontiers in immunology, 10.3389/fimmu.2022.960036, 13, 960036-960036, 2022.07, Regulatory T cells (Tregs) are normally born in the thymus and activated in secondary lymphoid tissues to suppress immune responses in the lymph node and at sites of inflammation. Tregs are also resident in various tissues or accumulate in damaged tissues, which are now called tissue Tregs, and contribute to homeostasis and tissue repair by interacting with non-immune cells. We have shown that Tregs accumulate in the brain during the chronic phase in a mouse cerebral infarction model, and these Tregs acquire the characteristic properties of brain Tregs and contribute to the recovery of neurological damage by interacting with astrocytes. However, the mechanism of tissue Treg development is not fully understood. We developed a culture method that confers brain Treg characteristics in vitro. Naive Tregs from the spleen were activated and efficiently amplified by T-cell receptor (TCR) stimulation in the presence of primary astrocytes. Furthermore, adding IL-33 and serotonin could confer part of the properties of brain Tregs, such as ST2, peroxisome proliferator-activated receptor γ (PPARγ), and serotonin receptor 7 (Htr7) expression. Transcriptome analysis revealed that in vitro generated brain Treg-like Tregs (induced brain Tregs; iB-Tregs) showed similar gene expression patterns as those in in vivo brain Tregs, although they were not identical. Furthermore, in Parkinson's disease models, in which T cells have been shown to be involved in disease progression, iB-Tregs infiltrated into the brain more readily and ameliorated pathological symptoms more effectively than splenic Tregs. These data indicate that iB-Tregs contribute to our understanding of brain Treg development and could also be therapeutic for inflammatory brain diseases.. |
| 12. |
Norio Kobayashi, Hiroaki Okae, Hitoshi Hiura, Naoto Kubota, Eri H Kobayashi, Shun Shibata, Akira Oike, Takeshi Hori, Chie Kikutake, Hirotaka Hamada, Hirokazu Kaji, Mikita Suyama, Marie-Line Bortolin-Cavaillé, Jérôme Cavaillé, Takahiro Arima, The microRNA cluster C19MC confers differentiation potential into trophoblast lineages upon human pluripotent stem cells., Nature communications, 10.1038/s41467-022-30775-w, 13, 1, 3071-3071, 2022.06, The first cell fate commitment during mammalian development is the specification of the inner cell mass and trophectoderm. This irreversible cell fate commitment should be epigenetically regulated, but the precise mechanism is largely unknown in humans. Here, we show that naïve human embryonic stem (hES) cells can transdifferentiate into trophoblast stem (hTS) cells, but primed hES cells cannot. Our transcriptome and methylome analyses reveal that a primate-specific miRNA cluster on chromosome 19 (C19MC) is active in naïve hES cells but epigenetically silenced in primed ones. Moreover, genome and epigenome editing using CRISPR/Cas systems demonstrate that C19MC is essential for hTS cell maintenance and C19MC-reactivated primed hES cells can give rise to hTS cells. Thus, we reveal that C19MC activation confers differentiation potential into trophoblast lineages on hES cells. Our findings are fundamental to understanding the epigenetic regulation of human early development and pluripotency.. |
| 13. |
Narumi Sakaguchi, Mikita Suyama, Pervasive occurrence of splice-site-creating mutations and their possible involvement in genetic disorders., NPJ genomic medicine, 10.1038/s41525-022-00294-0, 7, 1, 22-22, 2022.03, The search for causative mutations in human genetic disorders has mainly focused on mutations that disrupt coding regions or splice sites. Recently, however, it has been reported that mutations creating splice sites can also cause a range of genetic disorders. In this study, we identified 5656 candidate splice-site-creating mutations (SCMs), of which 3942 are likely to be pathogenic, in 4054 genes responsible for genetic disorders. Reanalysis of exome data obtained from ciliopathy patients led us to identify 38 SCMs as candidate causative mutations. We estimate that, by focusing on SCMs, the increase in diagnosis rate is approximately 5.9-8.5% compared to the number of already known pathogenic variants. This finding suggests that SCMs are mutations worth focusing on in the search for causative mutations of genetic disorders..  |
| 14. |
Shintaro Mise, Akinobu Matsumoto, Keisuke Shimada, Toshiaki Hosaka, Masatomo Takahashi, Kazuya Ichihara, Hideyuki Shimizu, Chisa Shiraishi, Daisuke Saito, Mikita Suyama, Tomoharu Yasuda, Toru Ide, Yoshihiro Izumi, Takeshi Bamba, Tomomi Kimura-Someya, Mikako Shirouzu, Haruhiko Miyata, Masahito Ikawa, Keiichi I Nakayama, Kastor and Polluks polypeptides encoded by a single gene locus cooperatively regulate VDAC and spermatogenesis., Nature communications, 10.1038/s41467-022-28677-y, 13, 1, 1071-1071, 2022.02, Although several long noncoding RNAs (lncRNAs) have recently been shown to encode small polypeptides, those in testis remain largely uncharacterized. Here we identify two sperm-specific polypeptides, Kastor and Polluks, encoded by a single mouse locus (Gm9999) previously annotated as encoding a lncRNA. Both Kastor and Polluks are inserted in the outer mitochondrial membrane and directly interact with voltage-dependent anion channel (VDAC), despite their different amino acid sequences. Male VDAC3-deficient mice are infertile as a result of reduced sperm motility due to an abnormal mitochondrial sheath in spermatozoa, and deficiency of both Kastor and Polluks also severely impaired male fertility in association with formation of a similarly abnormal mitochondrial sheath. Spermatozoa lacking either Kastor or Polluks partially recapitulate the phenotype of those lacking both. Cooperative function of Kastor and Polluks in regulation of VDAC3 may thus be essential for mitochondrial sheath formation in spermatozoa and for male fertility.. |
| 15. |
Zhuo Qu, Narumi Sakaguchi, Chie Kikutake, Mikita Suyama, Genome-wide identification of exon extension/shrinkage events induced by splice-site-creating mutations., RNA biology, 10.1080/15476286.2022.2139111, 19, 1, 1143-1152, 2022.01, Mutations that affect phenotypes have been identified primarily as those that directly alter amino acid sequences or disrupt splice sites. However, some mutations not located in functionally important sites can also affect phenotypes, such as splice-site-creating mutations (SCMs). To investigate how frequent exon extension/shrinkage events induced by SCMs occur in normal individuals, we used personal genome sequencing data and transcriptome data of the corresponding individuals and identified 371 exon extension/shrinkage events in normal individuals. This number was about three times higher than the number of pseudo-exon activation events identified in the previous study. The average numbers of exon extension and exon shrinkage events in each sample were 3.3 and 11.2, respectively. We also evaluated the impact of exon extension/shrinkage events on the resulting transcripts and their protein products and found that 40.2% of the identified events may have possible functional impacts by either generating premature termination codons in transcripts or affecting protein domains. Our results indicated that a certain fraction of SCMs identified in this study can be pathogenic mutations by creating novel splice sites.. |
| 16. |
Antonius Christianto, Takashi Baba, Fumiya Takahashi, Kai Inui, Miki Inoue, Mikita Suyama, Yusuke Ono, Yasuyuki Ohkawa, Ken-Ichirou Morohashi, Sex differences in metabolic pathways are regulated by Pfkfb3 and Pdk4 expression in rodent muscle., Communications biology, 10.1038/s42003-021-02790-y, 4, 1, 1264-1264, 2021.11, Skeletal muscles display sexually dimorphic features. Biochemically, glycolysis and fatty acid β-oxidation occur preferentially in the muscles of males and females, respectively. However, the mechanisms of the selective utilization of these fuels remains elusive. Here, we obtain transcriptomes from quadriceps type IIB fibers of untreated, gonadectomized, and sex steroid-treated mice of both sexes. Analyses of the transcriptomes unveil two genes, Pfkfb3 (phosphofructokinase-2) and Pdk4 (pyruvate dehydrogenase kinase 4), that may function as switches between the two sexually dimorphic metabolic pathways. Interestingly, Pfkfb3 and Pdk4 show male-enriched and estradiol-enhanced expression, respectively. Moreover, the contribution of these genes to sexually dimorphic metabolism is demonstrated by knockdown studies with cultured type IIB muscle fibers. Considering that skeletal muscles as a whole are the largest energy-consuming organs, our results provide insights into energy metabolism in the two sexes, during the estrus cycle in women, and under pathological conditions involving skeletal muscles.. |
| 17. |
Naoki Haratake, Qingjiang Hu, Tatsuro Okamoto, Tomoko Jogo, Gouji Toyokawa, Fumihiko Kinoshita, Tomoyoshi Takenaka, Tetsuzo Tagawa, Norifumi Iseda, Shinji Itoh, Yuichi Yamada, Yoshinao Oda, Mototsugu Shimokawa, Chie Kikutake, Mikita Suyama, Motoko Unoki, Hiroyuki Sasaki, Masaki Mori, Identification of SLC38A7 as a Prognostic Marker and Potential Therapeutic Target of Lung Squamous Cell Carcinoma., Annals of surgery, 10.1097/SLA.0000000000005001, 274, 3, 500-507, 2021.09, BACKGROUND: No effective molecular targeted therapy has been established for SCC. We conducted a comprehensive study of SCC patients using RNA-sequencing and TCGA dataset to clarify the driver oncogene of SCC. METHOD: Forty-six samples of 23 patients were totally analyzed with RNA-sequencing. We then searched for candidate-oncogenes of SCC using the TCGA database. To identify candidate oncogenes, we used the following 2 criteria: (1) the genes of interest were overexpressed in tumor tissues of SCC patients in comparison to normal tissues; and (2) using an integrated mRNA expression and DNA copy number profiling analysis using the TCGA dataset, the DNA copy number of the genes was positively correlated with the mRNA expression. RESULT: We identified 188 candidate-oncogenes. Among those, the high expression of SLC38A7 was a strong prognostic marker that was significantly associated with a poor prognosis in terms of both overall survival (OS) and recurrence-free survival in the TCGA dataset (P |
| 18. |
Akihiro Nita, Akinobu Matsumoto, Ronghao Tang, Chisa Shiraishi, Kazuya Ichihara, Daisuke Saito, Mikita Suyama, Tomoharu Yasuda, Gaku Tsuji, Masutaka Furue, Bumpei Katayama, Toshiyuki Ozawa, Teruasa Murata, Teruki Dainichi, Kenji Kabashima, Atsushi Hatano, Masaki Matsumoto, Keiichi I Nakayama, A ubiquitin-like protein encoded by the "noncoding" RNA TINCR promotes keratinocyte proliferation and wound healing., PLoS genetics, 10.1371/journal.pgen.1009686, 17, 8, e1009686, 2021.08, Although long noncoding RNAs (lncRNAs) are transcripts that do not encode proteins by definition, some lncRNAs actually contain small open reading frames that are translated. TINCR (terminal differentiation-induced ncRNA) has been recognized as a lncRNA that contributes to keratinocyte differentiation. However, we here show that TINCR encodes a ubiquitin-like protein that is well conserved among species and whose expression was confirmed by the generation of mice harboring a FLAG epitope tag sequence in the endogenous open reading frame as well as by targeted proteomics. Forced expression of this protein promoted cell cycle progression in normal human epidermal keratinocytes, and mice lacking this protein manifested a delay in skin wound healing associated with attenuated cell cycle progression in keratinocytes. We termed this protein TINCR-encoded ubiquitin-like protein (TUBL), and our results reveal a role for TINCR in the regulation of keratinocyte proliferation and skin regeneration that is dependent on TUBL.. |
| 19. |
Daiki Kajioka, Kentaro Suzuki, Shoko Matsushita, Shinjiro Hino, Tetsuya Sato, Shuji Takada, Kyoichi Isono, Toru Takeo, Mizuki Kajimoto, Naomi Nakagata, Mitsuyoshi Nakao, Mikita Suyama, Tony DeFalco, Shinichi Miyagawa, Gen Yamada, Sexual fate of murine external genitalia development: Conserved transcriptional competency for male-biased genes in both sexes., Proceedings of the National Academy of Sciences of the United States of America, 10.1073/pnas.2024067118, 118, 23, 2021.06, Testicular androgen is a master endocrine factor in the establishment of external genital sex differences. The degree of androgenic exposure during development is well known to determine the fate of external genitalia on a spectrum of female- to male-specific phenotypes. However, the mechanisms of androgenic regulation underlying sex differentiation are poorly defined. Here, we show that the genomic environment for the expression of male-biased genes is conserved to acquire androgen responsiveness in both sexes. Histone H3 at lysine 27 acetylation (H3K27ac) and H3K4 monomethylation (H3K4me1) are enriched at the enhancer of male-biased genes in an androgen-independent manner. Specificity protein 1 (Sp1), acting as a collaborative transcription factor of androgen receptor, regulates H3K27ac enrichment to establish conserved transcriptional competency for male-biased genes in both sexes. Genetic manipulation of MafB, a key regulator of male-specific differentiation, and Sp1 regulatory MafB enhancer elements disrupts male-type urethral differentiation. Altogether, these findings demonstrate conservation of androgen responsiveness in both sexes, providing insights into the regulatory mechanisms underlying sexual fate during external genitalia development.. |
| 20. |
Chie Kikutake, Minako Yoshihara, Mikita Suyama, Pan-cancer analysis of non-coding recurrent mutations and their possible involvement in cancer pathogenesis., NAR cancer, 10.1093/narcan/zcab008, 3, 1, zcab008, 2021.03, Cancer-related mutations have been mainly identified in protein-coding regions. Recent studies have demonstrated that mutations in non-coding regions of the genome could also be a risk factor for cancer. However, the non-coding regions comprise 98% of the total length of the human genome and contain a huge number of mutations, making it difficult to interpret their impacts on pathogenesis of cancer. To comprehensively identify cancer-related non-coding mutations, we focused on recurrent mutations in non-coding regions using somatic mutation data from COSMIC and whole-genome sequencing data from The Cancer Genome Atlas (TCGA). We identified 21 574 recurrent mutations in non-coding regions that were shared by at least two different samples from both COSMIC and TCGA databases. Among them, 580 candidate cancer-related non-coding recurrent mutations were identified based on epigenomic and chromatin structure datasets. One of such mutation was located in RREB1 binding site that is thought to interact with TEAD1 promoter. Our results suggest that mutations may disrupt the binding of RREB1 to the candidate enhancer region and increase TEAD1 expression levels. Our findings demonstrate that non-coding recurrent mutations and coding mutations may contribute to the pathogenesis of cancer.. |
| 21. |
Narumi Sakaguchi, Mikita Suyama, In silico identification of pseudo-exon activation events in personal genome and transcriptome data., RNA biology, 10.1080/15476286.2020.1809195, 18, 3, 382-390, 2021.03, Causative mutations for human genetic disorders have mainly been identified in exonic regions that code for amino acid sequences. Recently, however, it has been reported that mutations in deep intronic regions can also cause certain human genetic disorders by creating novel splice sites, leading to pseudo-exon activation. To investigate how frequently pseudo-exon activation events occur in normal individuals, we conducted in silico identification of such events using personal genome data and corresponding high-quality transcriptome data. With rather stringent conditions, on average, 2.6 pseudo-exon activation events per individual were identified. More pseudo-exon activation events were found in 5' donor splice sites than in 3' acceptor splice sites. Although pseudo-exon activation events have sporadically been reported as causative mutations in genetic disorders, it is revealed in this study that such events can be observed in normal individuals at a certain frequency. We estimate that human genomes typically contain on average at least 10 pseudo-exon activation events. The actual number should be higher than this, because we used stringent criteria to identify pseudo-exon activation events. This suggests that it is worth considering the possibility of pseudo-exon activation when searching for causative mutations of genetic disorders if candidate mutations are not identified in coding regions or RNA splice sites.. |
| 22. |
Hyeonjeong Kim, Minako Yoshihara, Mikita Suyama, Comparative genomic analysis of inbred rat strains reveals the existence of ancestral polymorphisms, Mammalian Genome, 10.1007/s00335-020-09831-7, 31, 3-4, 86-94, 2020.04, In an alignment of closely related genomic sequences, the existence of discordant mutation sites, which do not reflect the phylogenetic relationship of the genomes, is often observed. Although these discordant mutation sites are thought to have emerged by ancestral polymorphism or gene flow, their frequency and distribution in the genome have not yet been analyzed in detail. Using the genome sequences of all protein coding genes of 25 inbred rat strains, we analyzed the frequency and genome-wide distribution of the discordant mutation sites. From the comparison of different substrains, it was found that these loci are not substrain specific, but are common among different groups of substrains, suggesting that the discordant sites might have mainly emerged through ancestral polymorphism. It was also revealed that the discordant sites are not uniformly distributed along chromosomes, but are concentrated at certain genomic loci, such as RT1, major histocompatibility complex of rats, and olfactory receptors, indicating that genes known to be highly polymorphic tend to have more discordant sites. Our results also showed that loci with a high density of discordant sites are also rich in heterozygous variants, even though these are inbred strains.. |
| 23. |
Shinya Akatsuka, Guang Hua Li, Shinichi Kawaguchi, Takashi Takahashi, Minako Yoshihara, Mikita Suyama, Shinya Toyokuni, Augmented oxidative stress increases 8-oxoguanine preferentially in the transcriptionally active genomic regions, Free Radical Research, 10.1080/10715762.2020.1733548, 2020.01, 8-Oxoguanine (8-oxoG) is the most common DNA base modification in the mammalian genome, associated with oxidative stress. Here we analysed the alterations in the distribution of 8-oxoG across the entire murine genome, before and after an elevation of oxidative stress by the use of ferric nitrilotriacetate (Fe-NTA) as an oxidative stress inducer in the renal proximal tubules. We isolated DNA fragments containing 8-oxoGs with immunoprecipitation from the murine genome, and amplified them by PCR for a distribution analysis with microarray-based comparative genomic hybridisation. The distribution profiles revealed that frequencies of 8-oxoG fluctuated with a cycle of 1–10 Mb along the chromosomes and the amplitude of the fluctuation was reduced after Fe-NTA administration. The distributions of 8-oxoG along the entire genome in the control and oxidatively stressed conditions were negatively correlated with that of gene density but positively correlated with that of Lamin B1 interaction, which corresponds to lamina-associated domains. These results on the murine genome were consistent with those on the rat genome we previously reported. We further discovered a negative correlation between the distributions of 8-oxoG and transcriptional activity along the genome. Finally, a comparison of the distributions before and after Fe-NTA administration suggested that 8-oxoGs are generated in response to the augmented oxidative stress preferentially in the transcriptionally active genomic regions, where 8-oxoGs have been less accumulated in the control condition.. |
| 24. |
Naoto Kubota, Mikita Suyama, An integrated analysis of public genomic data unveils a possible functional mechanism of psoriasis risk via a long-range ERRFI1 enhancer., BMC medical genomics, 10.1186/s12920-020-0662-9, 13, 1, 8-8, 2020.01, BACKGROUND: Psoriasis is a chronic inflammatory skin disease, for which genome-wide association studies (GWAS) have identified many genetic variants as risk markers. However, the details of underlying molecular mechanisms, especially which variants are functional, are poorly understood. METHODS: We utilized a computational approach to survey psoriasis-associated functional variants that might affect protein functions or gene expression levels. We developed a pipeline by integrating publicly available datasets provided by GWAS Catalog, FANTOM5, GTEx, SNP2TFBS, and DeepBlue. To identify functional variants on exons or splice sites, we used a web-based annotation tool in the Ensembl database. To search for noncoding functional variants within promoters or enhancers, we used eQTL data calculated by GTEx. The data of variants lying on transcription factor binding sites provided by SNP2TFBS were used to predict detailed functions of the variants. RESULTS: We discovered 22 functional variant candidates, of which 8 were in noncoding regions. We focused on the enhancer variant rs72635708 (T > C) in the 1p36.23 region; this variant is within the enhancer region of the ERRFI1 gene, which regulates lipid metabolism in the liver and skin morphogenesis via EGF signaling. Further analysis showed that the ERRFI1 promoter spatially contacts with the enhancer, despite the 170 kb distance between them. We found that this variant lies on the AP-1 complex binding motif and may modulate binding levels. CONCLUSIONS: The minor allele rs72635708 (rs72635708-C) might affect the ERRFI1 promoter activity, which results in unstable expression of ERRFI1, enhancing the risk of psoriasis via disruption of lipid metabolism and skin cell proliferation. Our study represents a successful example of predicting molecular pathogenesis by integration and reanalysis of public data.. |
| 25. |
Masami Miki, Takamasa Oono, Nao Fujimori, Takehiro Takaoka, Ken Kawabe, Yoshihiro Miyasaka, Takao Ohtsuka, Daisuke Saito, Masafumi Nakamura, Yasuyuki Ohkawa, Yoshinao Oda, Mikita Suyama, Tetsuhide Ito, Yoshihiro Ogawa, CLEC3A, MMP7, and LCN2 as novel markers for predicting recurrence in resected G1 and G2 pancreatic neuroendocrine tumors., Cancer medicine, 10.1002/cam4.2232, 8, 8, 3748-3760, 2019.07, Although the postoperative recurrence rate for pancreatic neuroendocrine tumors (PNETs) is reported to be 13.5%-30%, the paucity of valuable biomarkers to predict recurrence poses a problem for the early detection of relapse. Hence, this study aimed to identify new biomarkers to predict the recurrence of PNETs. We performed RNA sequencing (RNA-Seq) on RNA isolated from frozen primary tumors sampled from all localized G1/G2 PNETs resected curatively from 1998 to 2015 in our institution. We calculated differentially expressed genes (DEGs) in tumor with and without recurrence (≥3 years) for the propensity-matched cohort. Gene ontology analysis for the identified DEGs was also performed. Furthermore, we evaluated the expression levels of candidate genes as recurrence predictors via immunostaining. Comparison of transcriptional levels in tumors with and without recurrence identified 166 DEGs. Up- and downregulated genes with high significance in these tumors were mainly related to extracellular organization and cell adhesion, respectively. We observed the top three upregulated genes, C-type lectin domain family 3 member A (CLEC3A), matrix metalloproteinase-7 (MMP7), and lipocalin2 (LCN2) immunohistochemically and compared their levels in recurrent and nonrecurrent tumors. Significantly higher recurrence rate was shown in patients with positive expression of CLEC3A (P = 0.028), MMP7 (P = 0.003), and LCN2 (P = 0.040) than that with negative expression. We identified CLEC3A, MMP7, and LCN2 known to be associated with the phosphatidylinositol-3-kinase/Akt pathway, as potential novel markers to predict the postoperative recurrence of PNETs.. |
| 26. |
Takashi Ishiuchi, Hiroaki Ohishi, Tetsuya Sato, Satoshi Kamimura, Masayoshi Yorino, Shusaku Abe, Atsushi Suzuki, Teruhiko Wakayama, Mikita Suyama, Hiroyuki Sasaki, Zfp281 Shapes the Transcriptome of Trophoblast Stem Cells and Is Essential for Placental Development, Cell Reports, 10.1016/j.celrep.2019.04.028, 27, 6, 1742-1754.e6, 2019.05, Placental development is a key event in mammalian reproduction and embryogenesis. However, the molecular basis underlying placental development is not fully understood. Here, we conduct a forward genetic screen to identify regulators for extraembryonic development and identify Zfp281 as a key factor. Zfp281 overexpression in mouse embryonic stem cells facilitates the induction of trophoblast stem-like cells. Zfp281 is preferentially expressed in the undifferentiated trophoblast stem cell population in an FGF-dependent manner, and disruption of Zfp281 in mice causes severe defects in early placental development. Consistently, Zfp281-depleted trophoblast stem cells exhibit defects in maintaining the transcriptome and differentiation capacity. Mechanistically, Zfp281 interacts with MLL or COMPASS subunits and occupies the promoters of its target genes. Importantly, ZNF281, the human ortholog of this factor, is required to stabilize the undifferentiated status of human trophoblast stem cells. Thus, we identify Zfp281 as a conserved factor for the maintenance of trophoblast stem cell plasticity. Ishiuchi et al. demonstrate that Zfp281 regulates gene expression through the interaction with MLL or COMPASS in trophoblast stem (TS) cells. Depletion of Zfp281 impairs TS cell plasticity in vitro and in vivo. Knockdown of ZNF281 downregulates a set of genes enriched in undifferentiated human TS cells, including ELF5 and LIN28A.. |
| 27. |
Yuko Katoh-Fukui, Takashi Baba, Tetsuya Sato, Hiroyuki Otake, Yuko Nagakui-Noguchi, Miyuki Shindo, Mikita Suyama, Yasuyuki Ohkawa, Hideki Tsumura, Ken-Ichirou Morohashi, Maki Fukami, Mouse polycomb group gene Cbx2 promotes osteoblastic but suppresses adipogenic differentiation in postnatal long bones, Bone, 10.1016/j.bone.2018.10.021, 120, 219-231, 2019.03, A set of key developmental genes is essential for skeletal growth from multipotent progenitor cells at weaning. Polycomb group proteins, which regulate such genes contributes to the cell lineage commitment and subsequent differentiation via epigenetic chromatin modification and remodeling. However, it is unclear which cell lineage and gene sets are targeted by polycomb proteins during skeletal growth. We now report that mice deficient in a polycomb group gene Cbx2cterm/cterm exhibited skeletal hypoplasia in the tibia, femur, and cranium. Long bone cavities in these mice contained fewer multipotent mesenchymal stromal cells. RNA-sequencing of bone marrow cells showed downregulation and upregulation of osteoblastic and adipogenic genes, respectively. Furthermore, the expression levels of genes specifically expressed in B-cell precursors were decreased. Forced expression of Cbx2 in Cbx2cterm/cterm bone marrow stromal cell recovered fibroblastic colony formation and suppressed adipogenic differentiation. Collectively, our results suggest that Cbx2 controls the maintenance and adipogenic differentiation of mesenchymal stromal cells in the bone marrow.. |
| 28. |
Mariko Kikuchi, Toshiya Nishimura, Daisuke Saito, Shuji Shigenobu, Ritsuko Takada, José Arturo Gutierrez-Triana, Juan Luis Mateo Cerdán, Shinji Takada, Joachim Wittbrodt, Mikita Suyama, Minoru Tanaka, Novel components of germline sex determination acting downstream of foxl3 in medaka., Developmental biology, 10.1016/j.ydbio.2018.10.019, 445, 1, 80-89, 2019.01, Germline sex determination is an essential process for the production of sexually dimorphic gametes. In medaka, Forkhead box L3 (foxl3) was previously identified as a germ cell-intrinsic regulator of sex determination that suppresses the initiation of spermatogenesis in female germ cells. To reveal the molecular mechanism of germline sex determination by foxl3, we conducted the following four analyses: Comparison of transcriptomes between wild-type and foxl3-mutant germ cells; epistatic analysis; identification of the FOXL3-binding motif; and ChIP-qPCR assay using a FOXL3-monoclonal antibody. We identified two candidate genes acting downstream of foxl3: Rec8a and fbxo47. It has been known that Rec8 regulates sister chromatid cohesion and Fbxo47 acts as a ubiquitin E3 ligase. These functions have not been, however, associated with sexual differentiation in germ cells. Our results uncover novel components acting downstream of foxl3, providing insights into the mechanism of germline sex determination.. |
| 29. |
Yuichi Shima, Kanako Miyabayashi, Tetsuya Sato, Mikita Suyama, Yasuyuki Ohkawa, Masao Doi, Hitoshi Okamura, Kentaro Suzuki, Fetal leydig cells dedifferentiate and serve as adult leydig stem cells, Development (Cambridge), 10.1242/dev.169136, 145, 23, 2018.12, Previous studies have established that fetal Leydig cells (FLCs) and adult Leydig cells (ALCs) show distinct functional characteristics. However, the lineage relationship between FLCs and ALCs has not been clarified yet. Here, we reveal that a subset of FLCs dedifferentiate at fetal stages to give rise to ALCs at the pubertal stage. Moreover, the dedifferentiated cells contribute to the peritubular myoid cell and vascular pericyte populations in the neonatal testis, and these nonsteroidogenic cells serve as potential ALC stem cells. We generated FLC lineage-specific Nr5a1 (Ad4BP/SF-1) gene-disrupted mice and mice lacking the fetal Leydig enhancer (FLE) of the Nr5a1 gene. Phenotypes of these mice support the conclusion that most of the ALCs arise from dedifferentiated FLCs, and that the FLE of the Nr5a1 gene is essential for both initial FLC differentiation and pubertal ALC redifferentiation.. |
| 30. |
Takashi Baba, Hiroyuki Otake, Miki Inoue, Tetsuya Sato, Yasuhiro Ishihara, Ju Yeon Moon, Megumi Tsuchiya, Kanako Miyabayashi, Hidesato Ogawa, Yuichi Shima, Lixiang Wang, Ryuichiro Sato, Takeshi Yamazaki, Mikita Suyama, Masatoshi Nomura, Man Ho Choi, Yasuyuki Ohkawa, Ken ichirou Morohashi, Ad4BP/SF-1 regulates cholesterol synthesis to boost the production of steroids, Communications Biology, 10.1038/s42003-018-0020-z, 1, 1, 2018.12, Housekeeping metabolic pathways such as glycolysis are active in all cell types. In addition, many types of cells are equipped with cell-specific metabolic pathways. To properly perform their functions, housekeeping and cell-specific metabolic pathways must function cooperatively. However, the regulatory mechanisms that couple metabolic pathways remain largely unknown. Recently, we showed that the steroidogenic cell-specific nuclear receptor Ad4BP/SF-1, which regulates steroidogenic genes, also regulates housekeeping glycolytic genes. Here, we identify cholesterogenic genes as the targets of Ad4BP/SF-1. Further, we reveal that Ad4BP/SF-1 regulates Hummr, a candidate mediator of cholesterol transport from endoplasmic reticula to mitochondria. Given that cholesterol is the starting material for steroidogenesis and is synthesized from acetyl-CoA, which partly originates from glucose, our results suggest that multiple biological processes involved in synthesizing steroid hormones are governed by Ad4BP/SF-1. To our knowledge, this study provides the first example where housekeeping and cell-specific metabolism are coordinated at the transcriptional level.. |
| 31. |
Yuichi Shima, Kanako Miyabayashi, Tetsuya Sato, Mikita Suyama, Yasuyuki Ohkawa, Masao Doi, Hitoshi Okamura, Kentaro Suzuki, Fetal Leydig cells dedifferentiate and serve as adult Leydig stem cells., Development (Cambridge, England), 10.1242/dev.169136, 145, 23, 2018.12, Previous studies have established that fetal Leydig cells (FLCs) and adult Leydig cells (ALCs) show distinct functional characteristics. However, the lineage relationship between FLCs and ALCs has not been clarified yet. Here, we reveal that a subset of FLCs dedifferentiate at fetal stages to give rise to ALCs at the pubertal stage. Moreover, the dedifferentiated cells contribute to the peritubular myoid cell and vascular pericyte populations in the neonatal testis, and these non-steroidogenic cells serve as potential ALC stem cells. We generated FLC lineage-specific Nr5a1 (Ad4BP/SF-1) gene-disrupted mice and mice lacking the fetal Leydig enhancer (FLE) of the Nr5a1 gene. Phenotypes of these mice support the conclusion that most of the ALCs arise from dedifferentiated FLCs, and that the FLE of the Nr5a1 gene is essential for both initial FLC differentiation and pubertal ALC redifferentiation.. |
| 32. |
Chie Kikutake, Minako Yoshihara, Tetsuya Sato, Daisuke Saito, Mikita Suyama, Pan-cancer analysis of intratumor heterogeneity associated with patient prognosis using multidimensional measures., Oncotarget, 10.18632/oncotarget.26485, 9, 102, 37689-37699, 2018.12, Human cancers accumulate various mutations during development and consist of highly heterogeneous cell populations. This phenomenon is called intratumor heterogeneity (ITH). ITH is known to be involved in tumor growth, progression, invasion, and metastasis, presenting obstacles to accurate diagnoses and effective treatments. Numerous studies have explored the dynamics of ITH, including constructions of phylogenetic trees in cancer samples using multiregional ultradeep sequencing and simulations of evolution using statistical models. Although ITH is associated with prognosis, it is still challenging to use the characteristics of ITH as prognostic factors because of difficulties in quantifying ITH precisely. In this study, we analyzed the relationship between patient prognosis and the distribution of variant allele frequencies (VAFs) in cancer samples (n = 6,064) across 16 cancer types registered in The Cancer Genome Atlas. To measure VAF distributions multidimensionally, we adopted parameters that define the shape of VAF distributions and evaluated the relationships between these parameters and prognosis. In seven cancer types, we found significant relationships between prognosis and VAF distributions. Moreover, we observed that samples with a larger amount of mutations were not necessarily linked to worse prognosis. By evaluating the ITH from multidimensional viewpoints, it will be possible to provide a more accurate prediction of cancer prognosis.. |
| 33. |
Chie Nakashima, Minako Yoshihara, Tetsuya Sato, Daisuke Saito, Mikita Suyama, Intratumor heterogeneity of HMCN1 mutant alleles associated with poor prognosis in patients with breast cancer, Oncotarget, 10.18632/oncotarget.26071, 9, 70, 33337-33347, 2018.09, Human breast cancers comprise a complex and highly heterogeneous population of tumor cells. Intratumor heterogeneity is an underlying cause of resistance to effective therapies and disease recurrence. To explore prognostic factors based on intratumor heterogeneity, we analyzed genomic mutations in breast cancer patients registered in The Cancer Genome Atlas. We calculated the variant allele frequency (VAF) at each mutation site and evaluated the associations of VAFs with the prognosis of breast cancer. VAFs of HMCN1 correlated with the prognosis and lymph node status. Although the detailed function of HMCN1 remains unknown, it is located in extracellular matrix and the mutation in the gene might be associated with cancer cell invasion and metastasis. This finding suggests that HMCN1 is a potential metastatic factor and can be a candidate gene for targeted breast cancer therapy.. |
| 34. |
Yasuyuki Kita, Yuta Katayama, Taichi Shiraishi, Takeru Oka, Tetsuya Sato, Mikita Suyama, Yasuyuki Ohkawa, Keishi Miyata, Yuichi Oike, Michiko Shirane, Masaaki Nishiyama, Keiichi I Nakayama, The Autism-Related Protein CHD8 Cooperates with C/EBPβ to Regulate Adipogenesis., Cell reports, 10.1016/j.celrep.2018.04.050, 23, 7, 1988-2000, 2018.05, The gene encoding the chromatin remodeler CHD8 is the most frequently mutated gene in individuals with autism spectrum disorder (ASD). Heterozygous mutations in CHD8 give rise to ASD that is often accompanied by macrocephaly, gastrointestinal complaints, and slender habitus. Whereas most phenotypes of CHD8 haploinsufficiency likely result from delayed neurodevelopment, the mechanism underlying slender habitus has remained unknown. Here, we show that CHD8 interacts with CCAAT/enhancer-binding protein β (C/EBPβ) and promotes its transactivation activity during adipocyte differentiation. Adipogenesis was impaired in Chd8-deleted preadipocytes, with the upregulation of C/EBPα and peroxisome-proliferator-activated receptor γ (PPARγ), two master regulators of this process, being attenuated in mutant cells. Furthermore, mice with CHD8 ablation in white preadipocytes had a markedly reduced white adipose tissue mass. Our findings reveal a mode of C/EBPβ regulation by CHD8 during adipogenesis, with CHD8 deficiency resulting in a defect in the development of white adipose tissue.. |
| 35. |
Ayako Suzuki, Shin Kawano, Toutai Mitsuyama, Mikita Suyama, Yae Kanai, Katsuhiko Shirahige, Hiroyuki Sasaki, Katsushi Tokunaga, Katsuya Tsuchihara, Sumio Sugano, Kenta Nakai, Yutaka Suzuki, DBTSS/DBKERO for integrated analysis of transcriptional regulation, Nucleic Acids Research, 10.1093/nar/gkx1001, 46, D1, D229-D238, 2018.01, DBTSS (Database of Transcriptional Start Sites)/DBKERO (Database of Kashiwa Encyclopedia for human genome mutations in Regulatory regions and their Omics contexts) is the database originally initiated with the information of transcriptional start sites and their upstream transcriptional regulatory regions. In recent years, we updated the database to assist users to elucidate biological relevance of the human genome variations or somatic mutations in cancers which may affect the transcriptional regulation. In this update, we facilitate interpretations of disease associated genomic variation, using the Japanese population as a model case. We enriched the genomic variation dataset consisting of the 13,368 individuals collected for various genome-wide association studies and the reference epigenome information in the surrounding regions using a total of 455 epigenome datasets (four tissue types from 67 healthy individuals) collected for the International Human Epigenome Consortium (IHEC). The data directly obtained from the clinical samples was associated with that obtained from various model systems, such as the drug perturbation datasets using cultured cancer cells. Furthermore, we incorporated the results obtained using the newly developed analytical methods, Nanopore/10x Genomics long-read sequencing of the human genome and single cell analyses. The database is made publicly accessible at the URL (http://dbtss.hgc.jp/).. |
| 36. |
Naoto Kubota, Toshifumi Yokoyama, Nobuhiko Hoshi, Mikita Suyama, Identification of a candidate enhancer for DMRT3 involved in spastic cerebral palsy pathogenesis, Biochemical and Biophysical Research Communications, 10.1016/j.bbrc.2018.01.011, 496, 1, 133-139, 2018.01, Cerebral palsy (CP) is a major neuronal disease and the most common movement disorder in children. Although environmental factors leading to CP have been greatly investigated, the genetic mechanism underlying CP is not well understood. Here we focused on two clinical reports that characterized a deletion involving the KANK1 gene locus in the 9p24.3 region. One report shows spastic CP and the other shows no spastic CP phenotype. Based on the epigenetic status and evolutionary conservation, we first found a functional genomic element at the noncoding region that was deleted only in patients with spastic CP. This element contains the retinoic acid receptor/retinoid X receptor (RAR/RXR) complex-binding motif that is widely conserved among placental mammals. RAR/RXR ChIP-seq data from mouse F9 embryonal carcinoma cells that were treated with trans-retinoic acids showed that the element has a binding ability. In addition, data regarding chromosome conformation capture from mouse neural progenitor and ES cells suggested that the element spatially interacts with the Doublesex and mab-3 related transcription factor 3 (Dmrt3) gene promoter that is located approximately 120 kb downstream of the RAR/RXR-binding site. Dmrt3 is detected in the developing mouse forebrain and in some interneurons in the spinal cord, and it works as a locomotion coordinator in horses and mice. Thus, the deletion of the cis-regulatory element for DMRT3 in humans may cause impaired development of the forebrain and gait abnormalities, resulting in spastic CP. In conclusion, this study provides new mechanistic insights into the genetic basis of CP.. |
| 37. |
Ayako Suzuki, Shin Kawano, Toutai Mitsuyama, Mikita Suyama, Yae Kanai, Katsuhiko Shirahige, Hiroyuki Sasaki, Katsushi Tokunaga, Katsuya Tsuchihara, Sumio Sugano, Kenta Nakai, Yutaka Suzuki, DBTSS/DBKERO for integrated analysis of transcriptional regulation., Nucleic acids research, 10.1093/nar/gkx1001, 46, D1, D229-D238, 2018.01, DBTSS (Database of Transcriptional Start Sites)/DBKERO (Database of Kashiwa Encyclopedia for human genome mutations in Regulatory regions and their Omics contexts) is the database originally initiated with the information of transcriptional start sites and their upstream transcriptional regulatory regions. In recent years, we updated the database to assist users to elucidate biological relevance of the human genome variations or somatic mutations in cancers which may affect the transcriptional regulation. In this update, we facilitate interpretations of disease associated genomic variation, using the Japanese population as a model case. We enriched the genomic variation dataset consisting of the 13,368 individuals collected for various genome-wide association studies and the reference epigenome information in the surrounding regions using a total of 455 epigenome datasets (four tissue types from 67 healthy individuals) collected for the International Human Epigenome Consortium (IHEC). The data directly obtained from the clinical samples was associated with that obtained from various model systems, such as the drug perturbation datasets using cultured cancer cells. Furthermore, we incorporated the results obtained using the newly developed analytical methods, Nanopore/10x Genomics long-read sequencing of the human genome and single cell analyses. The database is made publicly accessible at the URL (http://dbtss.hgc.jp/).. |
| 38. |
Hiroaki Okae, Hidehiro Toh, Tetsuya Sato, Hitoshi Hiura, Sota Takahashi, Kenjiro Shirane, Yuka Kabayama, Mikita Suyama, Hiroyuki Sasaki, Takahiro Arima, Derivation of Human Trophoblast Stem Cells., Cell stem cell, 10.1016/j.stem.2017.11.004, 22, 1, 50-63, 2018.01, Trophoblast cells play an essential role in the interactions between the fetus and mother. Mouse trophoblast stem (TS) cells have been derived and used as the best in vitro model for molecular and functional analysis of mouse trophoblast lineages, but attempts to derive human TS cells have so far been unsuccessful. Here we show that activation of Wingless/Integrated (Wnt) and EGF and inhibition of TGF-β, histone deacetylase (HDAC), and Rho-associated protein kinase (ROCK) enable long-term culture of human villous cytotrophoblast (CT) cells. The resulting cell lines have the capacity to give rise to the three major trophoblast lineages, which show transcriptomes similar to those of the corresponding primary trophoblast cells. Importantly, equivalent cell lines can be derived from human blastocysts. Our data strongly suggest that the CT- and blastocyst-derived cell lines are human TS cells, which will provide a powerful tool to study human trophoblast development and function.. |
| 39. |
Naoto Kubota, Toshifumi Yokoyama, Nobuhiko Hoshi, Mikita Suyama, Identification of a candidate enhancer for DMRT3 involved in spastic cerebral palsy pathogenesis., Biochemical and biophysical research communications, 10.1016/j.bbrc.2018.01.011, 496, 1, 133-139, 2018.01, Cerebral palsy (CP) is a major neuronal disease and the most common movement disorder in children. Although environmental factors leading to CP have been greatly investigated, the genetic mechanism underlying CP is not well understood. Here we focused on two clinical reports that characterized a deletion involving the KANK1 gene locus in the 9p24.3 region. One report shows spastic CP and the other shows no spastic CP phenotype. Based on the epigenetic status and evolutionary conservation, we first found a functional genomic element at the noncoding region that was deleted only in patients with spastic CP. This element contains the retinoic acid receptor/retinoid X receptor (RAR/RXR) complex-binding motif that is widely conserved among placental mammals. RAR/RXR ChIP-seq data from mouse F9 embryonal carcinoma cells that were treated with trans-retinoic acids showed that the element has a binding ability. In addition, data regarding chromosome conformation capture from mouse neural progenitor and ES cells suggested that the element spatially interacts with the Doublesex and mab-3 related transcription factor 3 (Dmrt3) gene promoter that is located approximately 120 kb downstream of the RAR/RXR-binding site. Dmrt3 is detected in the developing mouse forebrain and in some interneurons in the spinal cord, and it works as a locomotion coordinator in horses and mice. Thus, the deletion of the cis-regulatory element for DMRT3 in humans may cause impaired development of the forebrain and gait abnormalities, resulting in spastic CP. In conclusion, this study provides new mechanistic insights into the genetic basis of CP.. |
| 40. |
Kouhei Shimaji, Ryo Tanaka, Toru Maeda, Mamiko Ozaki, Hideki Yoshida, Yasuyuki Ohkawa, Tetsuya Sato, Mikita Suyama, Masamitsu Yamaguchi, Histone methyltransferase G9a is a key regulator of the starvation-induced behaviors in Drosophila melanogaster, Scientific Reports, 10.1038/s41598-017-15344-2, 7, 1, 2017.12, Organisms have developed behavioral strategies to defend themselves from starvation stress. Despite of their importance in nature, the underlying mechanisms have been poorly understood. Here, we show that Drosophila G9a (dG9a), one of the histone H3 Lys 9-specific histone methyltransferases, functions as a key regulator for the starvation-induced behaviors. RNA-sequencing analyses utilizing dG9a null mutant flies revealed that the expression of some genes relating to gustatory perception are regulated by dG9a under starvation conditions. Reverse transcription quantitative-PCR analyses showed that the expression of gustatory receptor genes for sensing sugar are up-regulated in starved dG9a null mutant. Consistent with this, proboscis extension reflex tests indicated that dG9a depletion increased the sensitivity to sucrose under starvation conditions. Furthermore, the locomotion activity was promoted in starved dG9a null mutant. We also found that dG9a depletion down-regulates the expression of insulin-like peptide genes that are required for the suppression of starvation-induced hyperactivity. Furthermore, refeeding of wild type flies after starvation conditions restores the hyperactivity and increased sensitivity to sucrose as well as dG9a expression level. These data suggest that dG9a functions as a key regulator for the decision of behavioral strategies under starvation conditions.. |
| 41. |
Shintaro Matsuba, Toshiki Yabe-Wada, Kazuya Takeda, Tetsuya Sato, Mikita Suyama, Toshiyuki Takai, Toshiaki Kikuchi, Toshihiro Nukiwa, Akira Nakamura, Identification of secretory leukoprotease inhibitor as an endogenous negative regulator in allergic effector cells, Frontiers in Immunology, 10.3389/fimmu.2017.01538, 8, NOV, 2017.11, Mast cells, basophils, and eosinophils are central effectors in allergic inflammatory disorders. These cells secrete abundant serine proteases as well as chemical mediators and cytokines; however, the expression profiles and functions of their endogenous inhibitors remain elusive. We found that murine secretory leukoprotease inhibitor (SLPI) is expressed in basophils and eosinophils but in not in mast cells. SLPI-deficient (Slpi-/-) basophils produce more cytokines than wild-type mice after IgE stimulation. Although the deletion of SLPI in basophils did not affect the release of chemical mediators upon IgE stimulation, the enzymatic activity of the serine protease tryptase was increased in Slpi-/- basophils. Mice transferred with Slpi-/- basophils were highly sensitive to IgE-mediated chronic allergic inflammation. Eosinophils lacking SLPI showed greater interleukin-6 secretion and invasive activity upon lipopolysaccharide stimulation, and the expression of matrix metalloproteinase-9 by these eosinophils was increased without stimulation. The absence of SLPI increases JNK1 phosphorylation at the steady state, and augments the serine phosphorylation of JNK1-downstream ETS transcriptional factor Elk-1 in eosinophils upon stimulation. Of note, SLPI interacts with a scaffold protein, JNK-interacting protein 3 (JIP3), that constitutively binds to the cytoplasmic domain of toll-like receptor (TLR) 4, suggesting that SLPI controls Elk-1 activation via binding to JIP3 in eosinophils. Mice transferred with Slpi-/- eosinophils showed the exacerbation of chitin-induced allergic inflammation. These findings showed that SLPI is a negative regulator in allergic effector cells and suggested a novel inhibitory role of SLPI in the TLR4 signaling pathways.. |
| 42. |
Kouhei Shimaji, Ryo Tanaka, Toru Maeda, Mamiko Ozaki, Hideki Yoshida, Yasuyuki Ohkawa, Tetsuya Sato, Mikita Suyama, Masamitsu Yamaguchi, Histone methyltransferase G9a is a key regulator of the starvation-induced behaviors in Drosophila melanogaster., Scientific reports, 10.1038/s41598-017-15344-2, 7, 1, 14763-14763, 2017.11, Organisms have developed behavioral strategies to defend themselves from starvation stress. Despite of their importance in nature, the underlying mechanisms have been poorly understood. Here, we show that Drosophila G9a (dG9a), one of the histone H3 Lys 9-specific histone methyltransferases, functions as a key regulator for the starvation-induced behaviors. RNA-sequencing analyses utilizing dG9a null mutant flies revealed that the expression of some genes relating to gustatory perception are regulated by dG9a under starvation conditions. Reverse transcription quantitative-PCR analyses showed that the expression of gustatory receptor genes for sensing sugar are up-regulated in starved dG9a null mutant. Consistent with this, proboscis extension reflex tests indicated that dG9a depletion increased the sensitivity to sucrose under starvation conditions. Furthermore, the locomotion activity was promoted in starved dG9a null mutant. We also found that dG9a depletion down-regulates the expression of insulin-like peptide genes that are required for the suppression of starvation-induced hyperactivity. Furthermore, refeeding of wild type flies after starvation conditions restores the hyperactivity and increased sensitivity to sucrose as well as dG9a expression level. These data suggest that dG9a functions as a key regulator for the decision of behavioral strategies under starvation conditions.. |
| 43. |
Takashi Kuramoto, Birger Voigt, Satoshi Nakanishi, Kazuhiro Kitada, Tadashi Nakamura, Kaori Wakamatsu, Minako Yoshihara, Mikita Suyama, Risa Uemura, Miyuu Tanaka, Mitsuru Kuwamura, Saki Shimizu, Yukihiro Ohno, Masashi Sasa, Tadao Serikawa, Identification of Candidate Genes for Generalized Tonic-Clonic Seizures in Noda Epileptic Rat., Behavior genetics, 10.1007/s10519-017-9870-2, 47, 6, 609-619, 2017.11, The Noda epileptic rat (NER) exhibits generalized tonic-clonic seizures (GTCS). A genetic linkage analysis identified two GTCS-associated loci, Ner1 on Chr 1 and Ner3 on Chr 5. The wild-type Ner1 and Ner3 alleles suppressed GTCS when combined in double-locus congenic lines, but not when present in single-locus congenic lines. Global expression analysis revealed that cholecystokinin B receptor (Cckbr) and suppressor of tumorigenicity 5 (St5), which map within Ner1, and PHD finger protein 24 (Phf24), which maps within Ner3, were significantly downregulated in NER. De novo BAC sequencing detected an insertion of an endogenous retrovirus sequence in intron 2 of the Phf24 gene in the NER genome, and PHF24 protein was almost absent in the NER brain. Phf24 encodes a Gαi-interacting protein involved in GABAB receptor signaling pathway. Based on these findings, we conclude that Cckbr, St5, and Phf24 are strong candidate genes for GTCS in NER.. |
| 44. |
Kanako Miyabayashi, Yuichi Shima, Miki Inoue, Tetsuya Sato, Takashi Baba, Yasuyuki Ohkawa, Mikita Suyama, Ken-Ichirou Morohashi, Alterations in Fetal Leydig Cell Gene Expression during Fetal and Adult Development, Sexual Development, 10.1159/000453323, 11, 2, 53-63, 2017.05, Fetal Leydig cells (FLCs) and adult Leydig cells (ALCs) develop in the mammalian prenatal and postnatal testes, respectively. In mice, FLCs emerge in the interstitial space of the testis as early as embryonic day 12.5 and thereafter increase in number during the fetal stage. We previously established a transgenic mouse line in which FLCs are labeled with EGFP and demonstrated that the EGFP-labeled FLCs were present even in adult testes. However, the characteristics of FLCs during postnatal stages remained unclear. In the present study, a comparison of the transcriptomes of FLCs from prenatal and postnatal testes and of ALCs from adult testes revealed that FLCs gradually alter their characteristics across developmental stages and come to roughly resemble ALCs. Many cholesterogenic genes simultaneously expressed a unique alternation pattern, while many oxidative phosphorylation and β-oxidation (both mitochondrial functions) genes showed a different unique pattern. These metabolic gene expression alterations might be triggered by milieu changes, such as nutrient and oxygen supply, from the prenatal to the postnatal period.. |
| 45. |
Minako Yoshihara, Tetsuya Sato, Daisuke Saito, Osamu Ohara, Takashi Kuramoto, Mikita Suyama, A deletion in the intergenic region upstream of Ednrb causes head spot in the rat strain KFRS4/Kyo, BMC Genetics, 10.1186/s12863-017-0497-3, 18, 1, 2017.03, Background: Head spot is one of the phenotypes identified in the KFRS4/Kyo rat strain. Although previous linkage analysis suggested that Ednrb, which is frequently involved in coat color variations in various animals, could be the gene responsible for this phenotype, no mutations have been identified in its coding region. Results: To identify mutations causative of this phenotype in KFRS4/Kyo, we analyzed target capture sequencing data that we recently generated. Our target capture method has a unique feature, i.e., it covers not only exonic regions but also conserved non-coding sequences (CNSs) among vertebrates; therefore, it has the potential to detect regulatory mutations. We identified a deletion of approximately 50kb in length approximately 50kb upstream of Ednrb. A comparative analysis with the epigenomic data in the corresponding region in humans and mice showed that one of the CNSs might be an enhancer. Further comparison with Hi-C data, which provide information about chromosome conformation, indicated that the putative enhancer is spatially close to the promoter of Ednrb, suggesting that it acts as an enhancer of Ednrb. Conclusions: These in silico data analyses strongly suggest that the identified deletion in the intergenic region upstream of Ednrb, which might contain a melanocyte-specific enhancer, is the mutation causative of the head spot phenotype in the KFRS4/Kyo rat strain.. |
| 46. |
Minako Yoshihara, Tetsuya Sato, Daisuke Saito, Osamu Ohara, Takashi Kuramoto, Mikita Suyama, A deletion in the intergenic region upstream of Ednrb causes head spot in the rat strain KFRS4/Kyo., BMC genetics, 10.1186/s12863-017-0497-3, 18, 1, 29-29, 2017.03, BACKGROUND: Head spot is one of the phenotypes identified in the KFRS4/Kyo rat strain. Although previous linkage analysis suggested that Ednrb, which is frequently involved in coat color variations in various animals, could be the gene responsible for this phenotype, no mutations have been identified in its coding region. RESULTS: To identify mutations causative of this phenotype in KFRS4/Kyo, we analyzed target capture sequencing data that we recently generated. Our target capture method has a unique feature, i.e., it covers not only exonic regions but also conserved non-coding sequences (CNSs) among vertebrates; therefore, it has the potential to detect regulatory mutations. We identified a deletion of approximately 50 kb in length approximately 50 kb upstream of Ednrb. A comparative analysis with the epigenomic data in the corresponding region in humans and mice showed that one of the CNSs might be an enhancer. Further comparison with Hi-C data, which provide information about chromosome conformation, indicated that the putative enhancer is spatially close to the promoter of Ednrb, suggesting that it acts as an enhancer of Ednrb. CONCLUSIONS: These in silico data analyses strongly suggest that the identified deletion in the intergenic region upstream of Ednrb, which might contain a melanocyte-specific enhancer, is the mutation causative of the head spot phenotype in the KFRS4/Kyo rat strain.. |
| 47. |
Bing Li, Takashi Baba, Kanako Miyabayashi, Tetsuya Sato, Yuichi Shima, Tomomi Ichinose, Daisuke Miura, Yasuyuki Ohkawa, Mikita Suyama, Ken-Ichirou Morohashi, Role of Ad4-binding protein/steroidogenic factor 1 in regulating NADPH production in adrenocortical Y-1 cells., Endocrine journal, 10.1507/endocrj.EJ16-0467, 64, 3, 315-324, 2017.03, Ad4-binding protein/steroidogenic factor 1 (Ad4BP/SF-1), a member of the nuclear receptor superfamily, is expressed in steroidogenic cells and regulates all steroidogenic gene expression. We recently employed mRNA and chromatin immunoprecipitation sequence (ChIP-seq) to demonstrate that Ad4BP/SF-1 directly regulates the expression of nearly all glycolytic genes. The pentose phosphate pathway (PPP) contributes to the production of nicotinamide adenine dinucleotide phosphate (NADPH). Although the expression of PPP genes and intracellular NADPH were decreased by Ad4BP/SF-1 knockdown, these genes were not the direct targets of Ad4BP/SF-1. This study therefore investigates whether Ad4BP/SF-1 directly regulates genes implicated in NADPH production. Examination of previously published data sets of mRNA sequence (mRNA-seq) and ChIP-seq strongly suggested a possibility that other NADPH-producing genes, such as malic enzyme 1 (Me1) and methylenetetrahydrofolate dehydrogenase 2 (Mthfd2), are the direct targets of Ad4BP/SF-1. Reporter gene assays and determination of intracellular NADPH concentration supported the notion that Ad4BP/SF-1 regulates NADPH production by regulating these genes. NADPH is required for macromolecule synthesis of compounds such as steroids, and for detoxification of reactive oxygen species. When synthesizing steroid hormones, steroidogenic cells consume NADPH through enzymatic reactions mediated by steroidogenic P450s. NADPH is also consumed through elimination of reactive oxygen species produced as the byproducts of the P450 reactions. Overall, Ad4BP/SF-1 potentially maintains the intracellular NADPH level through cooperative regulation of genes involved in the biological processes for consumption and supply.. |
| 48. |
Yurina Shishido, Takashi Baba, Tetsuya Sato, Yuichi Shima, Kanako Miyabayashi, Miki Inoue, Haruhiko Akiyama, Hiroshi Kimura, Yoshiakira Kanai, Yasuhiro Ishihara, Shogo Haraguchi, Akira Miyazaki, Damjana Rozman, Takeshi Yamazaki, Man-Ho Choi, Yasuyuki Ohkawa, Mikita Suyama, Ken-Ichirou Morohashi, Differential lactate and cholesterol synthetic activities in XY and XX Sertoli cells., Scientific reports, 10.1038/srep41912, 7, 41912-41912, 2017.02, SRY, a sex-determining gene, induces testis development in chromosomally female (XX) individuals. However, mouse XX Sertoli cells carrying Sry (XX/Sry Sertoli cells) are incapable of fully supporting germ cell development, even when the karyotype of the germ cells is XY. While it has therefore been assumed that XX/Sry Sertoli cells are not functionally equivalent to XY Sertoli cells, it has remained unclear which specific functions are affected. To elucidate the functional difference, we compared the gene expression of XY and XX/Sry Sertoli cells. Lactate and cholesterol metabolisms, essential for nursing the developing germ cells, were down-regulated in XX/Sry cells, which appears to be caused at least in part by the differential expression of histone modification enzymes SMCX/SMCY (H3K4me3 demethylase) and UTX/UTY (H3K27me3 demethylase) encoded by the sex chromosomes. We suggest that down-regulation of lactate and cholesterol metabolism that may be due to altered epigenetic modification affects the nursing functions of XX/Sry Sertoli cells.. |
| 49. |
Takanari Umegawachi, Hideki Yoshida, Hiromu Koshida, Momoko Yamada, Yasuyuki Ohkawa, Tetsuya Sato, Mikita Suyama, Henry M. Krause, Masamitsu Yamaguchi, Control of tissue size and development by a regulatory element in the yorkie 3'UTR, American Journal of Cancer Research, 7, 3, 673-687, 2017.01, Regulation of the Hippo pathway via phosphorylation of Yorkie (Yki), the Drosophila homolog of human Yes-associated protein 1, is conserved from Drosophila to humans. Overexpression of a non-phosphorylatable form of Yki induces severe overgrowth in adult fly eyes. Here, we show that yki mRNA associates with microsomal fractions and forms foci that partially colocalize to processing bodies in the vicinity of endoplasmic reticulum. This localization is dependent on a stem-loop (SL) structure in the 3' untranslated region of yki. Surprisingly, expression of SL deleted yki in eye imaginal discs also results in severe overgrowth phenotypes. When the structure of the SL is disrupted, Yki protein levels increase without a significant effect on RNA levels. When the SL is completely removed, protein levels drastically increase, but in this case, due to increased RNA stability. In the latter case, we show that the increased RNA accumulation is due to removal of a putative miR-8 seed sequence in the SL. These data demonstrate the function of two novel regulatory mechanisms, both controlled by the yki SL element, that are essential for proper Hippo pathway mediated growth regulation.. |
| 50. |
Bing Li, Takashi Baba, Kanako Miyabayashi, Tetsuya Sato, Yuichi Shima, Tomomi Ichinose, Daisuke Miura, Yasuyuki Ohkawa, Mikita Suyama, Ken-Ichirou Morohashi, Role of Ad4-binding protein/steroidogenic factor 1 in regulating NADPH production in adrenocortical Y-1 cells, Endocrine Journal, 10.1507/endocrj.EJ16-0467, 64, 3, 315-324, 2017.01, Ad4-binding protein/steroidogenic factor 1 (Ad4BP/SF-1), a member of the nuclear receptor superfamily, is expressed in steroidogenic cells and regulates all steroidogenic gene expression. We recently employed mRNA and chromatin immunoprecipitation sequence (ChIP-seq) to demonstrate that Ad4BP/SF-1 directly regulates the expression of nearly all glycolytic genes. The pentose phosphate pathway (PPP) contributes to the production of nicotinamide adenine dinucleotide phosphate (NADPH). Although the expression of PPP genes and intracellular NADPH were decreased by Ad4BP/SF-1 knockdown, these genes were not the direct targets of Ad4BP/SF-1. This study therefore investigates whether Ad4BP/SF-1 directly regulates genes implicated in NADPH production. Examination of previously published data sets of mRNA sequence (mRNA-seq) and ChIP-seq strongly suggested a possibility that other NADPH-producing genes, such as malic enzyme 1 (Me1) and methylenetetrahydrofolate dehydrogenase 2 (Mthfd2), are the direct targets of Ad4BP/SF-1. Reporter gene assays and determination of intracellular NADPH concentration supported the notion that Ad4BP/SF-1 regulates NADPH production by regulating these genes. NADPH is required for macromolecule synthesis of compounds such as steroids, and for detoxification of reactive oxygen species. When synthesizing steroid hormones, steroidogenic cells consume NADPH through enzymatic reactions mediated by steroidogenic P450s. NADPH is also consumed through elimination of reactive oxygen species produced as the byproducts of the P450 reactions. Overall, Ad4BP/SF-1 potentially maintains the intracellular NADPH level through cooperative regulation of genes involved in the biological processes for consumption and supply.. |
| 51. |
Hidehiro Toh, Kenjiro Shirane, Fumihito Miura, Naoki Kubo, Kenji Ichiyanagi, Katsuhiko Hayashi, Mitinori Saitou, Mikita Suyama, Takashi Ito, Hiroyuki Sasaki, Software updates in the illumina hiseq platform affect whole-genome bisulfite sequencing, BMC Genomics, 10.1186/s12864-016-3392-9, 18, 1, 2017.01, Background: Methylation of cytosine in genomic DNA is a well-characterized epigenetic modification involved in many cellular processes and diseases. Whole-genome bisulfite sequencing (WGBS), such as MethylC-seq and post-bisulfite adaptor tagging sequencing (PBAT-seq), uses the power of high-throughput DNA sequencers and provides genome-wide DNA methylation profiles at single-base resolution. However, the accuracy and consistency of WGBS outputs in relation to the operating conditions of high-throughput sequencers have not been explored. Results: We have used the Illumina HiSeq platform for our PBAT-based WGBS, and found that different versions of HiSeq Control Software (HCS) and Real-Time Analysis (RTA) installed on the system provided different global CpG methylation levels (approximately 5% overall difference) for the same libraries. This problem was reproduced multiple times with different WGBS libraries and likely to be associated with the low sequence diversity of bisulfite-converted DNA. We found that HCS was the major determinant in the observed differences. To determine which version of HCS is most suitable for WGBS, we used substrates with predetermined CpG methylation levels, and found that HCS v2.0.5 is the best among the examined versions. HCS v2.0.12 showed the poorest performance and provided artificially lower CpG methylation levels when 5-methylcytosine is read as guanine (first read of PBAT-seq and second read of MethylC-seq). In addition, paired-end sequencing of low diversity libraries using HCS v2.2.38 or the latest HCS v2.2.58 was greatly affected by cluster densities. Conclusions: Software updates in the Illumina HiSeq platform can affect the outputs from low-diversity sequencing libraries such as WGBS libraries. More recent versions are not necessarily the better, and HCS v2.0.5 is currently the best for WGBS among the examined HCS versions. Thus, together with other experimental conditions, special care has to be taken on this point when CpG methylation levels are to be compared between different samples by WGBS.. |
| 52. |
Yurin Shishido, Takashi Baba, Tetsuy Sato, Yuich Shima, Kanak Miyabayashi, Mik Inoue, Haruhik Akiyama, Hirosh Kimura, Yoshiakir Kanai, Yasuhir Ishihara, Shogo Haraguchi, Akir Miyazaki, Damjan Rozman, Takesh Yamazaki, Man Ho Choi, Yasuyuki Ohkawa, Mikita Suyama, Ken-Ichirou Morohashi, Differential lactate and cholesterol synthetic activities in XY and XX Sertoli cells, Scientific reports, 10.1038/srep41912, 7, 2017.01, SRY, a sex-determining gene, induces testis development in chromosomally female (XX) individuals. However, mouse XX Sertoli cells carrying Sry (XX/Sry Sertoli cells) are incapable of fully supporting germ cell development, even when the karyotype of the germ cells is XY. While it has therefore been assumed that XX/Sry Sertoli cells are not functionally equivalent to XY Sertoli cells, it has remained unclear which specific functions are affected. To elucidate the functional difference, we compared the gene expression of XY and XX/Sry Sertoli cells. Lactate and cholesterol metabolisms, essential for nursing the developing germ cells, were down-regulated in XX/Sry cells, which appears to be caused at least in part by the differential expression of histone modification enzymes SMCX/SMCY (H3K4me3 demethylase) and UTX/UTY (H3K27me3 demethylase) encoded by the sex chromosomes. We suggest that downregulation of lactate and cholesterol metabolism that may be due to altered epigenetic modification affects the nursing functions of XX/Sry Sertoli cells.. |
| 53. |
Hidehiro Toh, Kenjiro Shirane, Fumihito Miura, Naoki Kubo, Kenji Ichiyanagi, Katsuhiko Hayashi, Mitinori Saitou, Mikita Suyama, Takashi Ito, Hiroyuki Sasaki, Software updates in the Illumina HiSeq platform affect whole-genome bisulfite sequencing., BMC genomics, 10.1186/s12864-016-3392-9, 18, 1, 31-31, 2017.01, BACKGROUND: Methylation of cytosine in genomic DNA is a well-characterized epigenetic modification involved in many cellular processes and diseases. Whole-genome bisulfite sequencing (WGBS), such as MethylC-seq and post-bisulfite adaptor tagging sequencing (PBAT-seq), uses the power of high-throughput DNA sequencers and provides genome-wide DNA methylation profiles at single-base resolution. However, the accuracy and consistency of WGBS outputs in relation to the operating conditions of high-throughput sequencers have not been explored. RESULTS: We have used the Illumina HiSeq platform for our PBAT-based WGBS, and found that different versions of HiSeq Control Software (HCS) and Real-Time Analysis (RTA) installed on the system provided different global CpG methylation levels (approximately 5% overall difference) for the same libraries. This problem was reproduced multiple times with different WGBS libraries and likely to be associated with the low sequence diversity of bisulfite-converted DNA. We found that HCS was the major determinant in the observed differences. To determine which version of HCS is most suitable for WGBS, we used substrates with predetermined CpG methylation levels, and found that HCS v2.0.5 is the best among the examined versions. HCS v2.0.12 showed the poorest performance and provided artificially lower CpG methylation levels when 5-methylcytosine is read as guanine (first read of PBAT-seq and second read of MethylC-seq). In addition, paired-end sequencing of low diversity libraries using HCS v2.2.38 or the latest HCS v2.2.58 was greatly affected by cluster densities. CONCLUSIONS: Software updates in the Illumina HiSeq platform can affect the outputs from low-diversity sequencing libraries such as WGBS libraries. More recent versions are not necessarily the better, and HCS v2.0.5 is currently the best for WGBS among the examined HCS versions. Thus, together with other experimental conditions, special care has to be taken on this point when CpG methylation levels are to be compared between different samples by WGBS.. |
| 54. |
Minako Yoshihara, Tetsuya Sato, Daisuke Saito, Osamu Ohara, Takashi Kuramoto, Mikita Suyama, Application of target capture sequencing of exons and conserved non-coding sequences to 20 inbred rat strains, Genomics Data, 10.1016/j.gdata.2016.11.010, 10, 155-157, 2016.12, We report sequence data obtained by our recently devised target capture method TargetEC applied to 20 inbred rat strains. This method encompasses not only all annotated exons but also highly conserved non-coding sequences shared among vertebrates. The total length of the target regions covers 146.8 Mb. On an average, we obtained 31.7 × depth of target coverage and identified 154,330 SNVs and 24,368 INDELs for each strain. This corresponds to 470,037 unique SNVs and 68,652 unique INDELs among the 20 strains. The sequence data can be accessed at DDBJ/EMBL/GenBank under accession number PRJDB4648, and the identified variants have been deposited at http://bioinfo.sls.kyushu-u.ac.jp/rat_target_capture/20_strains.vcf.gz.. |
| 55. |
Minako Yoshihara, Tetsuya Sato, Daisuke Saito, Osamu Ohara, Takashi Kuramoto, Mikita Suyama, Application of target capture sequencing of exons and conserved non-coding sequences to 20 inbred rat strains., Genomics data, 10, 155-157, 2016.12, We report sequence data obtained by our recently devised target capture method TargetEC applied to 20 inbred rat strains. This method encompasses not only all annotated exons but also highly conserved non-coding sequences shared among vertebrates. The total length of the target regions covers 146.8 Mb. On an average, we obtained 31.7 × depth of target coverage and identified 154,330 SNVs and 24,368 INDELs for each strain. This corresponds to 470,037 unique SNVs and 68,652 unique INDELs among the 20 strains. The sequence data can be accessed at DDBJ/EMBL/GenBank under accession number PRJDB4648, and the identified variants have been deposited at http://bioinfo.sls.kyushu-u.ac.jp/rat_target_capture/20_strains.vcf.gz.. |
| 56. |
Hirotaka Hamada, Hiroaki Okae, Hidehiro Toh, Hatsune Chiba, Hitoshi Hiura, Kenjiro Shirane, Tetsuya Sato, Mikita Suyama, Nobuo Yaegashi, Hiroyuki Sasaki, Takahiro Arima, Allele-Specific Methylome and Transcriptome Analysis Reveals Widespread Imprinting in the Human Placenta, American Journal of Human Genetics, 10.1016/j.ajhg.2016.08.021, 99, 5, 1045-1058, 2016.11, DNA methylation is globally reprogrammed after fertilization, and as a result, the parental genomes have similar DNA-methylation profiles after implantation except at the germline differentially methylated regions (gDMRs). We and others have previously shown that human blastocysts might contain thousands of transient maternally methylated gDMRs (transient mDMRs), whose maternal methylation is lost in embryonic tissues after implantation. In this study, we performed genome-wide allelic DNA methylation analyses of purified trophoblast cells from human placentas and, surprisingly, found that more than one-quarter of the transient-in-embryo mDMRs maintained their maternally biased DNA methylation. RNA-sequencing-based allelic expression analyses revealed that some of the placenta-specific mDMRs were associated with expression of imprinted genes (e.g., TIGAR, SLC4A7, PROSER2-AS1, and KLHDC10), and three imprinted gene clusters were identified. This approach also identified some X-linked gDMRs. Comparisons of the data with those from other mammals revealed that genomic imprinting in the placenta is highly variable. These findings highlight the incomplete erasure of germline DNA methylation in the human placenta; understanding this erasure is important for understanding normal placental development and the pathogenesis of developmental disorders with imprinting effects.. |
| 57. |
Hirotaka Hamada, Hiroaki Okae, Hidehiro Toh, Hatsune Chiba, Hitoshi Hiura, Kenjiro Shirane, Tetsuya Sato, Mikita Suyama, Nobuo Yaegashi, Hiroyuki Sasaki, Takahiro Arima, Allele-Specific Methylome and Transcriptome Analysis Reveals Widespread Imprinting in the Human Placenta., American journal of human genetics, 10.1016/j.ajhg.2016.08.021, 99, 5, 1045-1058, 2016.11, DNA methylation is globally reprogrammed after fertilization, and as a result, the parental genomes have similar DNA-methylation profiles after implantation except at the germline differentially methylated regions (gDMRs). We and others have previously shown that human blastocysts might contain thousands of transient maternally methylated gDMRs (transient mDMRs), whose maternal methylation is lost in embryonic tissues after implantation. In this study, we performed genome-wide allelic DNA methylation analyses of purified trophoblast cells from human placentas and, surprisingly, found that more than one-quarter of the transient-in-embryo mDMRs maintained their maternally biased DNA methylation. RNA-sequencing-based allelic expression analyses revealed that some of the placenta-specific mDMRs were associated with expression of imprinted genes (e.g., TIGAR, SLC4A7, PROSER2-AS1, and KLHDC10), and three imprinted gene clusters were identified. This approach also identified some X-linked gDMRs. Comparisons of the data with those from other mammals revealed that genomic imprinting in the placenta is highly variable. These findings highlight the incomplete erasure of germline DNA methylation in the human placenta; understanding this erasure is important for understanding normal placental development and the pathogenesis of developmental disorders with imprinting effects.. |
| 58. |
Yuta Katayama, Masaaki Nishiyama, Hirotaka Shoji, Yasuyuki Ohkawa, Atsuki Kawamura, Tetsuya Sato, Mikita Suyama, Toru Takumi, Tsuyoshi Miyakawa, Keiichi I Nakayama, CHD8 haploinsufficiency results in autistic-like phenotypes in mice., Nature, 10.1038/nature19357, 537, 7622, 675-679, 2016.09, Autism spectrum disorder (ASD) comprises a range of neurodevelopmental disorders characterized by deficits in social interaction and communication as well as by restricted and repetitive behaviours. ASD has a strong genetic component with high heritability. Exome sequencing analysis has recently identified many de novo mutations in a variety of genes in individuals with ASD, with CHD8, a gene encoding a chromatin remodeller, being most frequently affected. Whether CHD8 mutations are causative for ASD and how they might establish ASD traits have remained unknown. Here we show that mice heterozygous for Chd8 mutations manifest ASD-like behavioural characteristics including increased anxiety, repetitive behaviour, and altered social behaviour. CHD8 haploinsufficiency did not result in prominent changes in the expression of a few specific genes but instead gave rise to small but global changes in gene expression in the mouse brain, reminiscent of those in the brains of patients with ASD. Gene set enrichment analysis revealed that neurodevelopment was delayed in the mutant mouse embryos. Furthermore, reduced expression of CHD8 was associated with abnormal activation of RE-1 silencing transcription factor (REST), which suppresses the transcription of many neuronal genes. REST activation was also observed in the brains of humans with ASD, and CHD8 was found to interact physically with REST in the mouse brain. Our results are thus consistent with the notion that CHD8 haploinsufficiency is a highly penetrant risk factor for ASD, with disease pathogenesis probably resulting from a delay in neurodevelopment.. |
| 59. |
Minako Yoshihara, Daisuke Saito, Tetsuya Sato, Osamu Ohara, Takashi Kuramoto, Mikita Suyama, Design and application of a target capture sequencing of exons and conserved non-coding sequences for the rat, BMC Genomics, 10.1186/s12864-016-2975-9, 17, 1, 2016.08, Background: Target capture sequencing is an efficient approach to directly identify the causative mutations of genetic disorders. To apply this strategy to laboratory rats exhibiting various phenotypes, we developed a novel target capture probe set, TargetEC (target capture for exons and conserved non-coding sequences), which can identify mutations not only in exonic regions but also in conserved non-coding sequences and thus can detect regulatory mutations. Results: TargetEC covers 1,078,129 regions spanning 146.8 Mb of the genome. We applied TargetEC to four inbred rat strains (WTC/Kyo, WTC-swh/Kyo, PVG/Seac, and KFRS4/Kyo) maintained by the National BioResource Project for the Rat in Japan, and successfully identified mutations associated with these phenotypes, including one mutation detected in a conserved non-coding sequence. Conclusions: The method developed in this study can be used to efficiently identify regulatory mutations, which cannot be detected using conventional exome sequencing, and will help to deepen our understanding of the relationships between regulatory mutations and associated phenotypes.. |
| 60. |
Minako Yoshihara, Daisuke Saito, Tetsuya Sato, Osamu Ohara, Takashi Kuramoto, Mikita Suyama, Design and application of a target capture sequencing of exons and conserved non-coding sequences for the rat., BMC genomics, 10.1186/s12864-016-2975-9, 17, 593-593, 2016.08, BACKGROUND: Target capture sequencing is an efficient approach to directly identify the causative mutations of genetic disorders. To apply this strategy to laboratory rats exhibiting various phenotypes, we developed a novel target capture probe set, TargetEC (target capture for exons and conserved non-coding sequences), which can identify mutations not only in exonic regions but also in conserved non-coding sequences and thus can detect regulatory mutations. RESULTS: TargetEC covers 1,078,129 regions spanning 146.8 Mb of the genome. We applied TargetEC to four inbred rat strains (WTC/Kyo, WTC-swh/Kyo, PVG/Seac, and KFRS4/Kyo) maintained by the National BioResource Project for the Rat in Japan, and successfully identified mutations associated with these phenotypes, including one mutation detected in a conserved non-coding sequence. CONCLUSIONS: The method developed in this study can be used to efficiently identify regulatory mutations, which cannot be detected using conventional exome sequencing, and will help to deepen our understanding of the relationships between regulatory mutations and associated phenotypes.. |
| 61. |
Peter E. Thijssen, Yuya Ito, Giacomo Grillo, Jun Wang, Guillaume Velasco, Hirohisa Nitta, Motoko Unoki, Minako Yoshihara, Mikita Suyama, Yu Sun, Richard J.L.F. Lemmers, Jessica C. De Greef, Andrew Gennery, Paolo Picco, Barbara Kloeckener-Gruissem, Tayfun Güngör, Ismail Reisli, Capucine Picard, Kamila Kebaili, Bertrand Roquelaure, Tsuyako Iwai, Ikuko Kondo, Takeo Kubota, Monique M. Van Ostaijen-Ten Dam, Maarten J.D. Van Tol, Corry Weemaes, Claire Francastel, Silvère M. Van Der Maarel, Hiroyuki Sasaki, Erratum
Mutations in CDCA7 and HELLS cause immunodeficiency-centromeric instability-facial anomalies syndrome (Nature Communications (2015) 6 (7870) DOI: 10.1038/ncomms8870), Nature Communications, 10.1038/ncomms12003, 7, 2016.06. |
| 62. |
Peter E Thijssen, Yuya Ito, Giacomo Grillo, Jun Wang, Guillaume Velasco, Hirohisa Nitta, Motoko Unoki, Minako Yoshihara, Mikita Suyama, Yu Sun, Richard J L F Lemmers, Jessica C de Greef, Andrew Gennery, Paolo Picco, Barbara Kloeckener-Gruissem, Tayfun Güngör, Ismail Reisli, Capucine Picard, Kamila Kebaili, Bertrand Roquelaure, Tsuyako Iwai, Ikuko Kondo, Takeo Kubota, Monique M van Ostaijen-Ten Dam, Maarten J D van Tol, Corry Weemaes, Claire Francastel, Silvère M van der Maarel, Hiroyuki Sasaki, Corrigendum: Mutations in CDCA7 and HELLS cause immunodeficiency-centromeric instability-facial anomalies syndrome., Nature communications, 10.1038/ncomms12003, 7, 12003-12003, 2016.06. |
| 63. |
Toshiki Yabe-Wada, Shintaro Matsuba, Kazuya Takeda, Tetsuya Sato, Mikita Suyama, Yasuyuki Ohkawa, Toshiyuki Takai, Haifeng Shi, Caroline C. Philpott, Akira Nakamura, TLR signals posttranscriptionally regulate the cytokine trafficking mediator sortilin, Scientific Reports, 10.1038/srep26566, 6, 2016.05, Regulating the transcription, translation and secretion of cytokines is crucial for controlling the appropriate balance of inflammation. Here we report that the sorting receptor sortilin plays a key role in cytokine production. We observed interactions of sortilin with multiple cytokines including IFN-α, and sortilin depletion in plasmacytoid dendritic cells (pDCs) led to a reduction of IFN-α secretion, suggesting a pivotal role of sortilin in the exocytic trafficking of IFN-α in pDCs. Moreover, sortilin mRNA was degraded posttranscriptionally upon stimulation with various TLR ligands. Poly-rC-binding protein 1 (PCBP1) recognized the C-rich element (CRE) in the 3' UTR of sortilin mRNA, and depletion of PCBP1 enhanced the degradation of sortilin transcripts, suggesting that PCBP1 can act as a trans-acting factor to stabilize sortilin transcripts. The nucleotide-binding ability of PCBP1 was impaired by zinc ions and alterations of intracellular zinc affect sortilin expression. PCBP1 may therefore control the stability of sortilin transcripts by sensing intracellular zinc levels. Collectively, our findings provide insights into the posttranslational regulation of cytokine production through the posttranscriptional control of sortilin expression by TLR signals.. |
| 64. |
Toshiki Yabe-Wada, Shintaro Matsuba, Kazuya Takeda, Tetsuya Sato, Mikita Suyama, Yasuyuki Ohkawa, Toshiyuki Takai, Haifeng Shi, Caroline C Philpott, Akira Nakamura, TLR signals posttranscriptionally regulate the cytokine trafficking mediator sortilin., Scientific reports, 10.1038/srep26566, 6, 26566-26566, 2016.05, Regulating the transcription, translation and secretion of cytokines is crucial for controlling the appropriate balance of inflammation. Here we report that the sorting receptor sortilin plays a key role in cytokine production. We observed interactions of sortilin with multiple cytokines including IFN-α, and sortilin depletion in plasmacytoid dendritic cells (pDCs) led to a reduction of IFN-α secretion, suggesting a pivotal role of sortilin in the exocytic trafficking of IFN-α in pDCs. Moreover, sortilin mRNA was degraded posttranscriptionally upon stimulation with various TLR ligands. Poly-rC-binding protein 1 (PCBP1) recognized the C-rich element (CRE) in the 3' UTR of sortilin mRNA, and depletion of PCBP1 enhanced the degradation of sortilin transcripts, suggesting that PCBP1 can act as a trans-acting factor to stabilize sortilin transcripts. The nucleotide-binding ability of PCBP1 was impaired by zinc ions and alterations of intracellular zinc affect sortilin expression. PCBP1 may therefore control the stability of sortilin transcripts by sensing intracellular zinc levels. Collectively, our findings provide insights into the posttranslational regulation of cytokine production through the posttranscriptional control of sortilin expression by TLR signals.. |
| 65. |
Miki Inoue, Yuichi Shima, Kanako Miyabayashi, Kaori Tokunaga, Tetsuya Sato, Takashi Baba, Yasuyuki Ohkawa, Haruhiko Akiyama, Mikita Suyama, Ken-Ichirou Morohashi, Isolation and characterization of Fetal Leydig progenitor cells of male mice, Endocrinology, 10.1210/en.2015-1773, 157, 3, 1222-1233, 2016.03, Fetal and adult Leydig cells develop in mammalian prenatal and postnatal testes, respectively. In mice, fetal Leydig cells (FLCs) emergeintheinterstitial spaceofthe testisatembryonic day 12.5 and thereafter increase in number, possibly through differentiation from progenitor cells. However, the progenitor cells have not yet been identified. Previously, we established transgenic mice in which FLCs are labeled strongly with enhanced green fluorescent protein (EGFP). Interestingly, fluorescence-activated cell sorting provided us with weakly EGFP-labeled cells as well as strongly EGFP-labeled FLCs. In vitro reconstruction of fetal testes demonstrated that weakly EGFP-labeled cells contain FLC progenitors. Transcriptome from the 2 cell populations revealed, as expected, marked differences in the expression of genes required for growth factor/receptor signaling and steroidogenesis. In addition, genes for energy metabolisms such as glycolytic pathways and the citrate cycle were activated in strongly EGFP-labeled cells, suggesting that metabolism is activated during FLC differentiation.. |
| 66. |
Miki Inoue, Yuichi Shima, Kanako Miyabayashi, Kaori Tokunaga, Tetsuya Sato, Takashi Baba, Yasuyuki Ohkawa, Haruhiko Akiyama, Mikita Suyama, Ken-ichirou Morohashi, Isolation and Characterization of Fetal Leydig Progenitor Cells of Male Mice., Endocrinology, 10.1210/en.2015-1773, 157, 3, 1222-33, 2016.03, Fetal and adult Leydig cells develop in mammalian prenatal and postnatal testes, respectively. In mice, fetal Leydig cells (FLCs) emerge in the interstitial space of the testis at embryonic day 12.5 and thereafter increase in number, possibly through differentiation from progenitor cells. However, the progenitor cells have not yet been identified. Previously, we established transgenic mice in which FLCs are labeled strongly with enhanced green fluorescent protein (EGFP). Interestingly, fluorescence-activated cell sorting provided us with weakly EGFP-labeled cells as well as strongly EGFP-labeled FLCs. In vitro reconstruction of fetal testes demonstrated that weakly EGFP-labeled cells contain FLC progenitors. Transcriptome from the 2 cell populations revealed, as expected, marked differences in the expression of genes required for growth factor/receptor signaling and steroidogenesis. In addition, genes for energy metabolisms such as glycolytic pathways and the citrate cycle were activated in strongly EGFP-labeled cells, suggesting that metabolism is activated during FLC differentiation.. |
| 67. |
Shoko Matsushita, Kentaro Suzuki, Yukiko Ogino, Shinjiro Hino, Tetsuya Sato, Mikita Suyama, Takahiro Matsumoto, Akiko Omori, Satoshi Inoue, Gen Yamada, Androgen regulates Mafb expression through its 3′UTR during mouse urethral masculinization, Endocrinology, 10.1210/en.2015-1586, 157, 2, 844-857, 2016.02, External genitalia are prominent organs showing hormone-dependent sexual differentiation. Androgen is an essential regulator of masculinization of the genital tubercle, which is the anlage of external genitalia. We have previously shown that v-maf avian musculoaponeurotic fibrosarcoma oncogene homolog B (MAFB) is an androgen-inducible regulator of embryonic urethral masculinization in mice. However, it remains unclear how androgen regulates Mafb expression. The current study suggests that the Mafb 3′ untranslated region (UTR) is an essential region for its regulation by androgen. We identified 2 functional androgen response elements (AREs) in Mafb 3′UTR. Androgen receptor is bound to such AREs in 3′UTR during urethral masculinization. In addition to 3′UTR, Mafb 5′UTR also showed androgen responsiveness. Moreover, we also demonstrated that β-catenin, one of genital tubercle masculinization factors, may be an additional regulator of Mafb expression during urethral masculinization. This study provides insights to elucidate mechanisms of gene regulation through AREs present in Mafb 3′UTR for a better understanding of the processes of urethral masculinization.. |
| 68. |
Shoko Matsushita, Kentaro Suzuki, Yukiko Ogino, Shinjiro Hino, Tetsuya Sato, Mikita Suyama, Takahiro Matsumoto, Akiko Omori, Satoshi Inoue, Gen Yamada, Androgen Regulates Mafb Expression Through its 3'UTR During Mouse Urethral Masculinization., Endocrinology, 10.1210/en.2015-1586, 157, 2, 844-57, 2016.02, External genitalia are prominent organs showing hormone-dependent sexual differentiation. Androgen is an essential regulator of masculinization of the genital tubercle, which is the anlage of external genitalia. We have previously shown that v-maf avian musculoaponeurotic fibrosarcoma oncogene homolog B (MAFB) is an androgen-inducible regulator of embryonic urethral masculinization in mice. However, it remains unclear how androgen regulates Mafb expression. The current study suggests that the Mafb 3' untranslated region (UTR) is an essential region for its regulation by androgen. We identified 2 functional androgen response elements (AREs) in Mafb 3'UTR. Androgen receptor is bound to such AREs in 3'UTR during urethral masculinization. In addition to 3'UTR, Mafb 5'UTR also showed androgen responsiveness. Moreover, we also demonstrated that β-catenin, one of genital tubercle masculinization factors, may be an additional regulator of Mafb expression during urethral masculinization. This study provides insights to elucidate mechanisms of gene regulation through AREs present in Mafb 3'UTR for a better understanding of the processes of urethral masculinization.. |
| 69. |
Yuta Katayama, Masaaki Nishiyama, Hirotaka Shoji, Yasuyuki Ohkawa, Atsuki Kawamura, Tetsuya Sato, Mikita Suyama, Toru Takumi, Tsuyoshi Miyakawa, Keiichi Nakayama, CHD8 haploinsufficiency results in autistic-like phenotypes in mice, Nature, 10.1038/nature19357, 537, 7622, 675-679, 2016.01, Autism spectrum disorder (ASD) comprises a range of neurodevelopmental disorders characterized by deficits in social interaction and communication as well as by restricted and repetitive behaviours. ASD has a strong genetic component with high heritability. Exome sequencing analysis has recently identified many de novo mutations in a variety of genes in individuals with ASD, with CHD8, a gene encoding a chromatin remodeller, being most frequently affected. Whether CHD8 mutations are causative for ASD and how they might establish ASD traits have remained unknown. Here we show that mice heterozygous for Chd8 mutations manifest ASD-like behavioural characteristics including increased anxiety, repetitive behaviour, and altered social behaviour. CHD8 haploinsufficiency did not result in prominent changes in the expression of a few specific genes but instead gave rise to small but global changes in gene expression in the mouse brain, reminiscent of those in the brains of patients with ASD. Gene set enrichment analysis revealed that neurodevelopment was delayed in the mutant mouse embryos. Furthermore, reduced expression of CHD8 was associated with abnormal activation of RE-1 silencing transcription factor (REST), which suppresses the transcription of many neuronal genes. REST activation was also observed in the brains of humans with ASD, and CHD8 was found to interact physically with REST in the mouse brain. Our results are thus consistent with the notion that CHD8 haploinsufficiency is a highly penetrant risk factor for ASD, with disease pathogenesis probably resulting from a delay in neurodevelopment.. |
| 70. |
Tetsuya Sato, Mikita Suyama, ChromContact
A web tool for analyzing spatial contact of chromosomes from Hi-C data, BMC Genomics, 10.1186/s12864-015-2282-x, 16, 1, 2015.12, Background: Hi-C analysis has revealed the three-dimensional architecture of chromosomes in the nucleus. Although Hi-C data contains valuable information on long-range interactions of chromosomes, the data is not yet widely utilized by molecular biologists because of the quantity of data. Results: We developed a web tool, ChromContact, to utilize the information obtained by Hi-C. The web tool is designed to be simple and easy to use. By specifying a locus of interest, ChromContact calculates contact profiles and generates links to the UCSC Genome Browser, enabling users to visually examine the contact information with various annotations. Conclusion: ChromContact provides wide-range of molecular biologists with a user-friendly means to access high-resolution Hi-C data. One of the possible applications of ChromContact is investigating novel long-range promoter-enhancer interactions. This facilitates the functional interpretation of statistically significant markers identified by GWAS or ChIP-seq peaks that are located far from any annotated genes. ChromContact is freely accessible at http://bioinfo.sls.kyushu-u.ac.jp/chromcontact/.. |
| 71. |
Daisuke Saito, Mikita Suyama, Linkage disequilibrium analysis of allelic heterogeneity in DNA methylation, Epigenetics, 10.1080/15592294.2015.1115176, 10, 12, 1093-1098, 2015.12, Heterogeneity of DNA methylation status among alleles is observed in various cell types and is involved in epigenetic gene regulation and cancer biology. However, the individual methylation profile within each allele has not yet been examined at the whole-genome level. In the present study, we applied linkage disequilibrium analysis to the DNA methylation data obtained from whole-genome bisulfite sequencing studies in mouse germline and other types of cells. We found that the methylation status of 2 consecutive CpG sites showed deviation from equilibrium frequency toward concordant linkage (both methylated or both unmethylated) in germline cells. In the imprinting loci where methylation of constituent alleles is known, our analysis detected the deviation toward the concordant linkage as expected. In addition, we applied this analysis to the transitional zone between methylated and unmethylated regions and to the cells undergoing epigenetic reprogramming. In both cases, deviation to the concordant-linked alleles was conspicuous, indicating that the methylation pattern is not random but rather concordant within each allele. These results will provide the key to understanding the mechanism underlying allelic heterogeneity.. |
| 72. |
Tetsuya Sato, Mikita Suyama, ChromContact: A web tool for analyzing spatial contact of chromosomes from Hi-C data., BMC genomics, 10.1186/s12864-015-2282-x, 16, 1060-1060, 2015.12, BACKGROUND: Hi-C analysis has revealed the three-dimensional architecture of chromosomes in the nucleus. Although Hi-C data contains valuable information on long-range interactions of chromosomes, the data is not yet widely utilized by molecular biologists because of the quantity of data. RESULTS: We developed a web tool, ChromContact, to utilize the information obtained by Hi-C. The web tool is designed to be simple and easy to use. By specifying a locus of interest, ChromContact calculates contact profiles and generates links to the UCSC Genome Browser, enabling users to visually examine the contact information with various annotations. CONCLUSION: ChromContact provides wide-range of molecular biologists with a user-friendly means to access high-resolution Hi-C data. One of the possible applications of ChromContact is investigating novel long-range promoter-enhancer interactions. This facilitates the functional interpretation of statistically significant markers identified by GWAS or ChIP-seq peaks that are located far from any annotated genes. ChromContact is freely accessible at http://bioinfo.sls.kyushu-u.ac.jp/chromcontact/ .. |
| 73. |
Kouhei Shimaji, Takahiro Konishi, Shintaro Tanaka, Hideki Yoshida, Yasuko Kato, Yasuyuki Ohkawa, Tetsuya Sato, Mikita Suyama, Hiroshi Kimura, Masamitsu Yamaguchi, Genomewide identification of target genes of histone methyltransferase dG9a during Drosophila embryogenesis, Genes to Cells, 10.1111/gtc.12281, 20, 11, 902-914, 2015.11, Post-translational modification of the histone plays important roles in epigenetic regulation of various biological processes. Among the identified histone methyltransferases (HMTases), G9a is a histone H3 Lys 9 (H3K9)-specific example active in euchromatic regions. Drosophila G9a (dG9a) has been reported to feature H3K9 dimethylation activity in vivo. Here, we show that the time required for hatching of a homozygous dG9a null mutant and heteroallelic combination of dG9a null mutants is delayed, suggesting that dG9a is at least partially responsible for progression of embryogenesis. Immunocytochemical analyses of the wild-type and the dG9a null mutant flies indicated that dG9a localizes in cytoplasm up to nuclear division cycle 7 where it is likely responsible for di-methylation of nucleosome-free H3K9. From cycles 8-11, dG9a moves into the nucleus and is responsible for di-methylating H3K9 in nucleosomes. RNA-sequence analysis utilizing early wild-type and dG9a mutant embryos showed that dG9a down-regulates expression of genes responsible for embryogenesis. RNA fluorescent in situ hybridization analysis further showed temporal and spatial expression patterns of these mRNAs did not significantly change in the dG9a mutant. These results indicate that dG9a controls transcription levels of some zygotic genes without changing temporal and spatial expression patterns of the transcripts of these genes.. |
| 74. |
Toyoshi Yanagihara, Fumiyuki Sanematsu, Tetsuya Sato, Takehito Uruno, Xuefeng Duan, Takahiro Tomino, Yosuke Harada, Mayuki Watanabe, Yuqing Wang, Yoshihiko Tanaka, Yoichi Nakanishi, Mikita Suyama, Fukui Yoshinori, Intronic regulation of Aire expression by Jmjd6 for self-tolerance induction in the thymus, Nature communications, 10.1038/ncomms9820, 6, 2015.11, The thymus has spatially distinct microenvironments, the cortex and the medulla, where the developing T-cells are selected to mature or die through the interaction with thymic stromal cells. To establish the immunological self in the thymus, medullary thymic epithelial cells (mTECs) express diverse sets of tissue-specific self-antigens (TSAs). This ectopic expression of TSAs largely depends on the transcriptional regulator Aire, yet the mechanism controlling Aire expression itself remains unknown. Here, we show that Jmjd6, a dioxygenase that catalyses lysyl hydroxylation of splicing regulatory proteins, is critical for Aire expression. Although Jmjd6 deficiency does not affect abundance of Aire transcript, the intron 2 of Aire gene is not effectively spliced out in the absence of Jmjd6, resulting in marked reduction of mature Aire protein in mTECs and spontaneous development of multi-organ autoimmunity in mice. These results highlight the importance of intronic regulation in controlling Aire protein expression.. |
| 75. |
Kouhei Shimaji, Takahiro Konishi, Shintaro Tanaka, Hideki Yoshida, Yasuko Kato, Yasuyuki Ohkawa, Tetsuya Sato, Mikita Suyama, Hiroshi Kimura, Masamitsu Yamaguchi, Genomewide identification of target genes of histone methyltransferase dG9a during Drosophila embryogenesis., Genes to cells : devoted to molecular & cellular mechanisms, 10.1111/gtc.12281, 20, 11, 902-14, 2015.11, Post-translational modification of the histone plays important roles in epigenetic regulation of various biological processes. Among the identified histone methyltransferases (HMTases), G9a is a histone H3 Lys 9 (H3K9)-specific example active in euchromatic regions. Drosophila G9a (dG9a) has been reported to feature H3K9 dimethylation activity in vivo. Here, we show that the time required for hatching of a homozygous dG9a null mutant and heteroallelic combination of dG9a null mutants is delayed, suggesting that dG9a is at least partially responsible for progression of embryogenesis. Immunocytochemical analyses of the wild-type and the dG9a null mutant flies indicated that dG9a localizes in cytoplasm up to nuclear division cycle 7 where it is likely responsible for di-methylation of nucleosome-free H3K9. From cycles 8-11, dG9a moves into the nucleus and is responsible for di-methylating H3K9 in nucleosomes. RNA-sequence analysis utilizing early wild-type and dG9a mutant embryos showed that dG9a down-regulates expression of genes responsible for embryogenesis. RNA fluorescent in situ hybridization analysis further showed temporal and spatial expression patterns of these mRNAs did not significantly change in the dG9a mutant. These results indicate that dG9a controls transcription levels of some zygotic genes without changing temporal and spatial expression patterns of the transcripts of these genes.. |
| 76. |
Toyoshi Yanagihara, Fumiyuki Sanematsu, Tetsuya Sato, Takehito Uruno, Xuefeng Duan, Takahiro Tomino, Yosuke Harada, Mayuki Watanabe, Yuqing Wang, Yoshihiko Tanaka, Yoichi Nakanishi, Mikita Suyama, Fukui Yoshinori, Intronic regulation of Aire expression by Jmjd6 for self-tolerance induction in the thymus., Nature communications, 10.1038/ncomms9820, 6, 8820-8820, 2015.11, The thymus has spatially distinct microenvironments, the cortex and the medulla, where the developing T-cells are selected to mature or die through the interaction with thymic stromal cells. To establish the immunological self in the thymus, medullary thymic epithelial cells (mTECs) express diverse sets of tissue-specific self-antigens (TSAs). This ectopic expression of TSAs largely depends on the transcriptional regulator Aire, yet the mechanism controlling Aire expression itself remains unknown. Here, we show that Jmjd6, a dioxygenase that catalyses lysyl hydroxylation of splicing regulatory proteins, is critical for Aire expression. Although Jmjd6 deficiency does not affect abundance of Aire transcript, the intron 2 of Aire gene is not effectively spliced out in the absence of Jmjd6, resulting in marked reduction of mature Aire protein in mTECs and spontaneous development of multi-organ autoimmunity in mice. These results highlight the importance of intronic regulation in controlling Aire protein expression.. |
| 77. |
Naoki Kubo, Hidehiro Toh, Kenjiro Shirane, Takayuki Shirakawa, Hisato Kobayashi, Tetsuya Sato, Hidetoshi Sone, Yasuyuki Sato, Shin Ichi Tomizawa, Yoshinori Tsurusaki, Hiroki Shibata, Hirotomo Saitsu, Yutaka Suzuki, Naomichi Matsumoto, Mikita Suyama, Tomohiro Kono, Kazuyuki Ohbo, Hiroyuki Sasaki, DNA methylation and gene expression dynamics during spermatogonial stem cell differentiation in the early postnatal mouse testis, BMC genomics, 10.1186/s12864-015-1833-5, 16, 1, 2015.08, Background: In the male germline, neonatal prospermatogonia give rise to spermatogonia, which include stem cell population (undifferentiated spermatogonia) that supports continuous spermatogenesis in adults. Although the levels of DNA methyltransferases change dynamically in the neonatal and early postnatal male germ cells, detailed genome-wide DNA methylation profiles of these cells during the stem cell formation and differentiation have not been reported. Results: To understand the regulation of spermatogonial stem cell formation and differentiation, we examined the DNA methylation and gene expression dynamics of male mouse germ cells at the critical stages: neonatal prospermatogonia, and early postntal (day 7) undifferentiated and differentiating spermatogonia. We found large partially methylated domains similar to those found in cancer cells and placenta in all these germ cells, and high levels of non-CG methylation and 5-hydroxymethylcytosines in neonatal prospermatogonia. Although the global CG methylation levels were stable in early postnatal male germ cells, and despite the reported scarcity of differential methylation in the adult spermatogonial stem cells, we identified many regions showing stage-specific differential methylation in and around genes important for stem cell function and spermatogenesis. These regions contained binding sites for specific transcription factors including the SOX family members. Conclusions: Our findings show a distinctive and dynamic regulation of DNA methylation during spermatogonial stem cell formation and differentiation in the neonatal and early postnatal testes. Furthermore, we revealed a unique accumulation and distribution of non-CG methylation and 5hmC marks in neonatal prospermatogonia. These findings contrast with the reported scarcity of differential methylation in adult spermatogonial stem cell differentiation and represent a unique phase of male germ cell development.. |
| 78. |
Naoki Kubo, Hidehiro Toh, Kenjiro Shirane, Takayuki Shirakawa, Hisato Kobayashi, Tetsuya Sato, Hidetoshi Sone, Yasuyuki Sato, Shin-ichi Tomizawa, Yoshinori Tsurusaki, Hiroki Shibata, Hirotomo Saitsu, Yutaka Suzuki, Naomichi Matsumoto, Mikita Suyama, Tomohiro Kono, Kazuyuki Ohbo, Hiroyuki Sasaki, DNA methylation and gene expression dynamics during spermatogonial stem cell differentiation in the early postnatal mouse testis., BMC genomics, 10.1186/s12864-015-1833-5, 16, 1, 624-624, 2015.08, BACKGROUND: In the male germline, neonatal prospermatogonia give rise to spermatogonia, which include stem cell population (undifferentiated spermatogonia) that supports continuous spermatogenesis in adults. Although the levels of DNA methyltransferases change dynamically in the neonatal and early postnatal male germ cells, detailed genome-wide DNA methylation profiles of these cells during the stem cell formation and differentiation have not been reported. RESULTS: To understand the regulation of spermatogonial stem cell formation and differentiation, we examined the DNA methylation and gene expression dynamics of male mouse germ cells at the critical stages: neonatal prospermatogonia, and early postntal (day 7) undifferentiated and differentiating spermatogonia. We found large partially methylated domains similar to those found in cancer cells and placenta in all these germ cells, and high levels of non-CG methylation and 5-hydroxymethylcytosines in neonatal prospermatogonia. Although the global CG methylation levels were stable in early postnatal male germ cells, and despite the reported scarcity of differential methylation in the adult spermatogonial stem cells, we identified many regions showing stage-specific differential methylation in and around genes important for stem cell function and spermatogenesis. These regions contained binding sites for specific transcription factors including the SOX family members. CONCLUSIONS: Our findings show a distinctive and dynamic regulation of DNA methylation during spermatogonial stem cell formation and differentiation in the neonatal and early postnatal testes. Furthermore, we revealed a unique accumulation and distribution of non-CG methylation and 5hmC marks in neonatal prospermatogonia. These findings contrast with the reported scarcity of differential methylation in adult spermatogonial stem cell differentiation and represent a unique phase of male germ cell development.. |
| 79. |
Toshiya Nishimura, Tetsuya Sato, Yasuhiro Yamamoto, Ikuko Watakabe, Yasuyuki Ohkawa, Mikita Suyama, Satoru Kobayashi, Minoru Tanaka, Foxl3 is a germ cell-intrinsic factor involved in sperm-egg fate decision in medaka, Science, 10.1126/science.aaa2657, 349, 6245, 328-331, 2015.07, Sex determination is an essential step in the commitment of a germ cell to a sperm or egg. However, the intrinsic factors that determine the sexual fate of vertebrate germ cells are unknown. Here, we show that foxl3, which is expressed in germ cells but not somatic cells in the gonad, is involved in sperm-egg fate decision in medaka fish. Adult XX medaka with disrupted foxl3 developed functional sperm in the expanded germinal epithelium of a histologically functional ovary. In chimeric medaka, mutant germ cells initiated spermatogenesis in female wild-type gonad. These results indicate that a germ cell-intrinsic cue for the sperm-egg fate decision is present in medaka and that spermatogenesis can proceed in a female gonadal environment.. |
| 80. |
Peter E. Thijssen, Yuya Ito, Giacomo Grillo, Jun Wang, Guillaume Velasco, Hirohisa Nitta, Motoko Unoki, Minako Yoshihara, Mikita Suyama, Yu Sun, Richard J.L.F. Lemmers, Jessica C. De Greef, Andrew Gennery, Paolo Picco, Barbara Kloeckener-Gruissem, Tayfun Güngör, Ismail Reisli, Capucine Picard, Kamila Kebaili, Bertrand Roquelaure, Tsuyako Iwai, Ikuko Kondo, Takeo Kubota, Monique M. Van Ostaijen-Ten Dam, Maarten J.D. Van Tol, Corry Weemaes, Claire Francastel, Silvère M. Van Der Maarel, Hiroyuki Sasaki, Mutations in CDCA7 and HELLS cause immunodeficiency-centromeric instability-facial anomalies syndrome, Nature communications, 10.1038/ncomms8870, 6, 2015.07, The life-threatening Immunodeficiency, Centromeric Instability and Facial Anomalies (ICF) syndrome is a genetically heterogeneous autosomal recessive disorder. Twenty percent of patients cannot be explained by mutations in the known ICF genes DNA methyltransferase 3B or zinc-finger and BTB domain containing 24. Here we report mutations in the cell division cycle associated 7 and the helicase, lymphoid-specific genes in 10 unexplained ICF cases. Our data highlight the genetic heterogeneity of ICF syndrome; however, they provide evidence that all genes act in common or converging pathways leading to the ICF phenotype.. |
| 81. |
Peter E Thijssen, Yuya Ito, Giacomo Grillo, Jun Wang, Guillaume Velasco, Hirohisa Nitta, Motoko Unoki, Minako Yoshihara, Mikita Suyama, Yu Sun, Richard J L F Lemmers, Jessica C de Greef, Andrew Gennery, Paolo Picco, Barbara Kloeckener-Gruissem, Tayfun Güngör, Ismail Reisli, Capucine Picard, Kamila Kebaili, Bertrand Roquelaure, Tsuyako Iwai, Ikuko Kondo, Takeo Kubota, Monique M van Ostaijen-Ten Dam, Maarten J D van Tol, Corry Weemaes, Claire Francastel, Silvère M van der Maarel, Hiroyuki Sasaki, Mutations in CDCA7 and HELLS cause immunodeficiency-centromeric instability-facial anomalies syndrome., Nature communications, 10.1038/ncomms8870, 6, 7870-7870, 2015.07, The life-threatening Immunodeficiency, Centromeric Instability and Facial Anomalies (ICF) syndrome is a genetically heterogeneous autosomal recessive disorder. Twenty percent of patients cannot be explained by mutations in the known ICF genes DNA methyltransferase 3B or zinc-finger and BTB domain containing 24. Here we report mutations in the cell division cycle associated 7 and the helicase, lymphoid-specific genes in 10 unexplained ICF cases. Our data highlight the genetic heterogeneity of ICF syndrome; however, they provide evidence that all genes act in common or converging pathways leading to the ICF phenotype.. |
| 82. |
Toshiya Nishimura, Tetsuya Sato, Yasuhiro Yamamoto, Ikuko Watakabe, Yasuyuki Ohkawa, Mikita Suyama, Satoru Kobayashi, Minoru Tanaka, Sex determination. foxl3 is a germ cell-intrinsic factor involved in sperm-egg fate decision in medaka., Science (New York, N.Y.), 10.1126/science.aaa2657, 349, 6245, 328-31, 2015.07, Sex determination is an essential step in the commitment of a germ cell to a sperm or egg. However, the intrinsic factors that determine the sexual fate of vertebrate germ cells are unknown. Here, we show that foxl3, which is expressed in germ cells but not somatic cells in the gonad, is involved in sperm-egg fate decision in medaka fish. Adult XX medaka with disrupted foxl3 developed functional sperm in the expanded germinal epithelium of a histologically functional ovary. In chimeric medaka, mutant germ cells initiated spermatogenesis in female wild-type gonad. These results indicate that a germ cell-intrinsic cue for the sperm-egg fate decision is present in medaka and that spermatogenesis can proceed in a female gonadal environment.. |
| 83. |
Tetsuya Sato, Mikita Suyama, GenomeCons
A web server for manipulating multiple genome sequence alignments and their consensus sequences, Bioinformatics, 10.1093/bioinformatics/btu803, 31, 8, 1293-1295, 2015.04, Summary: Genome sequence alignments provide valuable information on many aspects of molecular biological processes. In this study, we developed a web server, GenomeCons, for manipulating multiple genome sequence alignments and their consensus sequences for high-throughput genome sequence analyses. This server facilitates the visual inspection of multiple genome sequence alignments for a set of genomic intervals at a time. This allows the user to examine how these sites are evolutionarily conserved over time for their functional importance. The server also reports consensus sequences for the input genomic intervals, which can be applied to downstream analyses such as the identification of common motifs in the regions determined by ChIP-seq experiments. Availability and implementation: GenomeCons is freely accessible at http://bioinfo.sls.kyushu-u.ac.jp/genomecons/.. |
| 84. |
Hiroaki Okae, Hatsune Chiba, Hitoshi Hiura, Hirotaka Hamada, Akiko Sato, Takafumi Utsunomiya, Hiroyuki Kikuchi, Hiroaki Yoshida, Atsushi Tanaka, Mikita Suyama, Takahiro Arima, Genome-Wide Analysis of DNA Methylation Dynamics during Early Human Development, PLoS genetics, 10.1371/journal.pgen.1004868, 10, 12, 2014.12, DNA methylation is globally reprogrammed during mammalian preimplantation development, which is critical for normal development. Recent reduced representation bisulfite sequencing (RRBS) studies suggest that the methylome dynamics are essentially conserved between human and mouse early embryos. RRBS is known to cover 5–10% of all genomic CpGs, favoring those contained within CpG-rich regions. To obtain an unbiased and more complete representation of the methylome during early human development, we performed whole genome bisulfite sequencing of human gametes and blastocysts that covered>70% of all genomic CpGs. We found that the maternal genome was demethylated to a much lesser extent in human blastocysts than in mouse blastocysts, which could contribute to an increased number of imprinted differentially methylated regions in the human genome. Global demethylation of the paternal genome was confirmed, but SINE-VNTR-Alu elements and some other tandem repeat-containing regions were found to be specifically protected from this global demethylation. Furthermore, centromeric satellite repeats were hypermethylated in human oocytes but not in mouse oocytes, which might be explained by differential expression of de novo DNA methyltransferases. These data highlight both conserved and species-specific regulation of DNA methylation during early mammalian development. Our work provides further information critical for understanding the epigenetic processes underlying differentiation and pluripotency during early human development.. |
| 85. |
Minako Yoshihara, Li Jiang, Shinya Akatsuka, Mikita Suyama, Shinya Toyokuni, Genome-wide profiling of 8-oxoguanine reveals its association with spatial positioning in nucleus, DNA Research, 10.1093/dnares/dsu023, 21, 6, 603-612, 2014.12, 8-Oxoguanine (8-oxoG) is one of the most common DNA lesions generated by reactive oxygen species. In this study, we analysed the genome-wide distribution profile of 8-oxoG by combining immunoprecipitation by antibodies specific for the DNA fragments containing 8-oxoG with a microarray that covers rat genome. Genome-wide mapping of 8-oxoG in normal rat kidney revealed that 8-oxoG is preferentially located at gene deserts. We did not observe differences in 8-oxoG levels between groups of genes with high and low expression, possibly because of the generally low 8-oxoG levels in genic regions compared with gene deserts. The distribution of 8-oxoG and lamina-associated domains (LADs) were strongly correlated, suggesting that the spatial location of genomic DNA in the nucleus determines the susceptibility to oxidative modifications. One possible explanation for high 8-oxoG levels in LADs is that the nuclear periphery is more susceptible to the oxidative damage caused by the extra-nuclear factors. Moreover, LADs have a rather compact conformation, which may limit the recruitment of repair components to the modified bases.. |
| 86. |
Ladeana W. Hillier, Webb Miller, Ewan Birney, Wesley Warren, Ross C. Hardison, Chris P. Ponting, Peer Bork, David W. Burt, Martien A.M. Groenen, Mary E. Delany, Jerry B. Dodgson, Asif T. Chinwalla, Paul F. Cliften, Sandra W. Clifton, Kimberly D. Delehaunty, Catrina Fronick, Robert S. Fulton, Tina A. Graves, Colin Kremitzki, Dan Layman, Vincent Magrini, John D. McPherson, Tracie L. Miner, Patrick Minx, William E. Nash, Michael N. Nhan, Joanne O. Nelson, Lachlan G. Oddy, Craig S. Pohl, Jennifer Randall-Maher, Scott M. Smith, John W. Wallis, Shiaw Pyng Yang, Michael N. Romanov, Catherine M. Rondelli, Bob Paton, Jacqueline Smith, David Morrice, Laura Daniels, Helen G. Tempest, Lindsay Robertson, Julio S. Masabanda, Darren K. Griffin, Alain Vignal, Valerie Fillon, Lina Jacobbson, Susanne Kerje, Leif Andersson, Richard P.M. Crooijmans, Jan Aerts, Jan J. Van Der Poel, Hans Ellegren, Randolph B. Caldwell, Simon J. Hubbard, Darren V. Grafham, Andrzej M. Kierzek, Stuart R. McLaren, Ian M. Overton, Hiroshi Arakawa, Kevin J. Beattie, Yuri Bezzubov, Paul E. Boardman, James K. Bonfield, Michael D.R. Croning, Robert M. Davies, Matthew D. Francis, Sean J. Humphray, Carol E. Scott, Ruth G. Taylor, Cheryll Tickle, William R.A. Brown, Jane Rogers, Jean Marie Buerstedde, Stuart A. Wilson, Lisa Stubbs, Ivan Ovcharenko, Laurie Gordon, Susan Lucas, Marcia M. Miller, Hidetoshi Inoko, Takashi Shiina, Jim Kaufman, Jan Salomonsen, Karsten Skjoedt, Gane Ka Shu Wong, Jun Wang, Bin Liu, Jian Wang, Jun Yu, Huanming Yang, Mikhail Nefedov, Maxim Koriabine, Pieter J. DeJong, Leo Goodstadt, Caleb Webber, Nicholas J. Dickens, Ivica Letunic, Mikita Suyama, David Torrents, Christian Von Mering, Evgeny M. Zdobnov, Kateryna Makova, Anton Nekrutenko, Laura Elnitski, Pallavi Eswara, David C. King, Shan Yang, Svitlana Tyekucheva, Anusha Radakrishnan, Robert S. Harris, Francesca Chiaromonte, James Taylor, Jianbin He, Monique Rijnkels, Sam Griffiths-Jones, Abel Ureta-Vidal, Michael M. Hoffman, Jessica Severin, Stephen M.J. Searle, Andy S. Law, David Speed, Dave Waddington, Ze Cheng, Eray Tuzun, Evan Eichler, Zhirong Bao, Paul Flicek, David D. Shteynberg, Michael R. Brent, Jacqueline M. Bye, Elizabeth J. Huckle, Sourav Chatterji, Colin Dewey, Lior Pachter, Andrei Kouranov, Zissimos Mourelatos, Artemis G. Hatzigeorgiou, Andrew H. Paterson, Robert Ivarie, Mikael Brandstrom, Erik Axelsson, Niclas Backstrom, Sofia Berlin, Matthew T. Webster, Olivier Pourquie, Alexandre Reymond, Catherine Ucla, Stylianos E. Antonarakis, Manyuan Long, J. J. Emerson, Esther Betrán, Isabelle Dupanloup, Henrik Kaessmann, Angie S. Hinrichs, Gill Bejerano, Terrence S. Furey, Rachel A. Harte, Brian Raney, Adam Siepel, W. James Kent, David Haussler, Eduardo Eyras, Robert Castelo, Josep F. Abril, Sergi Castellano, Francisco Camara, Genis Parra, Roderic Guigo, Guillaume Bourque, Glenn Tesler, Pavel A. Pevzner, Arian Smit, Lucinda A. Fulton, Elaine R. Mardis, Richard K. Wilson,, Sequence and comparative analysis of the chicken genome provide unique perspectives on vertebrate evolution, Nature, 10.1038/nature03154, 423, 10, 695-777, 2014.12, We present here a draft genome sequence of the red jungle fowl, Gallus gallus. Because the chicken is a modern descendant of the dinosaurs and the first non-mammalian amniote to have its genome sequenced, the draft sequence of its genome - composed of approximately one billion base pairs of sequence and an estimated 20,000-23,000 genes - provides a new perspective on vertebrate genome evolution, while also improving the annotation of mammalian genomes. For example, the evolutionary distance between chicken and human provides high specificity in detecting functional elements, both non-coding and coding. Notably, many conserved non-coding sequences are far from genes and cannot be assigned to defined functional classes. In coding regions the evolutionary dynamics of protein domains and orthologous groups illustrate processes that distinguish the lineages leading to birds and mammals. The distinctive properties of avian microchromosomes, together with the inferred patterns of conserved synteny, provide additional insights into vertebrate chromosome architecture.. |
| 87. |
Hiroaki Okae, Hatsune Chiba, Hitoshi Hiura, Hirotaka Hamada, Akiko Sato, Takafumi Utsunomiya, Hiroyuki Kikuchi, Hiroaki Yoshida, Atsushi Tanaka, Mikita Suyama, Takahiro Arima, Genome-wide analysis of DNA methylation dynamics during early human development., PLoS genetics, 10.1371/journal.pgen.1004868, 10, 12, e1004868, 2014.12, DNA methylation is globally reprogrammed during mammalian preimplantation development, which is critical for normal development. Recent reduced representation bisulfite sequencing (RRBS) studies suggest that the methylome dynamics are essentially conserved between human and mouse early embryos. RRBS is known to cover 5-10% of all genomic CpGs, favoring those contained within CpG-rich regions. To obtain an unbiased and more complete representation of the methylome during early human development, we performed whole genome bisulfite sequencing of human gametes and blastocysts that covered>70% of all genomic CpGs. We found that the maternal genome was demethylated to a much lesser extent in human blastocysts than in mouse blastocysts, which could contribute to an increased number of imprinted differentially methylated regions in the human genome. Global demethylation of the paternal genome was confirmed, but SINE-VNTR-Alu elements and some other tandem repeat-containing regions were found to be specifically protected from this global demethylation. Furthermore, centromeric satellite repeats were hypermethylated in human oocytes but not in mouse oocytes, which might be explained by differential expression of de novo DNA methyltransferases. These data highlight both conserved and species-specific regulation of DNA methylation during early mammalian development. Our work provides further information critical for understanding the epigenetic processes underlying differentiation and pluripotency during early human development.. |
| 88. |
Minako Yoshihara, Li Jiang, Shinya Akatsuka, Mikita Suyama, Shinya Toyokuni, Genome-wide profiling of 8-oxoguanine reveals its association with spatial positioning in nucleus., DNA research : an international journal for rapid publication of reports on genes and genomes, 10.1093/dnares/dsu023, 21, 6, 603-12, 2014.12, 8-Oxoguanine (8-oxoG) is one of the most common DNA lesions generated by reactive oxygen species. In this study, we analysed the genome-wide distribution profile of 8-oxoG by combining immunoprecipitation by antibodies specific for the DNA fragments containing 8-oxoG with a microarray that covers rat genome. Genome-wide mapping of 8-oxoG in normal rat kidney revealed that 8-oxoG is preferentially located at gene deserts. We did not observe differences in 8-oxoG levels between groups of genes with high and low expression, possibly because of the generally low 8-oxoG levels in genic regions compared with gene deserts. The distribution of 8-oxoG and lamina-associated domains (LADs) were strongly correlated, suggesting that the spatial location of genomic DNA in the nucleus determines the susceptibility to oxidative modifications. One possible explanation for high 8-oxoG levels in LADs is that the nuclear periphery is more susceptible to the oxidative damage caused by the extra-nuclear factors. Moreover, LADs have a rather compact conformation, which may limit the recruitment of repair components to the modified bases.. |
| 89. |
Isao Tamura, Yasuyuki Ohkawa, Tetsuya Sato, Mikita Suyama, Kosuke Jozaki, Maki Okada, Lifa Lee, Ryo Maekawa, Hiromi Asada, Shun Sato, Yoshiaki Yamagata, Hiroshi Tamura, Norihiro Sugino, Genome-wide analysis of histone modifications in human endometrial stromal cells, Molecular Endocrinology, 10.1210/me.2014-1117, 28, 10, 1656-1669, 2014.10, Dramatic changes of gene expressions occur in human endometrial stromal cells (ESCs) during decidualization. The changes in gene expression are associated with changes of chromatin structure, which are regulated by histone modifications. Here we investigated genome-wide changes in histone modifications associated with decidualization in human ESCs using chromatin immunoprecipitation combined with next-generation sequencing. ESCs were incubated with estradiol and medroxyprogesterone acetate for 14 days to induce decidualization. The chromatin immunoprecipitation- sequence data showed that induction of decidualization increased H3K27ac and H3K4me3 signals in many genomic regions but decreased in only a few regions. Most of the H3K27ac-increased regions (80%) and half of the H3K4me3-increased regions were located in the distal promoter regions (more than 3 kb upstream or downstream of the transcription start site). RNA sequence showed that induction of decidualization up-regulated 881 genes, 223 of which had H3K27ac- or H3K4me3-increased regions in the proximal and distal promoter regions. Induction of decidualization increased themRNAlevels of these genes more than it increased themRNA levels of genes without H3K27ac- or H3K4me3-increased regions. Pathway analysis revealed that up-regulated genes with the H3K27ac- or H3K4me3-increased regions were associated with the insulin signaling, which may be involved in glucose uptake that is necessary for ESCs to undergo decidualization. These results show that histone modification statuses on a genome-wide basis change in human ESCs during decidualization. The main changes of histone modifications are increases of H3K27ac and H3K4me3 in both the proximal and distal promoter regions, which are involved in the up-regulation of gene expression that occurs during decidualization.. |
| 90. |
Saori Yamaguchi, Tomotoshi Marumoto, Takenobu Nii, Hirotaka Kawano, Jiyuan Liao, Yoko Nagai, Michiyo Okada, Atsushi Takahashi, Hiroyuki Inoue, Erika Sasaki, Hiroshi Fujii, Shinji Okano, Hayao Ebise, Tetsuya Sato, Mikita Suyama, Hideyuki Okano, Yoshie Miura, Kenzaburo Tani, Characterization of common marmoset dysgerminoma-like tumor induced by the lentiviral expression of reprogramming factors, Cancer Science, 10.1111/cas.12367, 105, 4, 402-408, 2014.04, Recent generation of induced pluripotent stem (iPSCs) has made a significant impact on the field of human regenerative medicine. Prior to the clinical application of iPSCs, testing of their safety and usefulness must be carried out using reliable animal models of various diseases. In order to generate iPSCs from common marmoset (CM; Callithrix jacchus), one of the most useful experimental animals, we have lentivirally transduced reprogramming factors, including POU5F1 (also known as OCT3/4), SOX2, KLF4, and c-MYC into CM fibroblasts. The cells formed round colonies expressing embryonic stem cell markers, however, they showed an abnormal karyotype denoted as 46, X, del(4q), +mar, and formed human dysgerminoma-like tumors in SCID mice, indicating that the transduction of reprogramming factors caused unexpected tumorigenesis of CM cells. Moreover, CM dysgerminoma-like tumors were highly sensitive to DNA-damaging agents, irradiation, and fibroblast growth factor receptor inhibitor, and their growth was dependent on c-MYC expression. These results indicate that DNA-damaging agents, irradiation, fibroblast growth factor receptor inhibitor, and c-MYC-targeted therapies might represent effective treatment strategies for unexpected tumors in patients receiving iPSC-based therapy. Reprogramming factors for iPSC generation can be oncogenic, and thus reprogramming factor transduced cells might have tumorigenic potential. Here we characterized common marmoset dysgerminoma like tumor, CM DGs, generated by the lentivirally transduced reprogramming factors into fibroblasts. The growth of CM DGs was dependent on c-MYC expression and bFGF signaling. Moreover CM DGs were highly sensitive to DNA damaging agents, irradiation and FGFR inhibitors. Therefore irradiation, DNA damaging agents and FGFR inhibitors might be effective for controlling reprogramming factor related tumors that may be found in patients treated with iPSC-based medicine.. |
| 91. |
Takashi Baba, Hiroyuki Otake, Tetsuya Sato, Kanako Miyabayashi, Yurina Shishido, Chia Yih Wang, Yuichi Shima, Hiroshi Kimura, Mikako Yagi, Yasuhiro Ishihara, Shinjiro Hino, Hidesato Ogawa, Mitsuyoshi Nakao, Takeshi Yamazaki, Dongchon Kang, Yasuyuki Ohkawa, Mikita Suyama, Bon Chu Chung, Ken Ichirou Morohashi, Glycolytic genes are targets of the nuclear receptor Ad4BP/SF-1, Nature communications, 10.1038/ncomms4634, 5, 2014.04, Genetic deficiencies in transcription factors can lead to the loss of certain types of cells and tissue. The steroidogenic tissue-specific nuclear receptor Ad4BP/SF-1 (NR5A1) is one such gene, because mice in which this gene is disrupted fail to develop the adrenal gland and gonads. However, the specific role of Ad4BP/SF-1 in these biological events remains unclear. Here we use chromatin immunoprecipitation sequencing to show that nearly all genes in the glycolytic pathway are regulated by Ad4BP/SF-1. Suppression of Ad4BP/SF-1 by small interfering RNA reduces production of the energy carriers ATP and nicotinamide adenine dinucleotide phosphate, as well as lowers expression of genes involved in glucose metabolism. Together, these observations may explain tissue dysgenesis as a result of Ad4BP/SF-1 gene disruption in vivo. Considering the function of estrogen-related receptor α, the present study raises the possibility that certain types of nuclear receptors regulate sets of genes involved in metabolic pathways to generate energy carriers.. |
| 92. |
Saori Yamaguchi, Tomotoshi Marumoto, Takenobu Nii, Hirotaka Kawano, Jiyuan Liao, Yoko Nagai, Michiyo Okada, Atsushi Takahashi, Hiroyuki Inoue, Erika Sasaki, Hiroshi Fujii, Shinji Okano, Hayao Ebise, Tetsuya Sato, Mikita Suyama, Hideyuki Okano, Yoshie Miura, Kenzaburo Tani, Characterization of common marmoset dysgerminoma-like tumor induced by the lentiviral expression of reprogramming factors., Cancer science, 10.1111/cas.12367, 105, 4, 402-8, 2014.04, Recent generation of induced pluripotent stem (iPSCs) has made a significant impact on the field of human regenerative medicine. Prior to the clinical application of iPSCs, testing of their safety and usefulness must be carried out using reliable animal models of various diseases. In order to generate iPSCs from common marmoset (CM; Callithrix jacchus), one of the most useful experimental animals, we have lentivirally transduced reprogramming factors, including POU5F1 (also known as OCT3/4), SOX2, KLF4, and c-MYC into CM fibroblasts. The cells formed round colonies expressing embryonic stem cell markers, however, they showed an abnormal karyotype denoted as 46, X, del(4q), +mar, and formed human dysgerminoma-like tumors in SCID mice, indicating that the transduction of reprogramming factors caused unexpected tumorigenesis of CM cells. Moreover, CM dysgerminoma-like tumors were highly sensitive to DNA-damaging agents, irradiation, and fibroblast growth factor receptor inhibitor, and their growth was dependent on c-MYC expression. These results indicate that DNA-damaging agents, irradiation, fibroblast growth factor receptor inhibitor, and c-MYC-targeted therapies might represent effective treatment strategies for unexpected tumors in patients receiving iPSC-based therapy.. |
| 93. |
Takashi Baba, Hiroyuki Otake, Tetsuya Sato, Kanako Miyabayashi, Yurina Shishido, Chia-Yih Wang, Yuichi Shima, Hiroshi Kimura, Mikako Yagi, Yasuhiro Ishihara, Shinjiro Hino, Hidesato Ogawa, Mitsuyoshi Nakao, Takeshi Yamazaki, Dongchon Kang, Yasuyuki Ohkawa, Mikita Suyama, Bon-Chu Chung, Ken-Ichirou Morohashi, Glycolytic genes are targets of the nuclear receptor Ad4BP/SF-1., Nature communications, 10.1038/ncomms4634, 5, 3634-3634, 2014.04, Genetic deficiencies in transcription factors can lead to the loss of certain types of cells and tissue. The steroidogenic tissue-specific nuclear receptor Ad4BP/SF-1 (NR5A1) is one such gene, because mice in which this gene is disrupted fail to develop the adrenal gland and gonads. However, the specific role of Ad4BP/SF-1 in these biological events remains unclear. Here we use chromatin immunoprecipitation sequencing to show that nearly all genes in the glycolytic pathway are regulated by Ad4BP/SF-1. Suppression of Ad4BP/SF-1 by small interfering RNA reduces production of the energy carriers ATP and nicotinamide adenine dinucleotide phosphate, as well as lowers expression of genes involved in glucose metabolism. Together, these observations may explain tissue dysgenesis as a result of Ad4BP/SF-1 gene disruption in vivo. Considering the function of estrogen-related receptor α, the present study raises the possibility that certain types of nuclear receptors regulate sets of genes involved in metabolic pathways to generate energy carriers.. |
| 94. |
Mikita Suyama, Mechanistic insights into mutually exclusive splicing in dynamin 1., Bioinformatics (Oxford, England), 10.1093/bioinformatics/btt368, 29, 17, 2084-7, 2013.09, Mutually exclusive splicing is a strictly regulated pattern of alternative splicing. A specific group of mutually exclusive splicing events has been shown to be regulated by the formation of specific RNA secondary structures. This type of regulation has been shown to exist only in arthropods. The present study involved a detailed sequence analysis of human gene structures that undergo mutually exclusive splicing, which showed that this type of regulation may also occur in dynamin 1 in mammals. A phylogenetic analysis revealed that the dynamin 1 orthologs in invertebrates did not share the same sequence features, which suggests that the regulatory mechanism has independently evolved in the mammalian lineage. Therefore, the emergence of this elaborate mechanism for mutually exclusive splicing may be attributable to mechanistic convergence.. |
| 95. |
Mikita Suyama, Mechanistic insights into mutually exclusive splicing in dynamin 1., Bioinformatics, 10.1093/bioinformatics/btt368, 29, 17, 2084-2087, 2013.06. |
| 96. |
Yasunobu Miyake, Kenji Toyonaga, Daiki Mori, Shigeru Kakuta, Yoshihiko Hoshino, Akiko Oyamada, Hisakata Yamada, Ken Ichiro Ono, Mikita Suyama, Yoichiro Iwakura, Yasunobu Yoshikai, Sho Yamasaki, C-type Lectin MCL Is an FcRγ-Coupled Receptor that Mediates the Adjuvanticity of Mycobacterial Cord Factor, Immunity, 10.1016/j.immuni.2013.03.010, 38, 5, 1050-1062, 2013.05, Cord factor, also called trehalose-6,6'-dimycolate (TDM), is a potent mycobacterial adjuvant. We herein report that the C-type lectin MCL (also called Clec4d) is a TDM receptor that is likely to arise from gene duplication of Mincle (also called Clec4e). Mincle isknown to be an inducible receptor recognizing TDM, whereas MCL was constitutively expressed in myeloid cells. To examine the contribution of MCL in response to TDM adjuvant, we generated MCL-deficient mice. TDM promoted innate immune responses, such as granuloma formation, which was severely impaired in MCL-deficient mice. TDM-induced acquired immune responses, such as experimental autoimmune encephalomyelitis (EAE), was almost completely dependent on MCL, but not Mincle. Furthermore, by generating Clec4egfp reporter mice, we found that MCL was also crucial for driving Mincle induction upon TDM stimulation. These results suggest that MCL is an FcRγ-coupled activating receptor that mediates the adjuvanticity ofTDM. •MCL is an ITAM-coupled TDM receptor that arises from gene duplication of Mincle•Innate and acquired immunity induced by TDM are impaired in MCL-deficient mice•MCL drives Mincle expression in dendritic cells upon TDM stimulation•MCL, but not Mincle, is critically involved in EAE induced by TDM adjuvant.. |
| 97. |
Yasunobu Miyake, Kenji Toyonaga, Daiki Mori, Shigeru Kakuta, Yoshihiko Hoshino, Akiko Oyamada, Hisakata Yamada, Ken-Ichiro Ono, Mikita Suyama, Yoichiro Iwakura, Yasunobu Yoshikai, Sho Yamasaki, C-type lectin MCL is an FcRγ-coupled receptor that mediates the adjuvanticity of mycobacterial cord factor., Immunity, 10.1016/j.immuni.2013.03.010, 38, 5, 1050-62, 2013.05, Cord factor, also called trehalose-6,6'-dimycolate (TDM), is a potent mycobacterial adjuvant. We herein report that the C-type lectin MCL (also called Clec4d) is a TDM receptor that is likely to arise from gene duplication of Mincle (also called Clec4e). Mincle is known to be an inducible receptor recognizing TDM, whereas MCL was constitutively expressed in myeloid cells. To examine the contribution of MCL in response to TDM adjuvant, we generated MCL-deficient mice. TDM promoted innate immune responses, such as granuloma formation, which was severely impaired in MCL-deficient mice. TDM-induced acquired immune responses, such as experimental autoimmune encephalomyelitis (EAE), was almost completely dependent on MCL, but not Mincle. Furthermore, by generating Clec4e(gfp) reporter mice, we found that MCL was also crucial for driving Mincle induction upon TDM stimulation. These results suggest that MCL is an FcRγ-coupled activating receptor that mediates the adjuvanticity of TDM.. |
| 98. |
M. Villacorte, K. Suzuki, A. Hirasawa, Yasuyuki Ohkawa, Mikita Suyama, T. Maruyama, D. Aoki, Y. Ogino, S. Miyagawa, T. Terabayashi, Y. Tomooka, N. Nakagata, G. Yamada, β-Catenin signaling regulates Foxa2 expression during endometrial hyperplasia formation., Oncogene, 10.1038/onc.2012.376, 2012.09. |
| 99. |
Chanavee Ratanajaraya, Hiroyuki Nishiyama, Meiko Takahashi, Takahisa Kawaguchi, Ryoichi Saito, Yoshiki Mikami, Mikita Suyama, Mark Lathrop, Ryo Yamada, Osamu Ogawa, Fumihiko Matsuda, A polymorphism of the POLG2 gene is genetically associated with the invasiveness of urinary bladder cancer in Japanese males., Journal of Human Genetics, 10.1038/jhg.2011.60, 56, 8, 572-576, 2011.08, Urinary bladder cancer (UBC) is a common cancer with male predominance. Pathologically it is classified into two distinct tumor entities related to the risk of patients. The low-grade tumors with relatively well-differentiated tumor histology (G1 and G2) at stage Ta are non-invasive and pose a minimal risk, whereas high-grade tumors (G2 and G3) with stages T1 to T4 are aggressive with invasion, and therefore, pose a serious risk for the patients. DNA repair and metabolic process genes may have major roles in cancer progression and development. To identify genes associated with invasiveness of UBC, we have extensively genotyped 802 single nucleotide polymorphisms in 114 genes related to DNA repair mechanisms and metabolic processes. A genetic association study was performed between non-invasive (G1 and G2 with Ta) and invasive (G2 and G3 with T1 to T4) groups of Japanese UBC patients. We found that rs17650301 in POLG2 showed marked difference in genotype distribution between the two groups in males (P=6.93 × 10−4), which was further confirmed in an independent sample set (overall P=1.67 × 10−4). We also found by an in silico analysis that the risk allele of rs17650301 increased the transcription of POLG2. In conclusion, rs17650301 is a good candidate marker for UBC invasiveness in Japanese males.. |
| 100. |
Mikita Suyama, Eoghan D Harrington, Svetlana Vinokourova, Magnus von Knebel Doeberitz, Osamu Ohara, Peer Bork, A network of conserved co-occurring motifs for the regulation of alternative splicing., Nucleic acids research, 10.1093/nar/gkq705, 38, 22, 7916-26, 2010.12, Cis-acting short sequence motifs play important roles in alternative splicing. It is now possible to identify such sequence motifs as conserved sequence patterns in genome sequence alignments. Here, we report the systematic search for motifs in the neighboring introns of alternatively spliced exons by using comparative analysis of mammalian genome alignments. We identified 11 conserved sequence motifs that might be involved in the regulation of alternative splicing. These motifs are not only significantly overrepresented near alternatively spliced exons, but they also co-occur with each other, thus, forming a network of cis-elements, likely to be the basis for context-dependent regulation. Based on this finding, we applied the motif co-occurrence to predict alternatively skipped exons. We verified exon skipping in 29 cases out of 118 predictions (25%) by EST and mRNA sequences in the databases. For the predictions not verified by the database sequences, we confirmed exon skipping in 10 additional cases by using both RT-PCR experiments and the publicly available RNA-Seq data. These results indicate that even more alternative splicing events will be found with the progress of large-scale and high-throughput analyses for various tissue samples and developmental stages.. |
| 101. |
Suyama M, Harrington ED, Vinokourova S, von Knebel Doeberitz M, Ohara O, Bork P., A network of conserved co-occurring motifs for the regulation of alternative splicing., Nucleic Acids Res., 10.1093/nar/gkq705, 38, 22, 7916-7926, 2010.08, Cis-acting short sequence motifs play important roles in alternative splicing. It is now possible to identify such sequence motifs as conserved sequence patterns in genome sequence alignments. Here, we report the systematic search for motifs in the neighboring introns of alternatively spliced exons by using comparative analysis of mammalian genome alignments. We identified 11 conserved sequence motifs that might be involved in the regulation of alternative splicing. These motifs are not only significantly overrepresented near alternatively spliced exons, but they also co-occur with each other, thus, forming a network of cis-elements, likely to be the basis for context-dependent regulation. Based on this finding, we applied the motif co-occurrence to predict alternatively skipped exons. We verified exon skipping in 29 cases out of 118 predictions (25%) by EST and mRNA sequences in the databases. For the predictions not verified by the database sequences, we confirmed exon skipping in 10 additional cases by using both RT-PCR experiments and the publicly available RNA-Seq data. These results indicate that even more alternative splicing events will be found with the progress of large-scale and high-throughput analyses for various tissue samples and developmental stages.. |
| 102. |
Marc Güell, Vera Van Noort, Eva Yus, Wei Hua Chen, Justine Leigh-Bell, Konstantinos Michalodimitrakis, Takuji Yamada, Manimozhiyan Arumugam, Tobias Doerks, Sebastian Kühner, Michaela Rode, Mikita Suyama, Sabine Schmidt, Anne Claude Gavin, Peer Bork, Luis Serrano, Transcriptome complexity in a genome-reduced bacterium, Science, 10.1126/science.1176951, 326, 5957, 1268-1271, 2009.11, To study basic principles of transcriptome organization in bacteria, we analyzed one of the smallest self-replicating organisms. Mycoplasma pneumoniae. We combined strand-specific tiling arrays, complemented by transcriptome sequencing, with more than 252 spotted arrays. We detected 117 previously undescribed, mostly noncoding transcripts, 89 of them in antisense configuration to known genes. We identified 341 operons, of which 139 are polycistronic; almost half of the latter show decaying expression in a staircase-like manner. Under various conditions, operons could be divided into 447 smaller transcriptional units, resulting in many alternative transcripts. Frequent antisense transcripts, alternative transcripts, and multiple regulators per gene imply a highly dynamic transcriptome, more similar to that of eukaryotes than previously thought.. |
| 103. |
Marc Güell, Vera van Noort, Eva Yus, Wei-Hua Chen, Justine Leigh-Bell, Konstantinos Michalodimitrakis, Takuji Yamada, Manimozhiyan Arumugam, Tobias Doerks, Sebastian Kühner, Michaela Rode, Mikita Suyama, Sabine Schmidt, Anne-Claude Gavin, Peer Bork, Luis Serrano, Transcriptome complexity in a genome-reduced bacterium., Science (New York, N.Y.), 10.1126/science.1176951, 326, 5957, 1268-71, 2009.11, To study basic principles of transcriptome organization in bacteria, we analyzed one of the smallest self-replicating organisms, Mycoplasma pneumoniae. We combined strand-specific tiling arrays, complemented by transcriptome sequencing, with more than 252 spotted arrays. We detected 117 previously undescribed, mostly noncoding transcripts, 89 of them in antisense configuration to known genes. We identified 341 operons, of which 139 are polycistronic; almost half of the latter show decaying expression in a staircase-like manner. Under various conditions, operons could be divided into 447 smaller transcriptional units, resulting in many alternative transcripts. Frequent antisense transcripts, alternative transcripts, and multiple regulators per gene imply a highly dynamic transcriptome, more similar to that of eukaryotes than previously thought.. |
| 104. |
Junji Itou, Mikita Suyama, Yukio Imamura, Tomonori Deguchi, Kazuhiro Fujimori, Shunsuke Yuba, Yutaka Kawarabayasi, Takashi Kawasaki, Functional and comparative genomics analyses of pmp22 in medaka fish, BMC Neuroscience, 10.1186/1471-2202-10-60, 10, 2009.06, Background: Pmp22, a member of the junction protein family Claudin/EMP/ PMP22, plays an important role in myelin formation. Increase of pmp22 transcription causes peripheral neuropathy, Charcot-Marie-Tooth disease type1A (CMT1A). The pathophysiological phenotype of CMT1A is aberrant axonal myelination which induces a reduction in nerve conduction velocity (NCV). Several CMT1A model rodents have been established by overexpressing pmp22. Thus, it is thought that pmp22 expression must be tightly regulated for correct myelin formation in mammals. Interestingly, the myelin sheath is also present in other jawed vertebrates. The purpose of this study is to analyze the evolutionary conservation of the association between pmp22 transcription level and vertebrate myelin formation, and to find the conserved non-coding sequences for pmp22 regulation by comparative genomics analyses between jawed fishes and mammals. Results: A transgenic pmp22 over-expression medaka fish line was established. The transgenic fish had approximately one fifth the peripheral NCV values of controls, and aberrant myelination of transgenic fish in the peripheral nerve system (PNS) was observed. We successfully confirmed that medaka fish pmp22 has the same exon-intron structure as mammals, and identified some known conserved regulatory motifs. Furthermore, we found novel conserved sequences in the first intron and 3'UTR. Conclusion: Medaka fish undergo abnormalities in the PNS when pmp22 transcription increases. This result indicates that an adequate pmp22 transcription level is necessary for correct myelination of jawed vertebrates. Comparison of pmp22 orthologs between distantly related species identifies evolutionary conserved sequences that contribute to precise regulation of pmp22 expression.. |
| 105. |
Carlos Quijano, Pavel Tomancak, Jesus Lopez-Marti, Mikita Suyama, Peer Bork, Marco Milan, David Torrents, Miguel Manzanares, Selective maintenance of Drosophila tandemly arranged duplicated genes during evolution, Genome biology, 10.1186/gb-2008-9-12-r176, 9, 12, 2008.12, Background: The physical organization and chromosomal localization of genes within genomes is known to play an important role in their function. Most genes arise by duplication and move along the genome by random shuffling of DNA segments. Higher order structuring of the genome occurs in eukaryotes, where groups of physically linked genes are co-expressed. However, the contribution of gene duplication to gene order has not been analyzed in detail, as it is believed that co-expression due to recent duplicates would obscure other domains of co-expression. Results: We have catalogued ordered duplicated genes in Drosophila melanogaster, and found that one in five of all genes is organized as tandem arrays. Furthermore, among arrays that have been spatially conserved over longer periods than would be expected on the basis of random shuffling, a disproportionate number contain genes encoding developmental regulators. Using in situ gene expression data for more than half of the Drosophila genome, we find that genes in these conserved clusters are co-expressed to a much higher extent than other duplicated genes. Conclusions: These results reveal the existence of functional constraints in insects that retain copies of genes encoding developmental and regulatory proteins as neighbors, allowing their co-expression. This co-expression may be the result of shared cis-regulatory elements or a shared need for a specific chromatin structure. Our results highlight the association between genome architecture and the gene regulatory networks involved in the construction of the body plan.. |
| 106. |
Giulia Soldà, Mikita Suyama, Paride Pelucchi, Silvia Boi, Alessandro Guffanti, Ermanno Rizzi, Peer Bork, Maria Luisa Tenchini, Francesca D. Ciccarelli, Non-random retention of protein-coding overlapping genes in Metazoa, BMC genomics, 10.1186/1471-2164-9-174, 9, 2008.04, Background: Although the overlap of transcriptional units occurs frequently in eukaryotic genomes, its evolutionary and biological significance remains largely unclear. Here we report a comparative analysis of overlaps between genes coding for well-annotated proteins in five metazoan genomes (human, mouse, zebrafish, fruit fly and worm). Results: For all analyzed species the observed number of overlapping genes is always lower than expected assuming functional neutrality, suggesting that gene overlap is negatively selected. The comparison to the random distribution also shows that retained overlaps do not exhibit random features: antiparallel overlaps are significantly enriched, while overlaps lying on the same strand and those involving coding sequences are highly underrepresented. We confirm that overlap is mostly species-specific and provide evidence that it frequently originates through the acquisition of terminal, non-coding exons. Finally, we show that overlapping genes tend to be significantly co-expressed in a breast cancer cDNA library obtained by 454 deep sequencing, and that different overlap types display different patterns of reciprocal expression. Conclusion: Our data suggest that overlap between protein-coding genes is selected against in Metazoa. However, when retained it may be used as a species-specific mechanism for the reciprocal regulation of neighboring genes. The tendency of overlaps to involve non-coding regions of the genes leads to the speculation that the advantages achieved by an overlapping arrangement may be optimized by evolving regulatory non-coding transcripts.. |
| 107. |
Mikita Suyama, Eoghan Harrington, Peer Bork, David Torrents, Identification and analysis of genes and pseudogenes within duplicated regions in the human and mouse genomes, PLoS Computational Biology, 10.1371/journal.pcbi.0020076, 2, 6, 627-636, 2006.07, The identification and classification of genes and pseudogenes in duplicated regions still constitutes a challenge for standard automated genome annotation procedures. Using an integrated homology and orthology analysis independent of current gene annotation, we have identified 9,484 and 9,017 gene duplicates in human and mouse, respectively. On the basis of the integrity of their coding regions, we have classified them into functional and inactive duplicates, allowing us to define the first consistent and comprehensive collection of 1,811 human and 1,581 mouse unprocessed pseudogenes. Furthermore, of the total of 14,172 human and mouse duplicates predicted to be functional genes, as many as 420 are not included in current reference gene databases and therefore correspond to likely novel mammalian genes. Some of these correspond to partial duplicates with less than half of the length of the original source genes, yet they are conserved and syntenic among different mammalian lineages. The genes and unprocessed pseudogenes obtained here will enable further studies on the mechanisms involved in gene duplication as well as of the fate of duplicated genes.. |
| 108. |
Mikita Suyama, David Torrents, Peer Bork, PAL2NAL
Robust conversion of protein sequence alignments into the corresponding codon alignments, Nucleic acids research, 10.1093/nar/gkl315, 34, WEB. SERV. ISS., W609-W612, 2006.07, PAL2NAL is a web server that constructs a multiple codon alignment from the corresponding aligned protein sequences. Such codon alignments can be used to evaluate the type and rate of nucleotide substitutions in coding DNA for a wide range of evolutionary analyses, such as the identification of levels of selective constraint acting on genes, or to perform DNA-based phylogenetic studies. The server takes a protein sequence alignment and the corresponding DNA sequences as input. In contrast to other existing applications, this server is able to construct codon alignments even if the input DNA sequence has mismatches with the input protein sequence, or contains untranslated regions and polyA tails. The server can also deal with frame shifts and inframe stop codons in the input models, and is thus suitable for the analysis of pseudogenes. Another distinct feature is that the user can specify a subregion of the input alignment in order to specifically analyze functional domains or exons of interest. The PAL2NAL server is available at http://www.bork.embl.de/pal2nal.. |
| 109. |
Mikita Suyama, David Torrents, Peer Bork, PAL2NAL: robust conversion of protein sequence alignments into the corresponding codon alignments., Nucleic acids research, 34, Web Server issue, W609-12, 2006.07, PAL2NAL is a web server that constructs a multiple codon alignment from the corresponding aligned protein sequences. Such codon alignments can be used to evaluate the type and rate of nucleotide substitutions in coding DNA for a wide range of evolutionary analyses, such as the identification of levels of selective constraint acting on genes, or to perform DNA-based phylogenetic studies. The server takes a protein sequence alignment and the corresponding DNA sequences as input. In contrast to other existing applications, this server is able to construct codon alignments even if the input DNA sequence has mismatches with the input protein sequence, or contains untranslated regions and polyA tails. The server can also deal with frame shifts and inframe stop codons in the input models, and is thus suitable for the analysis of pseudogenes. Another distinct feature is that the user can specify a subregion of the input alignment in order to specifically analyze functional domains or exons of interest. The PAL2NAL server is available at http://www.bork.embl.de/pal2nal.. |
| 110. |
Mikita Suyama, Eoghan Harrington, Peer Bork, David Torrents, Identification and analysis of genes and pseudogenes within duplicated regions in the human and mouse genomes., PLoS computational biology, 2, 6, e76, 2006.06, The identification and classification of genes and pseudogenes in duplicated regions still constitutes a challenge for standard automated genome annotation procedures. Using an integrated homology and orthology analysis independent of current gene annotation, we have identified 9,484 and 9,017 gene duplicates in human and mouse, respectively. On the basis of the integrity of their coding regions, we have classified them into functional and inactive duplicates, allowing us to define the first consistent and comprehensive collection of 1,811 human and 1,581 mouse unprocessed pseudogenes. Furthermore, of the total of 14,172 human and mouse duplicates predicted to be functional genes, as many as 420 are not included in current reference gene databases and therefore correspond to likely novel mammalian genes. Some of these correspond to partial duplicates with less than half of the length of the original source genes, yet they are conserved and syntenic among different mammalian lineages. The genes and unprocessed pseudogenes obtained here will enable further studies on the mechanisms involved in gene duplication as well as of the fate of duplicated genes.. |
| 111. |
Atsushi Kobayashi, Takashi Tonozuka, Kimihiko Sato, Mikita Suyama, Jun Sasaki, Batbold Nyamdawaa, Masayoshi Sakaguchi, Yoshiyuki Sakano, Molecular cloning and characterization of an enzyme hydrolyzing p-nitrophenyl α-D-glucoside from Bacillus stearothermophilus SA0301, Bioscience, Biotechnology and Biochemistry, 10.1271/bbb.70.495, 70, 2, 495-499, 2006.03, Bacillus stearothermophilus SA0301 produces an extracellular oligo-1,6-glucosidase (bsO16G) that also hydrolyzes p-nitrophenyl α-D-glucoside (Tonozuka et al., J. Appl. Glycosci., 45, 397-400 (1998)). We cloned a gene for an enzyme hydrolyzing p-nitrophenyl α-D-glucoside, which was different from the one mentioned above, from B. stearothermophilus SA0301. The k0/Km values of bsO16G for isomaltotriose and isomaltose were 13.2 and 1.39 s-1·mM-1 respectively, while the newly cloned enzyme did not hydrolyze isomaltotriose, and the k0/Km value for isomaltose was 0.81 s -1·mM-1. The primary structure of the cloned enzyme more closely resembled those of trehalose-6-phosphate hydrolases than those of oligo-1,6-glucosidases, and the cloned enzyme hydrolyzed trehalose 6-phosphate. An open reading frame encoding a protein homologous to the trehalose-specific IIBC component of the phopshotransferase system was also found upstream of the gene for this enzyme.. |
| 112. |
Atsushi Kobayashi, Takashi Tonozuka, Kimihiko Sato, Mikita Suyama, Jun Sasaki, Batbold Nyamdawaa, Masayoshi Sakaguchi, Yoshiyuki Sakano, Molecular cloning and characterization of an enzyme hydrolyzing p-nitrophenyl alpha-D-glucoside from Bacillus stearothermophilus SA0301., Bioscience, biotechnology, and biochemistry, 70, 2, 495-9, 2006.02, Bacillus stearothermophilus SA0301 produces an extracellular oligo-1,6-glucosidase (bsO16G) that also hydrolyzes p-nitrophenyl alpha-D-glucoside (Tonozuka et al., J. Appl. Glycosci., 45, 397-400 (1998)). We cloned a gene for an enzyme hydrolyzing p-nitrophenyl alpha-D-glucoside, which was different from the one mentioned above, from B. stearothermophilus SA0301. The k(0)/K(m) values of bsO16G for isomaltotriose and isomaltose were 13.2 and 1.39 s(-1).mM(-1) respectively, while the newly cloned enzyme did not hydrolyze isomaltotriose, and the k(0)/K(m) value for isomaltose was 0.81 s(-1).mM(-1). The primary structure of the cloned enzyme more closely resembled those of trehalose-6-phosphate hydrolases than those of oligo-1,6-glucosidases, and the cloned enzyme hydrolyzed trehalose 6-phosphate. An open reading frame encoding a protein homologous to the trehalose-specific IIBC component of the phopshotransferase system was also found upstream of the gene for this enzyme.. |
| 113. |
Mikita Suyama, Warren C. Lathe, Peer Bork, Palindromic repetitive DNA elements with coding potential in Methanocaldococcus jannaschii, FEBS Letters, 10.1016/j.febslet.2005.08.051, 579, 24, 5281-5286, 2005.10, We have identified 141 novel palindromic repetitive elements in the genome of euryarchaeon Methanocaldococcus jannaschii. The total length of these elements is 14.3 kb, which corresponds to 0.9% of the total genomic sequence and 6.3% of all extragenic regions. The elements can be divided into three groups (MJRE1-3) based on the sequence similarity. The low sequence identity within each of the groups suggests rather old origin of these elements in M. jannaschii. Three MJRE2 elements were located within the protein coding regions without disrupting the coding potential of the host genes, indicating that insertion of repeats might be a widespread mechanism to enhance sequence diversity in coding regions.. |
| 114. |
Mikita Suyama, Warren C Lathe 3rd, Peer Bork, Palindromic repetitive DNA elements with coding potential in Methanocaldococcus jannaschii., FEBS letters, 579, 24, 5281-6, 2005.10, We have identified 141 novel palindromic repetitive elements in the genome of euryarchaeon Methanocaldococcus jannaschii. The total length of these elements is 14.3kb, which corresponds to 0.9% of the total genomic sequence and 6.3% of all extragenic regions. The elements can be divided into three groups (MJRE1-3) based on the sequence similarity. The low sequence identity within each of the groups suggests rather old origin of these elements in M. jannaschii. Three MJRE2 elements were located within the protein coding regions without disrupting the coding potential of the host genes, indicating that insertion of repeats might be a widespread mechanism to enhance sequence diversity in coding regions.. |
| 115. |
, Tarjei S. Mikkelsen, Ladeana W. Hillier, Evan E. Eichler, Michael C. Zody, David B. Jaffe, Shiaw Pyng Yang, Wolfgang Enard, Ines Hellmann, Kerstin Lindblad-Toh, Tasha K. Altheide, Nicoletta Archidiacono, Peer Bork, Jonathan Butler, Jean L. Chang, Ze Cheng, Asif T. Chinwalla, Pieter Dejong, Kimberley D. Delehaunty, Catrina C. Fronick, Lucinda L. Fulton, Yoav Gilad, Gustavo Glusman, Sante Gnerre, Tina A. Graves, Toshiyuki Hayakawa, Karen E. Hayden, Xiaoqiu Huang, Hongkai Ji, W. James Kent, Mary Claire King, Edward J. Kulbokas, Ming K. Lee, Ge Liu, Carlos Lopez-Otin, Kateryna D. Makova, Orna Man, Elaine R. Mardis, Evan Mauceli, Tracie L. Miner, William E. Nash, Joanne O. Nelson, Svante Pääbo, Nick J. Patterson, Craig S. Pohl, Katherine S. Pollard, Kay Prüfer, Xose S. Puente, David Reich, Mariano Rocchi, Kate Rosenbloom, Initial sequence of the chimpanzee genome and comparison with the human genome, Nature, 10.1038/nature04072, 437, 7055, 69-87, 2005.09, Here we present a draft genome sequence of the common chimpanzee (Pan troglodytes). Through comparison with the human genome, we have generated a largely complete catalogue of the genetic differences that have accumulated since the human and chimpanzee species diverged from our common ancestor, constituting approximately thirty-five million single-nucleotide changes, five million insertion/deletion events, and various chromosomal rearrangements. We use this catalogue to explore the magnitude and regional variation of mutational forces shaping these two genomes, and the strength of positive and negative selection acting on their genes. In particular, we find that the patterns of evolution in human and chimpanzee protein-coding genes are highly correlated and dominated by the fixation of neutral and slightly deleterious alleles. We also use the chimpanzee genome as an outgroup to investigate human population genetics and identify signatures of selective sweeps in recent human evolution.. |
| 116. |
La Deana W. Hillier, Tina A. Graves, Robert S. Fulton, Lucinda A. Fulton, Kymberlie H. Pepin, Patrick Minx, Caryn Wagner-McPherson, Dan Layman, Kristine Wylie, Mandeep Sekhon, Michael C. Becker, Ginger A. Fewell, Kimberly D. Delehaunty, Tracie L. Miner, William E. Nash, Colin Kremitzki, Lachlan Oddy, Hui Du, Hui Sun, Holland Bradshaw-Cordum, Johar Ali, Jason Carter, Matt Cordes, Anthony Harris, Amber Isak, Andrew Van Brunt, Christine Nguyen, Feiyu Du, Laura Courtney, Joelle Kalicki, Philip Ozersky, Scott Abbott, Jon Armstrong, Edward A. Belter, Lauren Caruso, Maria Cedroni, Marc Cotton, Teresa Davidson, Anu Desai, Glendoria Elliott, Thomas Erb, Catrina Fronick, Tony Gaige, William Haakenson, Krista Haglund, Andrea Holmes, Richard Harkins, Kyung Kim, Scott S. Kruchowski, Cynthia Madsen Strong, Neenu Grewal, Ernest Goyea, Shunfang Hou, Andrew Levy, Scott Martinka, Kelly Mead, Michael D. McLellan, Rick Meyer, Jennifer Randall-Maher, Chad Tomlinson, Sara Dauphin-Kohlberg, Amy Kozlowicz-Reilly, Neha Shah, Sharhonda Swearengen-Shahid, Jacqueline Snider, Joseph T. Strong, Johanna Thompson, Martin Yoakum, Shawn Leonard, Charlene Pearman, Lee Trani, Maxim Radionenko, Jason E. Waligorski, Chunyan Wang, Susan M. Rock, Aye Mon Tin-Wollam, Rachel Maupin, Phil Latreille, Michael C. Wendl, Shiaw Pyng Yang, Craig Pohl, John W. Wallis, John Spieth, Tamberlyn A. Bieri, Nicolas Berkowicz, Joanne O. Nelson, John Osborne, Li Ding, Rekha Meyer, Aniko Sabo, Yoram Shotland, Prashant Sinha, Patricia E. Wohldmann, Lisa L. Cook, Matthew T. Hickenbotham, James Eldred, Donald Williams, Thomas A. Jones, Xinwei She, Francesca D. Ciccarelli, Elisa Izaurralde, James Taylor, Jeremy Schmutz, Richard M. Myers, David R. Cox, Xiaoqiu Huang, John D. McPherson, Elaine R. Mardis, Sandra W. Clifton, Wesley C. Warren, Asif T. Chinwalla, Sean R. Eddy, Marco A. Marra, Ivan Ovcharenko, Terrence S. Furey, Webb Miller, Evan E. Eichler, Peer Bork, Mikita Suyama, David Torrents, Robert H. Waterston, Richard K. Wilson, Generation and annotation of the DNA sequences of human chromosomes 2 and 4, Nature, 10.1038/nature03466, 434, 7034, 724-731, 2005.04, Human chromosome 2 is unique to the human lineage in being the product of a head-to-head fusion of two intermediate-sized ancestral chromosomes. Chromosome 4 has received attention primarily related to the search for the Huntington's disease gene, but also for genes associated with Wolf-Hirschhorn syndrome, polycystic kidney disease and a form of muscular dystrophy. Here we present approximately 237 million base pairs of sequence for chromosome 2, and 186 million base pairs for chromosome 4, representing more than 99.6% of their euchromatic sequences. Our initial analyses have identified 1,346 protein-coding genes and 1,239 pseudogenes on chromosome 2, and 796 protein-coding genes and 778 pseudogenes on chromosome 4. Extensive analyses confirm the underlying construction of the sequence, and expand our understanding of the structure and evolution of mammalian chromosomes, including gene deserts, segmental duplications and highly variant regions.. |
| 117. |
Ladeana W Hillier, Tina A Graves, Robert S Fulton, Lucinda A Fulton, Kymberlie H Pepin, Patrick Minx, Caryn Wagner-McPherson, Dan Layman, Kristine Wylie, Mandeep Sekhon, Michael C Becker, Ginger A Fewell, Kimberly D Delehaunty, Tracie L Miner, William E Nash, Colin Kremitzki, Lachlan Oddy, Hui Du, Hui Sun, Holland Bradshaw-Cordum, Johar Ali, Jason Carter, Matt Cordes, Anthony Harris, Amber Isak, Andrew van Brunt, Christine Nguyen, Feiyu Du, Laura Courtney, Joelle Kalicki, Philip Ozersky, Scott Abbott, Jon Armstrong, Edward A Belter, Lauren Caruso, Maria Cedroni, Marc Cotton, Teresa Davidson, Anu Desai, Glendoria Elliott, Thomas Erb, Catrina Fronick, Tony Gaige, William Haakenson, Krista Haglund, Andrea Holmes, Richard Harkins, Kyung Kim, Scott S Kruchowski, Cynthia Madsen Strong, Neenu Grewal, Ernest Goyea, Shunfang Hou, Andrew Levy, Scott Martinka, Kelly Mead, Michael D McLellan, Rick Meyer, Jennifer Randall-Maher, Chad Tomlinson, Sara Dauphin-Kohlberg, Amy Kozlowicz-Reilly, Neha Shah, Sharhonda Swearengen-Shahid, Jacqueline Snider, Joseph T Strong, Johanna Thompson, Martin Yoakum, Shawn Leonard, Charlene Pearman, Lee Trani, Maxim Radionenko, Jason E Waligorski, Chunyan Wang, Susan M Rock, Aye-Mon Tin-Wollam, Rachel Maupin, Phil Latreille, Michael C Wendl, Shiaw-Pyng Yang, Craig Pohl, John W Wallis, John Spieth, Tamberlyn A Bieri, Nicolas Berkowicz, Joanne O Nelson, John Osborne, Li Ding, Rekha Meyer, Aniko Sabo, Yoram Shotland, Prashant Sinha, Patricia E Wohldmann, Lisa L Cook, Matthew T Hickenbotham, James Eldred, Donald Williams, Thomas A Jones, Xinwei She, Francesca D Ciccarelli, Elisa Izaurralde, James Taylor, Jeremy Schmutz, Richard M Myers, David R Cox, Xiaoqiu Huang, John D McPherson, Elaine R Mardis, Sandra W Clifton, Wesley C Warren, Asif T Chinwalla, Sean R Eddy, Marco A Marra, Ivan Ovcharenko, Terrence S Furey, Webb Miller, Evan E Eichler, Peer Bork, Mikita Suyama, David Torrents, Robert H Waterston, Richard K Wilson, Generation and annotation of the DNA sequences of human chromosomes 2 and 4., Nature, 434, 7034, 724-31, 2005.04, Human chromosome 2 is unique to the human lineage in being the product of a head-to-head fusion of two intermediate-sized ancestral chromosomes. Chromosome 4 has received attention primarily related to the search for the Huntington's disease gene, but also for genes associated with Wolf-Hirschhorn syndrome, polycystic kidney disease and a form of muscular dystrophy. Here we present approximately 237 million base pairs of sequence for chromosome 2, and 186 million base pairs for chromosome 4, representing more than 99.6% of their euchromatic sequences. Our initial analyses have identified 1,346 protein-coding genes and 1,239 pseudogenes on chromosome 2, and 796 protein-coding genes and 778 pseudogenes on chromosome 4. Extensive analyses confirm the underlying construction of the sequence, and expand our understanding of the structure and evolution of mammalian chromosomes, including gene deserts, segmental duplications and highly variant regions.. |
| 118. |
Francesca D. Ciccarelli, Christian von Mering, Mikita Suyama, Eoghan D. Harrington, Elisa Izaurralde, Peer Bork, Complex genomic rearrangements lead to novel primate gene function, Genome Research, 10.1101/gr.3266405, 15, 3, 343-351, 2005.03, Orthologous genes that maintain a single-copy status in a broad range of species may indicate a selection against gene duplication. If this is the case, then duplicates of such genes that do survive may have escaped the dosage control by rapid and sizable changes in their function. To test this hypothesis and to develop a strategy for the identification of novel gene functions, we have analyzed 22 primate-specific intrachromosomal duplications of genes with a single-copy ortholog in all other completely sequenced metazoans. When comparing this set to genes not exposed to the single-copy status constraint, we observed a higher tendency of the former to modify their gene structure, often through complex genomic rearrangements. The analysis of the most dramatic of these duplications, affecting ∼10% of human Chromosome 2, enabled a detailed reconstruction of the events leading to the appearance of a novel gene family. The eight members of this family originated from the highly conserved nucleoporin RanBP2 by several genetic rearrangements such as segmental duplications, inversions, translocations, exon loss, and domain accretion. We have experimentally verified that at least one of the newly formed proteins has a cellular localization different from RanBP2's, and we show that positive selection did act on specific domains during evolution.. |
| 119. |
Francesca D Ciccarelli, Christian von Mering, Mikita Suyama, Eoghan D Harrington, Elisa Izaurralde, Peer Bork, Complex genomic rearrangements lead to novel primate gene function., Genome research, 15, 3, 343-51, 2005.03, Orthologous genes that maintain a single-copy status in a broad range of species may indicate a selection against gene duplication. If this is the case, then duplicates of such genes that do survive may have escaped the dosage control by rapid and sizable changes in their function. To test this hypothesis and to develop a strategy for the identification of novel gene functions, we have analyzed 22 primate-specific intrachromosomal duplications of genes with a single-copy ortholog in all other completely sequenced metazoans. When comparing this set to genes not exposed to the single-copy status constraint, we observed a higher tendency of the former to modify their gene structure, often through complex genomic rearrangements. The analysis of the most dramatic of these duplications, affecting approximately 10% of human Chromosome 2, enabled a detailed reconstruction of the events leading to the appearance of a novel gene family. The eight members of this family originated from the highly conserved nucleoporin RanBP2 by several genetic rearrangements such as segmental duplications, inversions, translocations, exon loss, and domain accretion. We have experimentally verified that at least one of the newly formed proteins has a cellular localization different from RanBP2's, and we show that positive selection did act on specific domains during evolution.. |
| 120. |
Mikita Suyama, David Torrents, Peer Bork, BLAST2GENE
A comprehensive conversion of BLAST output into independent genes and gene fragments, Bioinformatics, 10.1093/bioinformatics/bth172, 20, 12, 1968-1970, 2004.08, BLAST2GENE is a program that allows a detailed analysis of genomic regions containing completely or partially duplicated genes. From a BLAST (or BL2SEQ) comparison of a protein or nucleotide query sequence with any genomic region of interest, BLAST2GENE processes all high scoring pairwise alignments (HSPs) and provides the disposition all independent copies along the genomic fragment. The results are provided in text and PostScript formats to allow an automatic and visual evaluation of the respective region.. |
| 121. |
Mikita Suyama, Genome database, Tanpakushitsu kakusan koso. Protein, nucleic acid, enzyme, 49, 11 Suppl, 1841-1846, 2004.08. |
| 122. |
Mikita Suyama, David Torrents, Peer Bork, BLAST2GENE: a comprehensive conversion of BLAST output into independent genes and gene fragments., Bioinformatics (Oxford, England), 20, 12, 1968-70, 2004.08, SUMMARY: BLAST2GENE is a program that allows a detailed analysis of genomic regions containing completely or partially duplicated genes. From a BLAST (or BL2SEQ) comparison of a protein or nucleotide query sequence with any genomic region of interest, BLAST2GENE processes all high scoring pairwise alignments (HSPs) and provides the disposition of all independent copies along the genomic fragment. The results are provided in text and PostScript formats to allow an automatic and visual evaluation of the respective region. AVAILABILITY: The program is available upon request from the authors. A web server of BLAST2GENE is maintained at http://www.bork.embl.de/blast2gene. |
| 123. |
Hillier LW et al., Sequence and comparative analysis of the chicken genome provide unique perspectives on vertebrate evolution, Nature , 432, 695-716, 2004.06. |
| 124. |
Richard A. Gibbs, George M. Weinstock, Michael L. Metzker, Donna M. Muzny, Erica J. Sodergren, Steven Scherer, Graham Scott, David Steffen, Kim C. Worley, Paula E. Burch, Geoffrey Okwuonu, Sandra Hines, Lora Lewis, Christine Deramo, Oliver Delgado, Shannon Dugan-Rocha, George Miner, Margaret Morgan, Alicia Hawes, Rachel Gill, Robert A. Holt, Mark D. Adams, Peter G. Amanatides, Holly Baden-Tillson, Mary Barnstead, Soo Chin, Cheryl A. Evans, Steve Ferriera, Carl Fosler, Anna Glodek, Zhiping Gu, Don Jennings, Cheryl L. Kraft, Trixie Nguyen, Cynthia M. Pfannkoch, Cynthia Sitter, Granger G. Sutton, J. Craig Venter, Trevor Woodage, Douglas Smith, Hong Mei Lee, Erik Gustafson, Patrick Cahill, Arnold Kana, Lynn Doucette-Stamm, Keith Weinstock, Kim Fechtel, Robert B. Weiss, Diane M. Dunn, Eric D. Green, Robert W. Blakesley, Gerard G. Bouffard, Pieter J. de Jong, Kazutoyo Osoegawa, Baoli Zhu, Marco Marra, Jacqueline Schein, Ian Bosdet, Chris Fjell, Steven Jones, Martin Krzywinski, Carrie Mathewson, Asim Siddiqui, Natasja Wye, John McPherson, Shaying Zhao, Claire M. Fraser, Jyoti Shetty, Sofiya Shatsman, Keita Geer, Yixin Chen, Sofyia Abramzon, William C. Nierman, Richard A. Gibbs, George M. Weinstock, Paul H. Havlak, Rui Chen, K. James Durbin, Amy Egan, Yanru Ren, Xing Zhi Song, Bingshan Li, Yue Liu, Xiang Qin, Simon Cawley, George M. Weinstock, Kim C. Worley, A. J. Cooney, Richard A. Gibbs, Lisa M. D'Souza, Kirt Martin, Jia Qian Wu, Manuel L. Gonzalez-Garay, Andrew R. Jackson, Kenneth J. Kalafus, Michael P. McLeod, Aleksandar Milosavljevic, Davinder Virk, Andrei Volkov, David A. Wheeler, Zhengdong Zhang, Jeffrey A. Bailey, Evan E. Eichler, Eray Tuzun, Ewan Birney, Emmanuel Mongin, Abel Ureta-Vidal, Cara Woodwark, Evgeny Zdobnov, Peer Bork, Mikita Suyama, David Torrents, Marina Alexandersson, Barbara J. Trask, Janet M. Young, Douglas Smith, Hui Huang, Kim Fechtel, Huajun Wang, Heming Xing, Keith Weinstock, Sue Daniels, Darryl Gietzen, Jeanette Schmidt, Kristian Stevens, Ursula Vitt, Jim Wingrove, Francisco Camara, M. Mar Albà, Josep F. Abril, Roderic Guigo, Arian Smit, Inna Dubchak, Edward M. Rubin, Olivier Couronne, Alexander Poliakov, Norbert Hübner, Detlev Ganten, Claudia Goesele, Oliver Hummel, Thomas Kreitler, Young Ae Lee, Jan Monti, Herbert Schulz, Heike Zimdahl, Heinz Himmelbauer, Hans Lehrach, Howard J. Jacob, Susan Bromberg, Jo Gullings-Handley, Michael I. Jensen-Seaman, Anne E. Kwitek, Jozef Lazar, Dean Pasko, Peter J. Tonellato, Simon Twigger, Chris P. Ponting, Jose M. Duarte, Stephen Rice, Leo Goodstadt, Scott A. Beatson, Richard D. Emes, Eitan E. Winter, Caleb Webber, Petra Brandt, Gerald Nyakatura, Margaret Adetobi, Francesca Chiaromonte, Laura Elnitski, Pallavi Eswara, Ross C. Hardison, Minmei Hou, Diana Kolbe, Kateryna Makova, Webb Miller, Anton Nekrutenko, Cathy Riemer, Scott Schwartz, James Taylor, Shan Yang, Yi Zhang, Klaus Lindpaintner, T. Dan Andrews, Mario Caccamo, Michele Clamp, Laura Clarke, Valerie Curwen, Richard Durbin, Eduardo Eyras, Stephen M. Searle, Gregory M. Cooper, Serafim Batzoglou, Michael Brudno, Arend Sidow, Eric A. Stone, J. Craig Venter, Bret A. Payseur, Guillaume Bourque, Carlos López-Otín, Xose S. Puente, Kushal Chakrabarti, Sourav Chatterji, Colin Dewey, Lior Pachter, Nicolas Bray, Von Bing Yap, Anat Caspi, San Diego Glenn Tesler, Pavel A. Pevzner, Santa Cruz David Haussler, Krishna M. Roskin, Robert Baertsch, Hiram Clawson, Terrence S. Furey, Angie S. Hinrichs, Donna Karolchik, William J. Kent, Kate R. Rosenbloom, Heather Trumbower, Matt Weirauch, David N. Cooper, Peter D. Stenson, Bin Ma, Michael Brent, Manimozhiyan Arumugam, David Shteynberg, Richard R. Copley, Martin S. Taylor, Harold Riethman, Uma Mudunuri, Jane Peterson, Mark Guyer, Adam Felsenfeld, Susan Old, Stephen Mockrin, Francis Collins, Genome sequence of the Brown Norway rat yields insights into mammalian evolution, Nature, 10.1038/nature02426, 428, 6982, 493-520, 2004.04, The laboratory rat (Rattus norvegicus) is an indispensable tool in experimental medicine and drug development, having made inestimable contributions to human health. We report here the genome sequence of the Brown Norway (BN) rat strain. The sequence represents a high-quality 'draft' covering over 90% of the genome. The BN rat sequence is the third complete mammalian genome to be deciphered, and three-way comparisons with the human and mouse genomes resolve details of mammalian evolution. This first comprehensive analysis includes genes and proteins and their relation to human disease, repeated sequences, comparative genome-wide studies of mammalian orthologous chromosomal regions and rearrangement breakpoints, reconstruction of ancestral karyotypes and the events leading to existing species, rates of variation, and lineage-specific and lineage-independent evolutionary events such as expansion of gene families, orthology relations and protein evolution.. |
| 125. |
Richard A Gibbs, George M Weinstock, Michael L Metzker, Donna M Muzny, Erica J Sodergren, Steven Scherer, Graham Scott, David Steffen, Kim C Worley, Paula E Burch, Geoffrey Okwuonu, Sandra Hines, Lora Lewis, Christine DeRamo, Oliver Delgado, Shannon Dugan-Rocha, George Miner, Margaret Morgan, Alicia Hawes, Rachel Gill, Celera, Robert A Holt, Mark D Adams, Peter G Amanatides, Holly Baden-Tillson, Mary Barnstead, Soo Chin, Cheryl A Evans, Steve Ferriera, Carl Fosler, Anna Glodek, Zhiping Gu, Don Jennings, Cheryl L Kraft, Trixie Nguyen, Cynthia M Pfannkoch, Cynthia Sitter, Granger G Sutton, J Craig Venter, Trevor Woodage, Douglas Smith, Hong-Mei Lee, Erik Gustafson, Patrick Cahill, Arnold Kana, Lynn Doucette-Stamm, Keith Weinstock, Kim Fechtel, Robert B Weiss, Diane M Dunn, Eric D Green, Robert W Blakesley, Gerard G Bouffard, Pieter J De Jong, Kazutoyo Osoegawa, Baoli Zhu, Marco Marra, Jacqueline Schein, Ian Bosdet, Chris Fjell, Steven Jones, Martin Krzywinski, Carrie Mathewson, Asim Siddiqui, Natasja Wye, John McPherson, Shaying Zhao, Claire M Fraser, Jyoti Shetty, Sofiya Shatsman, Keita Geer, Yixin Chen, Sofyia Abramzon, William C Nierman, Paul H Havlak, Rui Chen, K James Durbin, Amy Egan, Yanru Ren, Xing-Zhi Song, Bingshan Li, Yue Liu, Xiang Qin, Simon Cawley, Kim C Worley, A J Cooney, Lisa M D'Souza, Kirt Martin, Jia Qian Wu, Manuel L Gonzalez-Garay, Andrew R Jackson, Kenneth J Kalafus, Michael P McLeod, Aleksandar Milosavljevic, Davinder Virk, Andrei Volkov, David A Wheeler, Zhengdong Zhang, Jeffrey A Bailey, Evan E Eichler, Eray Tuzun, Ewan Birney, Emmanuel Mongin, Abel Ureta-Vidal, Cara Woodwark, Evgeny Zdobnov, Peer Bork, Mikita Suyama, David Torrents, Marina Alexandersson, Barbara J Trask, Janet M Young, Hui Huang, Huajun Wang, Heming Xing, Sue Daniels, Darryl Gietzen, Jeanette Schmidt, Kristian Stevens, Ursula Vitt, Jim Wingrove, Francisco Camara, M Mar Albà, Josep F Abril, Roderic Guigo, Arian Smit, Inna Dubchak, Edward M Rubin, Olivier Couronne, Alexander Poliakov, Norbert Hübner, Detlev Ganten, Claudia Goesele, Oliver Hummel, Thomas Kreitler, Young-Ae Lee, Jan Monti, Herbert Schulz, Heike Zimdahl, Heinz Himmelbauer, Hans Lehrach, Howard J Jacob, Susan Bromberg, Jo Gullings-Handley, Michael I Jensen-Seaman, Anne E Kwitek, Jozef Lazar, Dean Pasko, Peter J Tonellato, Simon Twigger, Chris P Ponting, Jose M Duarte, Stephen Rice, Leo Goodstadt, Scott A Beatson, Richard D Emes, Eitan E Winter, Caleb Webber, Petra Brandt, Gerald Nyakatura, Margaret Adetobi, Francesca Chiaromonte, Laura Elnitski, Pallavi Eswara, Ross C Hardison, Minmei Hou, Diana Kolbe, Kateryna Makova, Webb Miller, Anton Nekrutenko, Cathy Riemer, Scott Schwartz, James Taylor, Shan Yang, Yi Zhang, Klaus Lindpaintner, T Dan Andrews, Mario Caccamo, Michele Clamp, Laura Clarke, Valerie Curwen, Richard Durbin, Eduardo Eyras, Stephen M Searle, Gregory M Cooper, Serafim Batzoglou, Michael Brudno, Arend Sidow, Eric A Stone, J Craig Venter, Bret A Payseur, Guillaume Bourque, Carlos López-Otín, Xose S Puente, Kushal Chakrabarti, Sourav Chatterji, Colin Dewey, Lior Pachter, Nicolas Bray, Von Bing Yap, Anat Caspi, Glenn Tesler, Pavel A Pevzner, David Haussler, Krishna M Roskin, Robert Baertsch, Hiram Clawson, Terrence S Furey, Angie S Hinrichs, Donna Karolchik, William J Kent, Kate R Rosenbloom, Heather Trumbower, Matt Weirauch, David N Cooper, Peter D Stenson, Bin Ma, Michael Brent, Manimozhiyan Arumugam, David Shteynberg, Richard R Copley, Martin S Taylor, Harold Riethman, Uma Mudunuri, Jane Peterson, Mark Guyer, Adam Felsenfeld, Susan Old, Stephen Mockrin, Francis Collins, Genome sequence of the Brown Norway rat yields insights into mammalian evolution., Nature, 428, 6982, 493-521, 2004.04, The laboratory rat (Rattus norvegicus) is an indispensable tool in experimental medicine and drug development, having made inestimable contributions to human health. We report here the genome sequence of the Brown Norway (BN) rat strain. The sequence represents a high-quality 'draft' covering over 90% of the genome. The BN rat sequence is the third complete mammalian genome to be deciphered, and three-way comparisons with the human and mouse genomes resolve details of mammalian evolution. This first comprehensive analysis includes genes and proteins and their relation to human disease, repeated sequences, comparative genome-wide studies of mammalian orthologous chromosomal regions and rearrangement breakpoints, reconstruction of ancestral karyotypes and the events leading to existing species, rates of variation, and lineage-specific and lineage-independent evolutionary events such as expansion of gene families, orthology relations and protein evolution.. |
| 126. |
David Torrents, Mikita Suyama, Evgeny Zdobnov, Peer Bork, A genome-wide survey of human pseudogenes, Genome Research, 10.1101/gr.1455503, 13, 12, 2559-2567, 2003.12, We screened all intergenic regions in the human genome to identify pseudogenes with a combination of homology searches and a functionality test using the ratio of silent to replacement nucleotide substitutions (KA/KS). We identified 19,724 regions of which 95% ± 3% are estimated to evolve neutrally and thus are likely to encode pseudogenes. Half of these have no detectable truncation in their pseudocoding regions and therefore are not identifiable by methods that require the presence of truncations to prove nonfunctionality. A comparative analysis with the mouse genome showed that 70% of these pseudogenes have a retrotranspositional origin (processed), and the rest arose by segmental duplication (nonprocessed). Although the spread of both types of pseudogenes correlates with chromosome size, nonprocessed pseudogenes appear to be enriched in regions with high gene density. It is likely that the human pseudogenes identified here represent only a small fraction of the total, which probably exceeds the number of genes.. |
| 127. |
David Torrents, Mikita Suyama, Evgeny Zdobnov, Peer Bork, A genome-wide survey of human pseudogenes., Genome research, 13, 12, 2559-67, 2003.12, We screened all intergenic regions in the human genome to identify pseudogenes with a combination of homology searches and a functionality test using the ratio of silent to replacement nucleotide substitutions (KA/KS). We identified 19,724 regions of which 95% +/- 3% are estimated to evolve neutrally and thus are likely to encode pseudogenes. Half of these have no detectable truncation in their pseudocoding regions and therefore are not identifiable by methods that require the presence of truncations to prove nonfunctionality. A comparative analysis with the mouse genome showed that 70% of these pseudogenes have a retrotranspositional origin (processed), and the rest arose by segmental duplication (nonprocessed). Although the spread of both types of pseudogenes correlates with chromosome size, nonprocessed pseudogenes appear to be enriched in regions with high gene density. It is likely that the human pseudogenes identified here represent only a small fraction of the total, which probably exceeds the number of genes.. |
| 128. |
La Deana W. Hillier, Robert S. Fulton, Lucinda A. Fulton, Tina A. Graves, Kymberlie H. Pepin, Caryn Wagner-McPherson, Dan Layman, Jason Maas, Sara Jaeger, Rebecca Walker, Kristine Wylie, Mandeep Sekhon, Michael C. Becker, Michelle D. O'Laughlin, Mark E. Schaller, Ginger A. Fewell, Kimberly D. Delehaunty, Tracie L. Miner, William E. Nash, Matt Cordes, Hui Du, Hui Sun, Jennifer Edwards, Holland Bradshaw-Cordum, Johar Ali, Stephanie Andrews, Amber Isak, Andrew Van Brunt, Christine Nguyen, Feiyu Du, Betty Lamar, Laura Courtney, Joelle Kalicki, Philip Ozersky, Lauren Bielicki, Kelsi Scott, Andrea Holmes, Richard Harkins, Anthony Harris, Cynthia Madsen Strong, Shunfang Hou, Chad Tomlinson, Sara Dauphin-Kohlberg, Amy Kozlowicz-Reilly, Shawn Leonard, Theresa Rohlfing, Susan M. Rock, Aye Mon Tin-Wollam, Amanda Abbott, Patrick Minx, Rachel Maupin, Catrina Strowmatt, Phil Latreille, Nancy Miller, Doug Johnson, Jennifer Murray, Jeffrey P. Woessner, Michael C. Wendl, Shiaw Pyng Yang, Brian R. Schultz, John W. Wallis, John Spieth, Tamberlyn A. Bieri, Joanne O. Nelson, Nicolas Berkowicz, Patricia E. Wohldmann, Lisa L. Cook, Matthew T. Hickenbotham, James Eldred, Donald Williams, Joseph A. Bedell, Elaine R. Mardis, Sandra W. Clifton, Stephanie L. Chissoe, Marco A. Marra, Christopher Raymond, Eric Haugen, Will Gillett, Yang Zhou, Rose James, Karen Phelps, Shawn Iadanoto, Kerry Bubb, Elizabeth Simms, Ruth Levy, James Clendenning, Rajinder Kaul, W. James Kent, Terrence S. Furey, Robert A. Baertsch, Michael R. Brent, Evan Keibler, Paul Flicek, Peer Borkk, Mikita Suyamak, Jeffrey A. Bailey, Matthew E. Portnoy, David Torrentsk, Asif T. Chinwalla, Warren R. Gish, Sean R. Eddy, John D. McPherson, Maynard V. Olson, Evan E. Eichler, Eric D. Green, Robert H. Waterston, Richard K. Wilson, The DNA sequence of human chromosome 7, Nature, 10.1038/nature01782, 424, 6945, 157-164, 2003.07, Human chromosome 7 has historically received prominent attention in the human genetics community, primarily related to the search for the cystic fibrosis gene and the frequent cytogenetic changes associated with various forms of cancer. Here we present more than 153 million base pairs representing 99.4% of the euchromatic sequence of chromosome 7, the first metacentric chromosome completed so far. The sequence has excellent concordance with previously established physical and genetic maps, and it exhibits an unusual amount of segmentally duplicated sequence (8.2%), with marked differences between the two arms. Our initial analyses have identified 1,150 protein-coding genes, 605 of which have been confirmed by complementary DNA sequences, and an additional 941 pseudogenes. Of genes confirmed by transcript sequences, some are polymorphic for mutations that disrupt the reading frame.. |
| 129. |
Ladeana W Hillier, Robert S Fulton, Lucinda A Fulton, Tina A Graves, Kymberlie H Pepin, Caryn Wagner-McPherson, Dan Layman, Jason Maas, Sara Jaeger, Rebecca Walker, Kristine Wylie, Mandeep Sekhon, Michael C Becker, Michelle D O'Laughlin, Mark E Schaller, Ginger A Fewell, Kimberly D Delehaunty, Tracie L Miner, William E Nash, Matt Cordes, Hui Du, Hui Sun, Jennifer Edwards, Holland Bradshaw-Cordum, Johar Ali, Stephanie Andrews, Amber Isak, Andrew Vanbrunt, Christine Nguyen, Feiyu Du, Betty Lamar, Laura Courtney, Joelle Kalicki, Philip Ozersky, Lauren Bielicki, Kelsi Scott, Andrea Holmes, Richard Harkins, Anthony Harris, Cynthia Madsen Strong, Shunfang Hou, Chad Tomlinson, Sara Dauphin-Kohlberg, Amy Kozlowicz-Reilly, Shawn Leonard, Theresa Rohlfing, Susan M Rock, Aye-Mon Tin-Wollam, Amanda Abbott, Patrick Minx, Rachel Maupin, Catrina Strowmatt, Phil Latreille, Nancy Miller, Doug Johnson, Jennifer Murray, Jeffrey P Woessner, Michael C Wendl, Shiaw-Pyng Yang, Brian R Schultz, John W Wallis, John Spieth, Tamberlyn A Bieri, Joanne O Nelson, Nicolas Berkowicz, Patricia E Wohldmann, Lisa L Cook, Matthew T Hickenbotham, James Eldred, Donald Williams, Joseph A Bedell, Elaine R Mardis, Sandra W Clifton, Stephanie L Chissoe, Marco A Marra, Christopher Raymond, Eric Haugen, Will Gillett, Yang Zhou, Rose James, Karen Phelps, Shawn Iadanoto, Kerry Bubb, Elizabeth Simms, Ruth Levy, James Clendenning, Rajinder Kaul, W James Kent, Terrence S Furey, Robert A Baertsch, Michael R Brent, Evan Keibler, Paul Flicek, Peer Bork, Mikita Suyama, Jeffrey A Bailey, Matthew E Portnoy, David Torrents, Asif T Chinwalla, Warren R Gish, Sean R Eddy, John D McPherson, Maynard V Olson, Evan E Eichler, Eric D Green, Robert H Waterston, Richard K Wilson, The DNA sequence of human chromosome 7., Nature, 424, 6945, 157-64, 2003.07, Human chromosome 7 has historically received prominent attention in the human genetics community, primarily related to the search for the cystic fibrosis gene and the frequent cytogenetic changes associated with various forms of cancer. Here we present more than 153 million base pairs representing 99.4% of the euchromatic sequence of chromosome 7, the first metacentric chromosome completed so far. The sequence has excellent concordance with previously established physical and genetic maps, and it exhibits an unusual amount of segmentally duplicated sequence (8.2%), with marked differences between the two arms. Our initial analyses have identified 1,150 protein-coding genes, 605 of which have been confirmed by complementary DNA sequences, and an additional 941 pseudogenes. Of genes confirmed by transcript sequences, some are polymorphic for mutations that disrupt the reading frame.. |
| 130. |
Mikita Suyama, Osamu Ohara, DomCut
Prediction of inter-domain linker regions in amino acid sequences, Bioinformatics, 10.1093/bioinformatics/btg031, 19, 5, 673-674, 2003.03, Summary: DomCut is a program to predict inter-domain linker regions solely by amino acid sequence information. The prediction is made by using linker index deduced from a data set of domain/linker segments. The linker preference profile, which is the averaged linker index along a sequence, can be visualized in the graphical interface.. |
| 131. |
Mikita Suyama, Osamu Ohara, DomCut: prediction of inter-domain linker regions in amino acid sequences., Bioinformatics (Oxford, England), 19, 5, 673-4, 2003.03, DomCut is a program to predict inter-domain linker regions solely by amino acid sequence information. The prediction is made by using linker index deduced from a data set of domain/linker segments. The linker preference profile, which is the averaged linker index along a sequence, can be visualized in the graphical interface.. |
| 132. |
R. H. Waterston, L. W. Hillier, L. A. Fulton, R. S. Fulton, T. A. Graves, K. H. Pepin, P. Bork, M. Suyama, D. Torrents, A. T. Chinwalla, E. R. Mardis, J. D. McPherson, R. K. Wilson, The human genome
Genes, pseudogenes, and variation on chromosome 7, Cold Spring Harbor Symposia on Quantitative Biology, 10.1101/sqb.2003.68.13, 68, 13-22, 2003.01. |
| 133. |
Robert H. Waterston, Kerstin Lindblad-Toh, Ewan Birney, Jane Rogers, Josep F. Abril, Pankaj Agarwal, Richa Agarwala, Rachel Ainscough, Marina Alexandersson, Peter An, Stylianos E. Antonarakis, John Attwood, Robert Baertsch, Jonathon Bailey, Karen Barlow, Stephan Beck, Eric Berry, Bruce Birren, Toby Bloom, Peer Bork, Marc Botcherby, Nicolas Bray, Michael R. Brent, Daniel G. Brown, Stephen D. Brown, Carol Bult, John Burton, Jonathan Butler, Robert D. Campbell, Piero Carninci, Simon Cawley, Francesca Chiaromonte, Asif T. Chinwalla, Deanna M. Church, Michele Clamp, Christopher Clee, Francis S. Collins, Lisa L. Cook, Richard R. Copley, Alan Coulson, Olivier Couronne, James Cuff, Val Curwen, Tim Cutts, Mark Daly, Robert David, Joy Davies, Kimberly D. Delehaunty, Justin Deri, Emmanouil T. Dermitzakis, Colin Dewey, Nicholas J. Dickens, Mark Diekhans, Sheila Dodge, Inna Dubchak, Diane M. Dunn, Sean R. Eddy, Laura Elnitski, Richard D. Emes, Pallavi Eswara, Eduardo Eyras, Adam Felsenfeld, Ginger A. Fewell, Paul Flicek, Karen Foley, Wayne N. Frankel, Lucinda A. Fulton, Robert S. Fulton, Terrence S. Furey, Diane Gage, Richard A. Gibbs, Gustavo Glusman, Sante Gnerre, Nick Goldman, Leo Goodstadt, Darren Grafham, Tina A. Graves, Eric D. Green, Simon Gregory, Roderic Guigó, Mark Guyer, Ross C. Hardison, David Haussler, Yoshihide Hayashizaki, Deana W. LaHillier, Angela Hinrichs, Wratko Hlavina, Timothy Holzer, Fan Hsu, Axin Hua, Tim Hubbard, Adrienne Hunt, Ian Jackson, David B. Jaffe, L. Steven Johnson, Matthew Jones, Thomas A. Jones, Ann Joy, Michael Kamal, Elinor K. Karlsson, Donna Karolchik, Arkadiusz Kasprzyk, Jun Kawai, Evan Keibler, Cristyn Kells, W. James Kent, Andrew Kirby, Diana L. Kolbe, Ian Korf, Raju S. Kucherlapati, Edward J. Kulbokas, David Kulp, Tom Landers, J. P. Leger, Steven Leonard, Ivica Letunic, Rosie Levine, Jia Li, Ming Li, Christine Lloyd, Susan Lucas, Bin Ma, Donna R. Maglott, Elaine R. Mardis, Lucy Matthews, Evan Mauceli, John H. Mayer, Megan McCarthy, W. Richard McCombie, Stuart McLaren, Kirsten McLay, John D. McPherson, Jim Meldrim, Beverley Meredith, Jill P. Mesirov, Webb Miller, Tracie L. Miner, Emmanuel Mongin, Kate T. Montgomery, Michael Morgan, Richard Mott, James C. Mullikin, Donna M. Muzny, William E. Nash, Joanne O. Nelson, Michael N. Nhan, Robert Nicol, Zemin Ning, Chad Nusbaum, Michael J. O'Connor, Yasushi Okazaki, Karen Oliver, Emma Overton-Larty, Lior Pachter, Genís Parra, Kymberlie H. Pepin, Jane Peterson, Pavel Pevzner, Robert Plumb, Craig S. Pohl, Alex Poliakov, Tracy C. Ponce, Chris P. Ponting, Simon Potter, Michael Quail, Alexandre Reymond, Bruce A. Roe, Krishna M. Roskin, Edward M. Rubin, Alistair G. Rust, Ralph Santos, Victor Sapojnikov, Brian Schultz, Jorg Schultz, Matthias S. Schwartz, Scott Schwartz, Carol Scott, Steven Seaman, Steve Searle, Ted Sharpe, Andrew Sheridan, Ratna Shownkeen, Sarah Sims, Jonathan B. Singer, Guy Slater, Arian Smit, Douglas R. Smith, Brian Spencer, Arne Stabenau, Nicole Stange-Thomann, Charles Sugnet, Mikita Suyama, Glenn Tesler, Johanna Thompson, David Torrents, Evanne Trevaskis, John Tromp, Catherine Ucla, Abel Ureta-Vidal, Jade P. Vinson, Andrew C. von Niederhausern, Claire M. Wade, Melanie Wall, Ryan J. Weber, Robert B. Weiss, Michael C. Wendl, Anthony P. West, Kris Wetterstrand, Raymond Wheeler, Simon Whelan, Jamey Wierzbowski, David Willey, Sophie Williams, Richard K. Wilson, Eitan Winter, Kim C. Worley, Dudley Wyman, Shan Yang, Shiaw Pyng Yang, Evgeny M. Zdobnov, Michael C. Zody, Eric S. Lander, Initial sequencing and comparative analysis of the mouse genome, Nature, 10.1038/nature01262, 420, 6915, 520-562, 2002.12, The sequence of the mouse genome is a key informational tool for understanding the contents of the human genome and a key experimental tool for biomedical research. Here, we report the results of an international collaboration to produce a high-quality draft sequence of the mouse genome. We also present an initial comparative analysis of the mouse and human genomes, describing some of the insights that can be gleaned from the two sequences. We discuss topics including the analysis of the evolutionary forces shaping the size, structure and sequence of the genomes; the conservation of large-scale synteny across most of the genomes; the much lower extent of sequence orthology covering less than half of the genomes; the proportions of the genomes under selection; the number of protein-coding genes; the expansion of gene families related to reproduction and immunity; the evolution of proteins; and the identification of intraspecies polymorphism.. |
| 134. |
Evgeny M. Zdobnov, Christian Von Mering, Ivica Letunic, David Torrents, Mikita Suyama, Richard R. Copley, George K. Christophides, Dana Thomasova, Robert A. Holt, G. Mani Subramanian, Hans Michael Mueller, George Dimopoulos, John H. Law, Michael A. Wells, Ewan Birney, Rosane Charlab, Aaron L. Halpern, Elena Kokoza, Cheryl L. Kraft, Zhongwu Lai, Suzanna Lewis, Christos Louis, Carolina Barillas-Mury, Deborah Nusskern, Gerald M. Rubin, Steven L. Salzberg, Granger G. Sutton, Pantelis Topalis, Ron Wides, Patrick Wincker, Mark Yandell, Frank H. Collins, Jose Ribeiro, William M. Gelbart, Fotis C. Kafatos, Peer Bork, Comparative genome and proteome analysis of Anopheles gambiae and Drosophila melanogaster, Science, 10.1126/science.1077061, 298, 5591, 149-159, 2002.10, Comparison of the genomes and proteomes of the two diptera Anopheles gambiae and Drosophila melanogaster, which diverged about 250 million years ago, reveals considerable similarities. However, numerous differences are also observed; some of these must reflect the selection and subsequent adaptation associated with different ecologies and life strategies. Almost half of the genes in both genomes are interpreted as orthologs and show an average sequence identity of about 56%, which is slightly lower than that observed between the orthologs of the pufferfish and human (diverged about 450 million years ago). This indicates that these two insects diverged considerably faster than vertebrates. Aligned sequences reveal that orthologous genes have retained only half of their intron/exon structure, indicating that intron gains or losses have occurred at a rate of about one per gene per 125 million years. Chromosomal arms exhibit significant remnants of homology between the two species, although only 34% of the genes colocalize in small "microsyntenic" clusters, and major interarm transfers as well as intra-arm shuffling of gene order are detected.. |
| 135. |
S. Yamamoto-Katayama, A. Sato, M. Ariyoshi, M. Suyama, K. Ishihara, T. Hirano, H. Nakamura, K. Morikawa, H. Jingami, Site-directed removal of N-glycosylation sites in BST-1/CD157
Effects on molecular and functional heterogeneity, Biochemical Journal, 10.1042/0264-6021:3570385, 357, 2, 385-392, 2001.07, Cyclic ADP ribose (cADPR) is a novel second messenger that releases calcium from intracellular calcium stores, but works independently of inositol 1,4,5-trisphosphate. In mammals ADP-ribosyl cyclase function is found in two membrane proteins, CD38 and bone marrow stromal cell antigen 1 (BST-1)/CD157. These enzymes are exposed extracellularly and also possess cADPR hydrolase activity, but an intracellular soluble ADP-ribosyl cyclase has been reported in human T-cells. Previously, a soluble form of BST-1/CD157 (sBST-1), which lacked the glycosylphosphatidylinositol-anchored portion, was expressed by a baculovirus-insect-cell system. In this study, we have purified the sBST-1, and it migrated as two major bands by SDS/PAGE, suggesting that it is post-translationally modified. BST-1 contains four putative N-glycosylation sites. Tunicamycin treatment reduced sBST-1 expression in the culture medium, indicating that N-glycosylation is essential for secretion. Site-directed mutagenesis was performed to generate sBST-1 mutants (N1-N4), each preserving a single N-glycosylation site. N1, N3 and N4 were well secreted into the medium, and were each detected as a single band. Although N3 and N4 retained the ADP-ribosyl cyclase activity, the cADPR-hydrolase activity was retained only in N4. We conclude that N-glycosylation of sBST-1 facilitates the folding of the nascent polypeptide chain into a conformation that is conductive for intracellular transport and enzymic activity. Furthermore a crystal has been obtained using the N4 mutant, but not the wild-type sBST-1. Thus the artificial engineering of N-glycosylation sites could be an effective method to generate homogeneous material for structural studies.. |
| 136. |
Mikita Suyama, Peer Bork, Evolution of prokaryotic gene order
Genome rearrangements in closely related species, Trends in Genetics, 10.1016/S0168-9525(00)02159-4, 17, 1, 10-13, 2001.01, Conservation of gene order in prokaryotes has become important in predicting protein function because, over the evolutionary timescale, genomes are shuffled so that local gene-order conservation reflects the functional constraints within the protein. Here, we compare closely related genomes to identify the rate with which gene order is disrupted and to infer the genes involved in the genome rearrangement.. |
| 137. |
A. Herold, M. Suyama, J. P. Rodrigues, I. C. Braun, U. Kutay, M. Carmo-Fonseca, P. Bork, E. Izaurralde, TAP (NXF1) belongs to a multigene family of putative RNA export factors with a conserved modular architecture, Molecular and cellular biology, 10.1128/MCB.20.23.8996-9008.2000, 20, 23, 8996-9008, 2000.12, Vertebrate TAP (also called NXF1) and its yeast orthologue, Mex67p, have been implicated in the export of mRNAs from the nucleus. The TAP protein includes a noncanonical RNP-type RNA binding domain, four leucine-rich repeats, an NTF2-like domain that allows heterodimerization with p15 (also called NXT1), and a ubiquitin-associated domain that mediates the interaction with nucleoporins. Here we show that TAP belongs to an evolutionarily conserved family of proteins that has more than one member in higher eukaryotes. Not only the overall domain organization but also residues important for p15 and nucleoporin interaction are conserved in most family members. We characterize two of four human TAP homologues and show that one of them, NXF2, binds RNA, localizes to the nuclear envelope, and exhibits RNA export activity. NXF3, which does not bind RNA or localize to the nuclear rim, has no RNA export activity. Database searches revealed that although only one p15 (nxt) gene is present in the Drosophila melanogaster and Caenorhabditis elegans genomes, there is at least one additional p15 homologue (p15-2 [also called NXT2]) encoded by the human genome. Both human p15 homologues bind TAP, NXF2, and NXF3. Together, our results indicate that the TAP-p15 mRNA export pathway has diversified in higher eukaryotes compared to yeast, perhaps reflecting a greater substrate complexity.. |
| 138. |
Thomas Dandekar, Martijn Huynen, Jörg Thomas Regula, Barbara Ueberle, Carl Ulrich Zimmermann, Miguel A. Andrade, Tobias Doerks, Luis Sánchez-Pulido, Berend Snel, Mikita Suyama, Yan P. Yuan, Richard Herrmann, Peer Bork, Re-annotating the Mycoplasma pneumoniae genome sequence
Adding value, function and reading frames, Nucleic acids research, 28, 17, 3278-3288, 2000.09, Four years after the original sequence submission, we have re-annotated the genome of Mycoplasma pneumoniae to incorporate novel data. The total number of ORFss has been increased from 677 to 688 (10 new proteins were predicted in intergenic regions, two further were newly identified by mass spectrometry and one protein ORF was dismissed) and the number of RNAs from 39 to 42 genes. For 19 of the now 35 tRNAs and for six other functional RNAs the exact genome positions were re-annotated and two new tRNA(Leu) and a small 200 nt RNA were identified. Sixteen protein reading frames were extended and eight shortened. For each ORF a consistent annotation vocabulary has been introduced. Annotation reasoning, annotation categories and comparisons to other published data on M.pneumoniae functional assignments are given. Experimental evidence includes 2-dimensional gel electrophoresis in combination with mass spectrometry as well as gene expression data from this study. Compared to the original annotation, we increased the number of proteins with predicted functional features from 349 to 458. The increase includes 36 new predictions and 73 protein assignments confirmed by the published literature. Furthermore, there are 23 reductions and 30 additions with respect to the previous annotation. mRNA expression data support transcription of 184 of the functionally unassigned reading frames.. |
| 139. |
Reiko Kikuno, Takahiro Nagase, Mikita Suyama, Mina Waki, Makoto Hirosawa, Osamu Ohara, HUGE
A database for human large proteins identified in the Kazusa cDNA sequencing project, Nucleic acids research, 28, 1, 331-332, 2000.01, HUGE is a database for human large proteins newly identified in the Kazusa cDNA project, the aim of which is to predict the primary structure of proteins from the sequences of human large cDNAs (> 4 kb). In particular, cDNA clones capable of coding for large proteins (> 50 kDa) are the current targets of the project. HUGE contains > 1100 cDNA sequences and detailed information obtained through analysis of the sequences of cDNAs and the predicted proteins. Besides an increase in the number of cDNA entries, the amount of experimental data for expression profiling has been largely increased and data on chromosomal locations have been newly added. All of the protein-coding regions were examined by GeneMark analysis, and the results of a motif/domain search of each predicted protein sequence against the Pfam database have been newly added. HUGE is available through the WWW at http://www.kazusa.or.jp/huge.. |
| 140. |
Mikita Suyama, Tobias Doerks, Isabelle C. Braun, Michael Sattler, Elisa Izaurralde, Peer Bork, Prediction of structural domains of TAP reveals details of its interaction with p15 and nucleoporins, EMBO Reports, 10.1093/embo-reports/kvd009, 1, 1, 53-58, 2000.01, Vertebrate TAP is a nuclear mRNA export factor homologous to yeast Mex67p. The middle domain of TAP binds directly to p15, a protein related to the nuclear transport factor 2 (NTF2), whereas its C-terminal domain interacts with various nucleoporins, the components of the nuclear pore complex (NPC). Here, we report that the middle domain of TAP is also similar to NTF2, as well as to regions in Ras-GAP SH3 domain binding protein (G3BP) and some plant protein kinases. Based on the known three-dimensional structure of NTF2 homodimer, a heterodimerization model of TAP and p15 could be inferred. This model was confirmed by site-directed mutagenesis of residues located at the dimer interface. Furthermore, the C-terminus of TAP was found to contain a ubiquitin-associated (UBA) domain. By site-directed mutagenesis we show that a conserved loop in this domain plays an essential role in mediating TAP-nucleoporin interaction.. |
| 141. |
Evgeny M Zdobnov, Christian von Mering, Ivica Letunic, David Torrents, Mikita Suyama, Richard R Copley, George K Christophides, Dana Thomasova, Robert A Holt, G Mani Subramanian, Hans-Michael Mueller, George Dimopoulos, John H Law, Michael A Wells, Ewan Birney, Rosane Charlab, Aaron L Halpern, Elena Kokoza, Cheryl L Kraft, Zhongwu Lai, Suzanna Lewis, Christos Louis, Carolina Barillas-Mury, Deborah Nusskern, Gerald M Rubin, Steven L Salzberg, Granger G Sutton, Pantelis Topalis, Ron Wides, Patrick Wincker, Mark Yandell, Frank H Collins, Jose Ribeiro, William M Gelbart, Fotis C Kafatos, Peer Bork, Comparative genome and proteome analysis of Anopheles gambiae and Drosophila melanogaster., Science (New York, N.Y.), 298, 5591, 149-59, 2002.10, Comparison of the genomes and proteomes of the two diptera Anopheles gambiae and Drosophila melanogaster, which diverged about 250 million years ago, reveals considerable similarities. However, numerous differences are also observed; some of these must reflect the selection and subsequent adaptation associated with different ecologies and life strategies. Almost half of the genes in both genomes are interpreted as orthologs and show an average sequence identity of about 56%, which is slightly lower than that observed between the orthologs of the pufferfish and human (diverged about 450 million years ago). This indicates that these two insects diverged considerably faster than vertebrates. Aligned sequences reveal that orthologous genes have retained only half of their intron/exon structure, indicating that intron gains or losses have occurred at a rate of about one per gene per 125 million years. Chromosomal arms exhibit significant remnants of homology between the two species, although only 34% of the genes colocalize in small "microsyntenic" clusters, and major interarm transfers as well as intra-arm shuffling of gene order are detected.. |
| 142. |
Robert H Waterston, Kerstin Lindblad-Toh, Ewan Birney, Jane Rogers, Josep F Abril, Pankaj Agarwal, Richa Agarwala, Rachel Ainscough, Marina Alexandersson, Peter An, Stylianos E Antonarakis, John Attwood, Robert Baertsch, Jonathon Bailey, Karen Barlow, Stephan Beck, Eric Berry, Bruce Birren, Toby Bloom, Peer Bork, Marc Botcherby, Nicolas Bray, Michael R Brent, Daniel G Brown, Stephen D Brown, Carol Bult, John Burton, Jonathan Butler, Robert D Campbell, Piero Carninci, Simon Cawley, Francesca Chiaromonte, Asif T Chinwalla, Deanna M Church, Michele Clamp, Christopher Clee, Francis S Collins, Lisa L Cook, Richard R Copley, Alan Coulson, Olivier Couronne, James Cuff, Val Curwen, Tim Cutts, Mark Daly, Robert David, Joy Davies, Kimberly D Delehaunty, Justin Deri, Emmanouil T Dermitzakis, Colin Dewey, Nicholas J Dickens, Mark Diekhans, Sheila Dodge, Inna Dubchak, Diane M Dunn, Sean R Eddy, Laura Elnitski, Richard D Emes, Pallavi Eswara, Eduardo Eyras, Adam Felsenfeld, Ginger A Fewell, Paul Flicek, Karen Foley, Wayne N Frankel, Lucinda A Fulton, Robert S Fulton, Terrence S Furey, Diane Gage, Richard A Gibbs, Gustavo Glusman, Sante Gnerre, Nick Goldman, Leo Goodstadt, Darren Grafham, Tina A Graves, Eric D Green, Simon Gregory, Roderic Guigó, Mark Guyer, Ross C Hardison, David Haussler, Yoshihide Hayashizaki, LaDeana W Hillier, Angela Hinrichs, Wratko Hlavina, Timothy Holzer, Fan Hsu, Axin Hua, Tim Hubbard, Adrienne Hunt, Ian Jackson, David B Jaffe, L Steven Johnson, Matthew Jones, Thomas A Jones, Ann Joy, Michael Kamal, Elinor K Karlsson, Donna Karolchik, Arkadiusz Kasprzyk, Jun Kawai, Evan Keibler, Cristyn Kells, W James Kent, Andrew Kirby, Diana L Kolbe, Ian Korf, Raju S Kucherlapati, Edward J Kulbokas, David Kulp, Tom Landers, J P Leger, Steven Leonard, Ivica Letunic, Rosie Levine, Jia Li, Ming Li, Christine Lloyd, Susan Lucas, Bin Ma, Donna R Maglott, Elaine R Mardis, Lucy Matthews, Evan Mauceli, John H Mayer, Megan McCarthy, W Richard McCombie, Stuart McLaren, Kirsten McLay, John D McPherson, Jim Meldrim, Beverley Meredith, Jill P Mesirov, Webb Miller, Tracie L Miner, Emmanuel Mongin, Kate T Montgomery, Michael Morgan, Richard Mott, James C Mullikin, Donna M Muzny, William E Nash, Joanne O Nelson, Michael N Nhan, Robert Nicol, Zemin Ning, Chad Nusbaum, Michael J O'Connor, Yasushi Okazaki, Karen Oliver, Emma Overton-Larty, Lior Pachter, Genís Parra, Kymberlie H Pepin, Jane Peterson, Pavel Pevzner, Robert Plumb, Craig S Pohl, Alex Poliakov, Tracy C Ponce, Chris P Ponting, Simon Potter, Michael Quail, Alexandre Reymond, Bruce A Roe, Krishna M Roskin, Edward M Rubin, Alistair G Rust, Ralph Santos, Victor Sapojnikov, Brian Schultz, Jörg Schultz, Matthias S Schwartz, Scott Schwartz, Carol Scott, Steven Seaman, Steve Searle, Ted Sharpe, Andrew Sheridan, Ratna Shownkeen, Sarah Sims, Jonathan B Singer, Guy Slater, Arian Smit, Douglas R Smith, Brian Spencer, Arne Stabenau, Nicole Stange-Thomann, Charles Sugnet, Mikita Suyama, Glenn Tesler, Johanna Thompson, David Torrents, Evanne Trevaskis, John Tromp, Catherine Ucla, Abel Ureta-Vidal, Jade P Vinson, Andrew C Von Niederhausern, Claire M Wade, Melanie Wall, Ryan J Weber, Robert B Weiss, Michael C Wendl, Anthony P West, Kris Wetterstrand, Raymond Wheeler, Simon Whelan, Jamey Wierzbowski, David Willey, Sophie Williams, Richard K Wilson, Eitan Winter, Kim C Worley, Dudley Wyman, Shan Yang, Shiaw-Pyng Yang, Evgeny M Zdobnov, Michael C Zody, Eric S Lander, Initial sequencing and comparative analysis of the mouse genome., Nature, 420, 6915, 520-62, 2002.12, The sequence of the mouse genome is a key informational tool for understanding the contents of the human genome and a key experimental tool for biomedical research. Here, we report the results of an international collaboration to produce a high-quality draft sequence of the mouse genome. We also present an initial comparative analysis of the mouse and human genomes, describing some of the insights that can be gleaned from the two sequences. We discuss topics including the analysis of the evolutionary forces shaping the size, structure and sequence of the genomes; the conservation of large-scale synteny across most of the genomes; the much lower extent of sequence orthology covering less than half of the genomes; the proportions of the genomes under selection; the number of protein-coding genes; the expansion of gene families related to reproduction and immunity; the evolution of proteins; and the identification of intraspecies polymorphism.. |
| 143. |
Mikita Suyama, Takahiro Nagase, Osamu Ohara, HUGE
A database for human large proteins identified by Kazusa cDNA sequencing project, Nucleic acids research, 10.1093/nar/27.1.338, 27, 1, 338-339, 1999.01, HUGE is a database for human large proteins newly identified by Kazusa cDNA project, which aims to predict protein primary structures from sequences of human large cDNAs (> 4 kb). In particular, cDNA clones capable of coding for large proteins (> 50 kDa) are current targets of the project. More than 700 sequences of human cDNAs (average size, 5.1 kb) have been determined to date and deposited in the public databases. Notable information implied from the cDNAs and the predicted protein sequences can be obtained through HUGE via the World Wide Web at URL http://www.kazusa.or.jp/huge.. |
| 144. |
Tonozuka T, Sato K, Sakaguchi M, Suyama M, Sekine K, Sakano Y, Purification and some properties of an extracellular oligo-1,6-glucosidase from Bacillus stearothermophilus SA0301, Journal of Applied Glycoscience, 45, 397-400, 1998.06. |
| 145. |
Takahiro Nagase, Ken Ichi Ishikawa, Mikita Suyama, Reiko Kikuno, Makoto Hirosawa, Nobuyuki Miyajima, Ayako Tanaka, Hirokazu Kotani, Nobuo Nomura, Osamu Ohara, Prediction of the coding sequences of unidentified human genes. XIII. The complete sequences of 100 new cDNA clones from brain which code for large proteins in vitro, DNA Research, 10.1093/dnares/6.1.63, 6, 1, 63-70, 1999.01, As a part of our cDNA project for deducing the coding sequence of unidentified human genes, we newly determined the sequences of 100 cDNA clones from a set of size-fractionated human brain cDNA libraries, and predicted the coding sequences of the corresponding genes, named KIAA0919 to KIAA1018. The sequencing of these clones revealed that the average sizes of the inserts and corresponding open reading frames were 4.9 kb and 2.6 kb (882 amino acid residues), respectively. A computer search of the sequences against the public databases indicated that predicted coding sequences of 87 genes contained sequences similar to known genes, 53% of which (46 genes) were categorized as proteins relating to cell signaling/communication, cell structure/motility and nucleic acid management. The chromosomal locations of the genes were determined by using human-rodent hybrid panels unless their mapping data were already available in the public databases. The expression profiles of all the genes among 10 human tissues, 8 brain regions (amygdala, corpus callosum, cerebellum, caudate nucleus, hippocampus, substania nigra, subthalamic nucleus, and thalamus), spinal cord, fetal brain and fetal liver were also examined by reverse transcription-coupled polymerase chain reaction, products of which were quantified by enzyme-linked immunosorbent assay.. |
| 146. |
Katsuhisa Horimoto, Mikita Suyama, Hiroyuki Toh, Kentaro Mori, Jinya Otsuka, A method for comparing circular genomes from gene locations
Application to mitochondrial genomes, Bioinformatics, 10.1093/bioinformatics/14.9.789, 14, 9, 789-802, 1998.01, Motivation: Data on the entire structures of organelle and bacterial genomes, most of which are known to be circular, have accumulated at a rapid pace. This information enables us to utilize the locations of homologous gene pairs for measuring the dissimilarity between complete genomic structures. Results: A macroscopic distance is presented for comparing circular genomes from their overall structures, on the basis of the locations of two pairs of homologous genes on the compared genomes. The novel aspect of our method is that the comparison between the genomes automatically reveals a relationship based on the information on all gene locations, by incorporating the mobility of each gene, which includes not only the gene order, but also the relative location between gene pairs. The plausibility of the newly defined distances is evaluated by means of 44 mitochondrial genomes. The genome distance shows high performance for quantitatively describing the differences between the gene organizations of the genomes. Availability: Since the programs implementing these calculations require well-arranged gene organization data, they have not been released yet. However, one of the authors will analyse circular genomes upon request. Data on the gene organizations may be submitted electronically to the address below. Contact: horimoto@@@ged.saga-med.ac.jp.. |
| 147. |
Takahiro Nagase, Ken Ichi Ishikawa, Mikita Suyama, Reiko Kikuno, Makoto Hirosawa, Nobuyuki Miyajima, Ayako Tanaka, Hirokazu Kotani, Nobuo Nomura, Osamu Ohara, Prediction of the coding sequences of unidentified human genes. XII. The complete sequences of 100 new cDNA clones from brain which code for large proteins in vitro, DNA Research, 10.1093/dnares/5.6.355, 5, 6, 355-364, 1998.01, In this paper, we report the sequences of 100 cDNA clones newly determined from a set of size-fractionated human brain cDNA libraries and predict the coding sequences of the corresponding genes, named KIAA0819 to KIAA0918. These cDNA clones were selected on the basis of their coding potentials of large proteins (50 kDa and more) by using in vitro transcription/translation assays. The sequence data showed that the average sizes of the inserts and corresponding open reading frames are 4.4 kb and 2.5 kb (831 amino acid residues), respectively. Homology and motif/domain searches against the public databases indicated that the predicted coding sequences of 83 genes were similar to those of known genes, 59% of which (49 genes) were categorized as coding for proteins functionally related to cell signaling/communication, cell structure/motility and nucleic acid management. The chromosomal locations and the expression profiles of all the genes were also examined. For 54 clones including brain-specific ones, the mRNA levels were further examined among 8 brain regions (amygdala, corpus callosum, cerebellum, caudate nucleus, hippocampus, substantia nigra, subthalamic nucleus, and thalamus), spinal cord, and fetal brain.. |
| 148. |
Ken Ichi Ishikawa, Takahiro Nagase, Mikita Suyama, Nobuyuki Miyajima, Ayako Tanaka, Hirokazu Kotani, Nobuo Nomura, Osamu Ohara, Prediction of the coding sequences of unidentified human genes. X. The complete sequences of 100 new cDNA clones from brain which can code for large proteins in vitro, DNA Research, 10.1093/dnares/5.3.169, 5, 3, 169-176, 1998.01, As an extension of our cDNA analysis for deducing the coding sequences of unidentified human genes, we have newly determined the sequences of 100 cDNA clones from a set of size-fractionated human brain cDNA libraries, and predicted the coding sequences of the corresponding genes, named KIAA0611 to KIAA0710. In vitro transcription-coupled translation assay was applied as the first screening to select cDNA clones which produce proteins with apparent molecular mass of 50 kDa and over. One hundred unidentified cDNA clones thus selected were then subjected to sequencing of entire inserts. The average size of the inserts and corresponding open reading frames was 4.9 kb and 2.8 kb (922 amino acid residues), respectively. Computer search of the sequences against the public databases indicated that predicted coding sequences of 87 genes were similar to those of known genes, 62% of which (54 genes) were categorized as proteins related to cell signaling/communication, cell structure/motility and nucleic acid management. The expression profiles in 10 human tissues of all the clones characterized in this study were examined by reverse transcription-coupled polymerase chain reaction and the chromosomal locations of the clones were determined by using human-rodent hybrid panels.. |
| 149. |
Takahiro Nagase, Ken Ichi Ishikawa, Mikita Suyama, Reiko Kikuno, Nobuyuki Miyajima, Ayako Tanaka, Hirokazu Kotani, Nobuo Nomura, Osamu Ohara, Prediction of the coding sequences of unidentified human genes. XI. The complete sequences of 100 new cDNA clones from brain which code for large proteins in vitro, DNA Research, 10.1093/dnares/5.5.277, 5, 5, 277-286, 1998.01, In our series of projects for accumulating sequence information on the coding sequences of unidentified human genes, we have newly determined the sequences of 100 cDNA clones from a set of size-fractionated human brain cDNA libraries, and predicted the coding sequences of the corresponding genes, named KIAA0711 to KIAA0810. These cDNA clones were selected according to their coding potentials of large proteins (50 kDa and more) in vitro. The average sizes of the inserts and corresponding open reading frames were 4.3 kb and 2.6 kb (869 amino acid residues), respectively. Sequence analyses against the public databases indicated that the predicted coding sequences of 78 genes were similar to those of known genes, 64% of which (50 genes) were categorized as proteins functionally related to cell signaling/communication, cell structure/motility and nucleic acid management. As additional information concerning genes characterized in this study, the chromosomal locations of the clones were determined by using human-rodent hybrid panels and the expression profiles among 10 human tissues were examined by reverse transcription-coupled polymerase chain reaction which was substantially improved by enzyme-linked immunosorbent assay.. |
| 150. |
Mikita Suyama, Yo Matsuo, Ken Nishikawa, Comparison of protein structures using 3D profile alignment, Journal of Molecular Evolution, 10.1007/PL00000065, 44, SUPPL. 1, S163-S173, 1997.03, A novel method for protein structure comparison using 3D profile alignment is presented. The 3D profile is a position-dependent scoring matrix derived from three-dimensional structures and is basically used to estimate sequence-structure compatibility for prediction of protein structure. Our idea is to compare two 3D profiles using a dynamic programming algorithm to obtain optimal alignment and a similarity score between them. When the 3D profile of hemoglobin was compared with each of the profiles in the library, which contained 325 profiles of representative structures, all the profiles of other globins were detected with relatively high scores, and proteins in the same structural class followed the globins. Exhaustive comparison of 3D profiles in the library was also performed to depict protein relatedness in the structure space. Using multidimensional scaling, a planar projection of points in the protein structure space revealed an overall grouping in terms of structural classes, i.e., all-α, all-β, α/β, and α+β. These results differ in implication from those obtained by the conventional structure- structure comparison method. Differences are discussed with respect to the structural divergence of proteins in the course of molecular evolution.. |
| 151. |
Mikita Suyama, Takaaki Nishioka, Jun'ichi Oda, Searching for common sequence patterns among distantly related proteins, Protein Engineering, Design and Selection, 10.1093/protein/8.11.1075, 8, 11, 1075-1080, 1995.11, We have developed a program Gap Allowing Pattern Explorer (GAPE) to extract amino acid sequence motifs conserved among distantly related proteins. The GAPE program is designed to allow gaps in the sequences. First, this program generates all possible amino acid patterns comprising up to five amino acids. Sequences containing the amino acid residues in the same order as a generated pattern are selected as subsequences, where the differences in the distances between two consecutive amino acids are ignored. Next, the motifs are extracted from the subsequences under conditions in which all four distances between the five amino acids are fixed. At this stage, motifs with gaps in their subsequence are also found by relaxing one of the four fixed distances. The statistical significance for a motif obtained is calculated based on the amino acid composition of the sequences under consideration. When the GAPE program was applied to 59 pyridoxal-phosphaterelated sequences and 64 ATP (AMP-forming)-related sequences, motifs extracted with a low expectation of occurrence contained some of the amino acid residues chemically proved to be involved in the ligand recognition.. |
| 152. |
Mikita Suyama, Atsushi Ogiwara, Takaaki Nishioka, Jun'ichi Oda, Searching for amino acid sequence motifs among enzymes
The enzyme-reaction database, Bioinformatics, 10.1093/bioinformatics/9.1.9, 9, 1, 9-15, 1993.02, Recently we have constructed a database-the Enzyme-Reaction Database-which links a chemical structure to amino acid sequences of enzymes that recognize the chemical structure as their ligand. The total number of enzymes registered in the database is 1103 with 6668 NBRF-PIR entry codes and 1756 chemical compounds. The chemical structures and chemical names for 842 compounds are registered in the Chemical-Structure Database on the MACCS system. For each enzyme, the sequences were divided into clusters, and multiply aligned in each cluster to extract a conserved sequence. A total of 158 781 five-residue-long fragments were constructed from 433 conserved sequences and compared among different clusters of different enzymes. One of these motifs shared by different enzymes S-G-G-L-D. The motif was conserved in both argininosuccinate synthase (EC 6.3.4.5) and asparagine synthase (glutamine-hydrolysing) (EC 6.3.5.4). This result showed that the database was useful for the analysis of the relationship between chemical structures and amino acid sequence motifs.. |
