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Yoshimichi Nakatsu Last modified date:2021.07.15

Associate Professor / Bioregulation
Department of Basic Medicine
Faculty of Medical Sciences

Graduate School
Undergraduate School

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Department of Medical Biophysics and Radiation Biology, Faculty of Medical Sciences .
Academic Degree
Doctor of Science
Country of degree conferring institution (Overseas)
Field of Specialization
Molecular Biology
Total Priod of education and research career in the foreign country
Research Interests
  • Elucidation of environmental and genetic factors that enhance microsatellite instability in mismatch repair deficient human cells
    keyword : Genomic destabilization, mismatch repair, colorectal cancer
  • Biological effects of non-thermal atmospheric pressure plasma
    keyword : ROS, cell death, mutation, DNA damages, epigenome
  • Mutagenesis and carcinogenesis caused by the failure of the mechanisms for DNA damage response
    keyword : oxidative stress DNA repair mismatch repair cell-death
  • Molecular mechanisms for DNA damage response
    keyword : mutation cell-death adaptive response mismatch repair
  • Molecular mechanisms for prevention and elimination of oxidative DNA damages
    keyword : oxidative DNA damages, nucleotide, mutagenesis, cell death, carcinogenesis, gene-knockout mice, transgenic mice
Current and Past Project
  • Non-thermal atmospheric pressure plasma induces selective death for cancer cells and promotes endoderm formation in embryonic body derived from embryonic stem cells. Plasma-treatment of plant seeds also promotes germination and subsequent growth. Since plasma irradiation causes DNA damage responses, active molecular species generated by plasma act on chromatin, and It is possible that the resulting chromatin-modifications could cause cell death and epigenome changes leading to the promotion of endoderm formation in the embryoid body and the growth promotion of plants. In this study, we examine the possibility that non-thermal atmospheric pressure plasma can generate modified bases in genomic DNA and induce cell death and epigenome changes.
  • The mutation rate and microsatellite sequence stability are analyzed in wild type and mutant human iPS cells, human cancer cell lines, and mouse fetal fibroblasts, cultured under hypoxic conditions.
  • The short tandem repeat (STR) is a difficult-to-reproduce sequence in which errors frequently occur during DNA replication, but a large number of STR are distributed on the eukaryotic genome. The physiological function of STR is unknown so far, but its alterations are associated with some human cancers and neuromuscular diseases. With the aim of elucidating the molecular mechanism of STR changes and to identify the molecular abnormalities that lead to repeat instability, the mutations in DNA mismatch repair gene and DNA polymerase δ gene found in human diseases are introduced into human cells by means of genome editing technique, and the mutation rate, mutation spectrum and changes in microsatellite sequences of those cells are analyzed.
  • Plasma exhibits wound healing and carcinostatic effects, but on the other hand, it may induce DNA damages and cause carcinogenesis and aging. For safety use of plasma medicine, it is necessary to establish safe dosage and usage. The purpose of this study is to analyze the DNA damages and mutations caused by plasma, and to provide a research base for the safety evaluation of plasma medicine.
  • The MUTYH gene is known to be associated with Familial adenomatous polyposis (MAP: MUTYH -associated polyposis), and encodes a DNA repair protein that suppresses mutations caused by oxidative stress in mammalian cells. MUTYH is involved not only in the suppression of mutations but also in the cell death induced by oxidative stress. In this study, I investigate the role of MUTYH associated with mismatch repair proteins in the induction of cell death caused by oxidative stress.
  • We developed an experimental system for oxidative stress-inducing mutagenesis and carcinogenesis in the intestine of mice. In oder to identify the common genes mutated in the adenomas/carcinomas induced by using this system, we analyze the genomes of adenomas/carcinomas induced in the same animals.
  • Mutagenesis caused by oxidized nucleotides and it preventing mechanisms.
    Analysis of cell death pathway evoked by oxidized nucleotides.
  • Oxygen-induced DNA damage and its repair mechanism
Academic Activities
1. Yoshimichi Nakatsu, Mutso Sekiguchi, Oxidative Damage to Nucleotide: Consequences and Preventive Mechanism, Imperial College Press, "Oxidative Stress, Disease and Cancer" Singh, KK (Ed), pp221-252, 2006.01.
1. Tsuzuki T, Nakatsu Y, Nakabeppu Y, Significance of error-avoiding mechanisms for oxidative DNA damage in carcinogenesis, Cancer Science, 2007.08.
1. Yusuke Matsuno, Yuko Atsumi, Md Alauddin, Md Masud Rana, Haruka Fujimori, Mai Hyodo, Atsuhiro Shimizu, Tomoki Ikuta, Hiroko Tani, Hidetaka Torigoe, Yoshimichi Nakatsu, Teruhisa Tsuzuki, Michio Komai, Hitoshi Shirakawa, Ken ichi Yoshioka, Resveratrol and its Related Polyphenols Contribute to the Maintenance of Genome Stability, Scientific reports, 10.1038/s41598-020-62292-5, 10, 1, 2020.12, Genomic destabilisation is associated with the induction of mutations, including those in cancer-driver genes, and subsequent clonal evolution of cells with abrogated defence systems. Such mutations are not induced when genome stability is maintained; however, the mechanisms involved in genome stability maintenance remain elusive. Here, resveratrol (and related polyphenols) is shown to enhance genome stability in mouse embryonic fibroblasts, ultimately protecting the cells against the induction of mutations in the ARF/p53 pathway. Replication stress-associated DNA double-strand breaks (DSBs) that accumulated with genomic destabilisation were effectively reduced by resveratrol treatment. In addition, resveratrol transiently stabilised the expression of histone H2AX, which is involved in DSB repair. Similar effects on the maintenance of genome stability were observed for related polyphenols. Accordingly, we propose that polyphenol consumption can contribute to the suppression of cancers that develop with genomic instability, as well as lifespan extension..
2. Haruka Fujimori, Mai Hyodo, Yusuke Matsuno, Atsuhiro Shimizu, Yusuke Minakawa, Yuko Atsumi, Yoshimichi Nakatsu, Teruhisa Tsuzuki, Yasufumi Murakami, Ken ichi Yoshioka, Mismatch repair dependence of replication stress-associated DSB recognition and repair, Heliyon, 10.1016/j.heliyon.2019.e03057, 5, 12, 2019.12, Most cancers develop with one of two types of genomic instability, namely, chromosomal instability (CIN) or microsatellite instability (MSI). Both are induced by replication stress-associated DNA double-strand breaks (DSBs). The type of genomic instability that arises is dependent on the choice of DNA repair pathway. Specifically, MSI is induced via a PolQ-dependent repair pathway called microhomology-mediated end joining (MMEJ) in a mismatch repair (MMR)-deficient background. However, it is unclear how the MMR status determines the choice of DSB repair pathway. Here, we show that replication stress-associated DSBs initially targeted by the homologous recombination (HR) system were subsequently hijacked by PolQ-dependent MMEJ in MMR-deficient cells, but persisted as HR intermediates in MMR-proficient cells. PolQ interacting with MMR factors was effectively loaded onto damaged chromatin in an MMR-deficient background, in which merged MRE11/γH2AX foci also effectively formed. Thus, the choice of DNA repair pathway according to the MMR status determines whether CIN or MSI is induced. Biological sciences; Cell biology; Genetics; DNA repair; Molecular biology; Genomic instability; DNA repair; Mismatch repair; Microhomology-mediated end joining; DNA polymerase theta.
3. Yusuke Matsuno, Yuko Atsumi, Atsuhiro Shimizu, Kotoe Katayama, Haruka Fujimori, Mai Hyodo, Yusuke Minakawa, Yoshimichi Nakatsu, Syuzo Kaneko, Ryuji Hamamoto, Teppei Shimamura, Satoru Miyano, Teruhisa Tsuzuki, Fumio Hanaoka, Ken ichi Yoshioka, Replication stress triggers microsatellite destabilization and hypermutation leading to clonal expansion in vitro, Nature communications, 10.1038/s41467-019-11760-2, 10, 1, 2019.12, Mismatch repair (MMR)-deficient cancers are characterized by microsatellite instability (MSI) and hypermutation. However, it remains unclear how MSI and hypermutation arise and contribute to cancer development. Here, we show that MSI and hypermutation are triggered by replication stress in an MMR-deficient background, enabling clonal expansion of cells harboring ARF/p53-module mutations and cells that are resistant to the anti-cancer drug camptothecin. While replication stress-associated DNA double-strand breaks (DSBs) caused chromosomal instability (CIN) in an MMR-proficient background, they induced MSI with concomitant suppression of CIN via a PARP-mediated repair pathway in an MMR-deficient background. This was associated with the induction of mutations, including cancer-driver mutations in the ARF/p53 module, via chromosomal deletions and base substitutions. Immortalization of MMR-deficient mouse embryonic fibroblasts (MEFs) in association with ARF/p53-module mutations was ~60-fold more efficient than that of wild-type MEFs. Thus, replication stress-triggered MSI and hypermutation efficiently lead to clonal expansion of cells with abrogated defense systems..
4. Genki Hayashida, Seijiro Shioi, Kyoko Hidaka, Ryosuke Fujikane, Masumi Hidaka, Toshiki Tsurimoto, Teruhisa Tsuzuki, Shinya Oda, Yoshimichi Nakatsu, Differential genomic destabilisation in human cells with pathogenic MSH2 mutations introduced by genome editing, Experimental Cell Research, 10.1016/j.yexcr.2019.02.020, 377, 1-2, 24-35, 2019.04, Repeat destabilisation is variously associated with human disease. In neoplastic diseases, microsatellite instability (MSI) has been regarded as simply reflecting DNA mismatch repair (MMR) deficiency. However, several discrepancies have been pointed out. Firstly, the MSI + phenotype is not uniform in human neoplasms. Established classification utilises the frequency of microsatellite changes, i.e. MSI-H (high) and -L (low), the former regarded as an authentic MMR-defective phenotype. In addition, we have observed the qualitatively distinct modes of MSI, i.e. Type A and Type B. One discrepancy we previously pointed out is that tumours occurring in MMR gene knockout mice exhibited not drastic microsatellite changes typical in MSI-H tumours (i.e. Type B mode) but minor and more subtle alterations (i.e. Type A mode). In the present study, MSH2 mutations reported in Lynch syndrome (LS) kindred have been introduced into HeLa cells using the CRISPR/Cas9 system. The established mutant clones clearly exhibited MMR-defective phenotypes with alkylating agent-tolerance and elevated mutation frequencies. Nevertheless, microsatellites were not markedly destabilised as in MSI-H tumours occurring in LS patients, and all the observed alterations were uniformly Type A, which confirms the results in mice. Our findings suggest added complexities to the molecular mechanisms underlying repeat destabilisation in human genome..
5. Matsubara Y, Matsumoto T, Yoshiya K,Yoshida A,Ikeda S, Furuyama T, Nakatsu Y, Tsuzuki T,Nomura M, Maehara Y., Budding Uninhibited by Benzimidazole-1 Insufficiency Prevents Acute Renal Failure in Severe Sepsis by Maintaining Anticoagulant Functions of Vascular Endothelial Cells., SHOCK, 10.1097/SHK.0000000000001147, 51, 3, 364-371, 2019.03.
6. Tetsuya Suzuki, Yuri Yanai, Natsuki Nishigaki, Yoshimichi Nakatsu, Teruhisa Tsuzuki, Hiroyuki Kamiya, Effects of mismatches distant from the target position on gene correction with a 5′-tailed duplex, Journal of Bioscience and Bioengineering, 10.1016/j.jbiosc.2017.12.017, 125, 5, 619-623, 2018.05, The introduction of a 5′-tailed duplex (5′-TD) fragment into cells corrects a base-substitution mutation in a target DNA. We previously reported that the gene correction efficiency was improved when a frameshift type of second mismatch was present ∼330 bases distant from the target position, between the target DNA and the 5′-TD fragment. In this study, the effects of the second mismatches on the gene correction were further examined. Base–base mismatches 332 bases distant from the target position slightly enhanced gene correction, but less efficiently than the previously studied frameshift mismatches. The gene correction efficiency was also increased when the distance between the target position and the second frameshift mismatch was changed to ∼270 bases. These results suggested that the introduction of an appropriate second frameshift mismatch into the 5′-TD fragment improves the gene correction efficiency..
7. Issei Egashira, Fumi Takahashi-Yanaga, Risa Nishida, masaki arioka, Kazunobu Igawa, Katsuhiko Tomooka, Yoshimichi Nakatsu, Teruhisa Tsuzuki, Yusaku Nakabeppu, Takanari Kitazono, Toshiyuki Sasaguri, Celecoxib and 2,5-dimethylcelecoxib inhibit intestinal cancer growth by suppressing the Wnt/β-catenin signaling pathway, Cancer Science, 10.1111/cas.13106, 108, 1, 108-115, 2017.01, We previously reported that celecoxib, a selective COX-2 inhibitor, strongly inhibited human colon cancer cell proliferation by suppressing the Wnt/β-catenin signaling pathway. 2,5-Dimethylcelecoxib (DM-celecoxib), a celecoxib analog that does not inhibit COX-2, has also been reported to have an antitumor effect. In the present study, we elucidated whether DM-celecoxib inhibits intestinal cancer growth, and its underlying mechanism of action. First, we compared the effect of DM-celecoxib with that of celecoxib on the human colon cancer cell lines HCT-116 and DLD-1. 2,5-Dimethylcelecoxib suppressed cell proliferation and inhibited T-cell factor 7-like 2 expression with almost the same strength as celecoxib. 2,5-Dimethylcelecoxib also inhibited the T-cell factor-dependent transcription activity and suppressed the expression of Wnt/β-catenin target gene products cyclin D1 and survivin. Subsequently, we compared the in vivo effects of celecoxib and DM-celecoxib using the Mutyh−/− mouse model, in which oxidative stress induces multiple intestinal carcinomas. Serum concentrations of orally administered celecoxib and DM-celecoxib elevated to the levels enough to suppress cancer cell proliferation. Repeated treatment with celecoxib and DM-celecoxib markedly reduced the number and size of the carcinomas without showing toxicity. These results suggest that the central mechanism for the anticancer effect of celecoxib derivatives is the suppression of the Wnt/β-catenin signaling pathway but not the inhibition of COX-2, and that DM-celecoxib might be a better lead compound candidate than celecoxib for the development of novel anticancer drugs..
8. Daisuke Matsuda, Takuya Matsumoto, Kenichi Honma, Ayae Ikawa-Yoshida, Mitsuho Onimaru, Tadashi Furuyama, Yoshimichi Nakatsu, Teruhisa Tsuzuki, Yoshihiko Maehara, BUBR1 insufficiency in mice increases their sensitivity to oxidative stress, In Vivo, 10.21873/invivo.10993, 30, 6, 769-776, 2016.11, Background/Aim: Budding uninhibited by benzimidazole-related 1 (BUBR1) plays an important role in the spindle assembly checkpoint to prevent chromosome missegregation and aneuploidy during mitosis. We previously generated mutant mice that express BUBR1 at only 20% of the normal level (BubR1L/L mice). Here, we examined the effect of low BUBR1 expression on oxidative stress-induced carcinogenesis in mice. Materials and Methods: We orally administered either a potassium bromate (KBrO3) solution (2 g/l) or tap water to BubR1L/L and wild-type (BubR1+/+) mice for 16 weeks and examined the subsequent incidence of tumours. Results: KBrO3-treated BubR1L/L mice showed significantly higher mortality than the KBrO3-treated BubR1+/+ and control tap water-treated mice (p=0.0082). Histopathological and immunohistochemical analyses revealed that the spleens of surviving BubR1L/L mice were occupied by non-B-, non-T-cells with high proliferative potential. Conclusion: Our results indicate that low BUBR1 expression increases oxidative stress-induced mortality in mice, possibly caused by splenic neoplasms..
9. Ayae Ikawa-Yoshida, Takuya Matsumoto, Shinji Okano, Yukihiko Aoyagi, Yutaka Matsubara, Tadashi Furuyama, Yoshimichi Nakatsu, Teruhisa Tsuzuki, Mitsuho Onimaru, Tomoko Ohkusa, Masatoshi Nomura, Yoshihiko Maehara, BubR1 Insufficiency Impairs Liver Regeneration in Aged Mice after Hepatectomy through Intercalated Disc Abnormality, Scientific Reports, 10.1038/srep32399, 6, 2016.08, A delay in liver regeneration after partial hepatectomy (PHx) leads to acute liver injury, and such delays are frequently observed in aged patients. BubR1 (budding uninhibited by benzimidazole-related 1) controls chromosome mitotic segregation through the spindle assembly checkpoint, and BubR1 down-regulation promotes aging-associated phenotypes. In this study we investigated the effects of BubR1 insufficiency on liver regeneration in mice. Low-BubR1-expressing mutant (BubR1L/L) mice had a delayed recovery of the liver weight-to-body weight ratio and increased liver deviation enzyme levels after PHx. Microscopic observation of BubR1L/L mouse liver showed an increased number of necrotic hepatocytes and intercalated disc anomalies, resulting in widened inter-hepatocyte and perisinusoidal spaces, smaller hepatocytes and early-stage microvilli atrophy. Up-regulation of desmocollin-1 (DSC1) was observed in wild-type, but not BubR1L/L, mice after PHx. In addition, knockdown of BubR1 expression caused down-regulation of DSC1 in a human keratinocyte cell line. BubR1 insufficiency results in the impaired liver regeneration through weakened microstructural adaptation against PHx, enhanced transient liver failure and delayed hepatocyte proliferation. Thus, our data suggest that a reduction in BubR1 levels causes failure of liver regeneration through the DSC1 abnormality..
10. Hiroyuki Kamiya, Natsuki Nishigaki, Akihiro Ikeda, Seiya Yukawa, Yukiko Morita, Yoshimichi Nakatsu, Teruhisa Tsuzuki, Hideyoshi Harashima, Insertion and deletion mismatches distant from the target position improve gene correction with a tailed duplex, Nucleosides, Nucleotides and Nucleic Acids, 10.1080/15257770.2016.1163384, 35, 7, 379-388, 2016.07, A 5′-tailed duplex (TD) DNA corrects a base-substitution mutation. In this study, the effects of insertion and deletion (indel) mismatches distant from the target position on the gene correction were examined. Three target plasmid DNAs with and without indel mismatches ∼330 bases distant from the correction target position were prepared, and introduced into HeLa cells together with the TD. The indel mismatches improved the gene correction efficiency and specificity without sequence conversions at the indel mismatch site. These results suggested that the gene correction efficiency and specificity are increased when an appropriate second mismatch is introduced into the TD fragment..
11. Hiroyuki Yamamoto, Masataka Ishimura, Masayuki Ochiai, Hidetoshi Takada, K. Kusuhara, Yoshimichi Nakatsu, Teruhisa Tsuzuki, K. Mitani, Toshiro Hara, BTK gene targeting by homologous recombination using a helper-dependent adenovirus/adeno-associated virus hybrid vector, 10.1038/gt.2015.91, 23, 2, 205-213, 2015.08.
12. Rie Kanao, Masayuki Yokoi, Tsuyoshi Ohkumo, Yasutaka Sakurai, Kantaro Dotsu, Shinobu Kura, Yoshimichi Nakatsu, Teruhisa Tsuzuki, Chikahide Masutani, Fumio Hanaoka, UV-induced mutations in epidermal cells of mice defective in DNA polymerase η and/or ι, DNA Repair, 10.1016/j.dnarep.2015.02.006, 29, 139-146, 2015.05, Xeroderma pigmentosum variant (XP-V) is a human rare inherited recessive disease, predisposed to sunlight-induced skin cancer, which is caused by deficiency in DNA polymerase η (Polη). Polη catalyzes accurate translesion synthesis (TLS) past pyrimidine dimers, the most prominent UV-induced lesions. DNA polymerase ι (Polι) is a paralog of Polη that has been suggested to participate in TLS past UV-induced lesions, but its function in vivo remains uncertain. We have previously reported that Polη-deficient and Polη/Polι double-deficient mice showed increased susceptibility to UV-induced carcinogenesis. Here, we investigated UV-induced mutation frequencies and spectra in the epidermal cells of Polη- and/or Polι-deficient mice. While Polη-deficient mice showed significantly higher UV-induced mutation frequencies than wild-type mice, Polι deficiency did not influence the frequencies in the presence of Polη. Interestingly, the frequencies in Polη/Polι double-deficient mice were statistically lower than those in Polη-deficient mice, although they were still higher than those of wild-type mice. Sequence analysis revealed that most of the UV-induced mutations in Polη-deficient and Polη/Polι double-deficient mice were base substitutions at dipyrimidine sites. An increase in UV-induced mutations at both G:C and A:T pairs associated with Polη deficiency suggests that Polη contributes to accurate TLS past both thymine- and cytosine-containing dimers in vivo. A significant decrease in G:C to A:T transition in Polη/Polι double-deficient mice when compared with Polη-deficient mice suggests that Polι is involved in error-prone TLS past cytosine-containing dimers when Polη is inactivated..
13. Naoya Kubokura, Fumi Takahashi, Masaki Ariuoka, Yoshihara Tatsuya, Kazunobu Igawa, Katsuhiko Tomooka, Sachio Morimoto, Yoshimichi Nakatsu, Teruhisa Tsuzuki, Yusaku Nakabeppu, Takayuki Matsumoto, Kitazono T, Toshiyuki Sasaguri, Differentiation-inducing factor-3 inhibits intestinal tumor growth in vitro and in vivo., 10.1016/j.jphs.2015.03.005, 127, 4, 446-455, 2015.04.
14. Ryoichi Kyuragi, Takuya Matsumoto, Yui Harada, Satoru Saito, Mitsuho Onimaru, Yoshimichi Nakatsu, Teruhisa Tsuzuki, Masatoshi Nomura, Yoshikazu Yonemitsu, Yoshihiko Maehara, BubR1 insufficiency inhibits neointimal hyperplasia through impaired vascular smooth muscle cell proliferation in mice, Arterioscler. Thromb. Vasc. Biol., 10.1161/ATVBAHA.114.304737, 35, 341-347, 2015.02, Objective—BubR1, a cell cycle–related protein, is an essential component of the spindle checkpoint that regulates cell division. Mice with BubR1 expression reduced to 10% of the normal level display a phenotype characterized by progeria; however, the involvement of BubR1 in vascular diseases is still unknown. We generated mice in which BubR1 expression was reduced to 20% (BubR1L/L mice) of that in wild-type mice (BubR1+/+) to investigate the effects of BubR1 on arterial intimal hyperplasia.
Approach and Results—Ten-week-old male BubR1L/L and age-matched wild-type littermates (BubR1+/+) were used in this study. The left common carotid artery was ligated, and histopathologic examinations were conducted 4 weeks later. Bone marrow transplantation was also performed. Vascular smooth muscle cells (VSMCs) were isolated from the thoracic aorta to examine cell proliferation, migration, and cell cycle progression. Severe neointimal hyperplasia was observed after artery ligation in BubR1+/+ mice, whereas BubR1L/L mice displayed nearly complete inhibition of neointimal hyperplasia. Bone marrow transplantation from all donors did not affect the reconstitution of 3 hematopoietic lineages, and neointimal hyperplasia was still suppressed after bone marrow transplantation from BubR1+/+ mice to BubR1L/L mice. VSMC proliferation was impaired in BubR1L/L mice because of delayed entry into the S phase. VSMC migration was unaffected in these BubR1L/L mice. p38 mitogen–activated protein kinase–inhibited VSMCs showed low expression of BubR1, and BubR1-inhibited VSMCs showed low expression of p38.
Conclusions—BubR1 may represent a new target molecule for treating pathological states of vascular remodeling, such as restenosis after angioplasty..
15. Takuro Isoda, Yoshimichi Nakatsu, Kazumi Yamauchi, Takashi Yao, Jingshu Piao, Hiroshi Honda, Yusaku Nakabeppu, Teruhisa Tsuzuki, Abnormality in Wnt Signaling is Causatively Associated with Oxidative Stress-Induced Intestinal Tumorigenesis in MUTYH-Null Mice, Int. J. Biol. Sci., 10.7150/ijbs.9241, 10, 8, 940-947, 2014.08.
16. Fumi Takahashi, Yoshihara Tatsuya, Kentaro Jingushi, Kazunobu Igawa, KATSUHIKO TOMOOKA, Yutaka Watanabe, Sachio Morimoto, Yoshimichi Nakatsu, Teruhisa Tsuzuki, Yusaku Nakabeppu, Toshiyuki Sasaguri, DIF-1 inhibits tumor growth in vivo reducing phosphorylation of GSK-3β and expressions of cyclin D1 and TCF7L2 in cancer model mice, Biochemical pharmacology, 10.1016/j.bcp.2014.03.006, 89, 340-348, 2014.06.
17. Jingshu Piao, Yoshimichi Nakatsu, Mizuki Ohno, Ken-ichi Taguchi, Teruhisa Tsuzuki, Mismatch repair deficient mice show susceptibility to oxidative stress-induced intestinal carcinogenesis, International Journal of Biological Sciences, 10.7150/ijbs.5750, 10, 1, 73-79, 2013.12.
18. Teik How Lim, Ryosuke Fujikane, Shiori Sano, Ryuji Sakagami, Yoshimichi Nakatsu, Teruhisa Tsuzuki, Mutsuo Sekiguchi, Masumi Hidaka, Activation of AMP-activated protein kinase by MAPO1 and FLCN induces apoptosis triggered by alkylated base mismatch in DNA, DNA Repair, 10.1016/j.dnarep.2011.11.006, 11, 3, 259-266, 2012.03, O 6-Methylguanine produced in DNA by the action of simple alkylating agents, such as N-methyl-N-nitrosourea (MNU), causes base-mispairing during DNA replication, thus leading to mutations and cancer. To prevent such outcomes, the cells carrying O 6-methylguanine undergo apoptosis in a mismatch repair protein-dependent manner. We previously identified MAPO1 as one of the components required for the induction of apoptosis triggered by O 6-methylguanine. MAPO1, also known as FNIP2 and FNIPL, forms a complex with AMP-activated protein kinase (AMPK) and folliculin (FLCN), which is encoded by the BHD tumor suppressor gene. We describe here the involvement of the AMPK-MAPO1-FLCN complex in the signaling pathway of apoptosis induced by O 6-methylguanine. By the introduction of siRNAs specific for these genes, the transition of cells to a population with sub-G 1 DNA content following MNU treatment was significantly suppressed. After MNU exposure, phosphorylation of AMPKα occurred in an MLH1-dependent manner, and this activation of AMPK was not observed in cells in which the expression of either the Mapo1 or the Flcn gene was downregulated. When cells were treated with AICA-ribose (AICAR), a specific activator of AMPK, activation of AMPK was also observed in a MAPO1- and FLCN-dependent manner, thus leading to cell death which was accompanied by the depolarization of the mitochondrial membrane, a hallmark of the apoptosis induction. It is therefore likely that MAPO1, in its association with FLCN, may regulate the activation of AMPK to control the induction of apoptosis triggered by O 6-methylguanine..
19. Ito R, Sekiguchi M, Setoyama D, Nakatsu Y, Yamagata Y, Hayakawa H., Cleavage of Oxidized Guanine Nucleotide and ADP-sugar by Human NUDT5 Protein, The Journal of Biochemistry, 149, 6, 731-738, 2011.06.
20. Teruhisa Tsuzuki, Jingshu Piao, Takuro Isoda, Kunihiko Sakumi, Yusaku Nakabeppu, Yoshimichi Nakatsu, Oxidative stress-induced tumorigenesis in the small intestine of various types of DNA repair-deficient mice, Health Physics, 100, 293-294, 2011.05.
21. Komori K, Takagi Y, Sanada M, Lim TH, Nakatsu Y, Tsuzuki T, Sekiguchi M, Hidaka M, A novel protein, MAPO1, that functions in apoptosis triggered by O6-methylguanine mispair in DNA, Oncogene, 28(8):1142-1150, 2009.02.
22. Kuraoka I, Ito S, Wada T, Hayashida M, Lee L, Saijo M, Nakatsu Y, Matsumoto M, Matsunaga T, Handa H, Qin J, Nakatani Y, Tanaka K, Isolation of XAB2 complex involved in pre-mRNA splicing, transcription, and transcription-coupled repair, J Biol Chem, 283(2):940-950, 2008.01.
23. Sakamoto K, Tominaga Y, Yamauchi K, Nakatsu Y, Sakumi K, Yoshiyama K, Egashira A, Kura S, Yao T, Tsuneyoshi M, Maki H, Nakabeppu Y, Tsuzuki T, MUTYH-null mice are susceptible to spontaneous and oxidative stress induced intestinal tumorigenesis, Cancer Research, 67(14):6599-6604, 2007.07.
1. 織田 信弥、中津 可道, Mononucleotide microsatellite instability caused by DNA Polymerase defects
, 第 79 回日本癌学会学術総会, 2020.10, DNA mismatch repair (MMR) and DNA polymerase proofreading (PR) are the two major mechanisms counteracting replication errors (RER) and suppress mutations occurring on the genome. Germline mutations in MMR genes are known to cause an inherited cancer predisposition, Lynch syndrome (LS), and, in addition, those in the PR domains of the polymerase genes have also been reported in LS-like kindred, recently. A human colorectal cancer cell line, DLD-1, which carries a polymerase delta PR domain mutation, In prokaryotes and yeasts, DNA polymerase proofreading (PPR) and DNA mismatch repair (MMR) cooperatively counteracts replication errors leading to repeat sequence destabilization (i.e. insertions/deletions of repeat units). However, PPR has not thus far been regarded as a mechanism stabilizing repeat sequences in higher eukaryotic cells. In a human cancer cell line, DLD-1, which carries mutations in both MSH6 and the Exo domain of POLD1, we previously observed that mononucleotide microsatellites were markedly destabilized whereas being stable in the simple MMR-defective backgrounds. In this study, we introduced the Exo domain mutation found in DLD-1 cells into MSH2-null HeLa cell clones, using CRISPR/Cas9 system. In the established Exo-/MMR-mutated HeLa clones, mononucleotide repeat sequences were remarkably destabilized as in DLD-1 cells. In contrast, dinucleotide microsatellites were readily destabilized in the parental MMR-deficient backgrounds, and the instability was not notably increased in the genome-edited HeLa clones. Here, we show an involvement of the Exo domain functions of DNA polymerase delta in mononucleotide repeat stabilization in human cells, which also suggests a possible role division between DNA polymerase Exo domain and MMR in repeat maintenance in the human genome..
2. Mizuki OHNO, Kunihiko SAKUMI, Noriko TAKANO, Kosuke TESHIMA, Kyoko HIDAKA, Yoshimichi NAKATSU, Teruhisa TSUZUKI , Detection of de novo germline mutations in DNA repair-deficient mice lines, The Joint Meeting of The 6th Asian Congress on Environmental Mutagens(ACEM) and the 48th Annual Meeting of the Japanese Environmental Mutagen Society(JEMS), 2019.11, It is imperative to assess the trans generational effect of environmental mutagens such as radiation or chemicals. However, evaluation of the mutagenic effect, especially for low-dose mutagens, on de novo germline mutation (dGM) is still challenging because of low mutation rate in wild-type animals under natural conditions. Here, we use DNA repair-deficient mice lines for the detection of spontaneous dGMs in order to understand the molecular mechanisms behind germline genome integrity. We generated several mutant-mice family lines, deficient in 8-oxoguanine-related repair pathway, deficient in DNA mismatch repair (MMR) pathway or deficient in nucleotide excision repair (NER) pathway. Genomic DNA of parents and offspring were subjected to whole-exome or whole-genome sequencing for the detection of dGMs. Mth1/Ogg1/Mutyh-triple gene knocked out mice (8-oxoguaniene-induced mutations are usually well suppressed by MTH1, OGG1, MUTYH) exhibited 37-lold higher dGM rate than that in wild-type mice with a typical mutational pattern induced by 8-oxoguanine. In the MMR deficient mice, insertions/deletions at tandem repeat sequences were frequently observed in addition to high rate of G>A base substitution. On the other hand, dGM rate in NER deficient mice was almost same level as that in wild-type mice, suggesting that the repair targets of NER may not be frequently generated in the germ cells. Our results suggesting that dGM rate and spectra may fluctuate with the frequency of spontaneous DNA damages and DNA repair efficacy..
3. Yoshimichi Nakatsu, Noriko Takano, Mizuki Ohno, Satoshi Kitazaki, Kazunori Koga, Akiyo Tanaka, Masaharu Shiratani, Application of transgenic mice to analyze genotoxic effects induced by non-thermal atmospheric air plasma
, 10th Anniversary International Symposium on Advanced Plasma Science and its Applications for Nitrides and Nanomaterials /11th International Conference on Plasma-Nano Technology & Science. (ISPlasma2018 / IC-PLANTS2018), 2018.03, Non-thermal atmospheric plasma has been used for biomedical applications such as blood coagulation, wound healing, and decontamination [1]. In addition, the plasma has been proposed as a potential tool for cancer therapy because of its ability to induce apoptosis in cancer cells.
Non-thermal atmospheric plasma as well as ionizing radiation generates reactive oxygen species (ROS) in the liquid phase [2]. ROS such as hydroxyl radical induce DNA double-strand break (DSB) and lead to cell death. The apoptotic induction of cancer cells by plasma treatment could be mediated by ROS. The cytotoxic activity was also observed in plasma-treated medium, so called plasma-activated medium (PAM) [3]. PAM has been shown to have a selective cytotoxicity in several cancer cell lines, even chemotherapeutic agent resistant cell lines, providing a potential treatment for novel anti-cancer therapies [4]. The cytotoxic effect of PAM is suggested to be mediated by ROS such as H2O2.
DSBs can be repaired by two different pathways: homologous recombination (HR) and non-homologous end joining (NHEJ) [5]. HR is an error-free repair pathway, but it can be operated only in S and G2 phase of cell cycle because HR needs the sister chromatid as a repair template that is synthesized by DNA replication during the S phase. NHEJ is the predominant repair pathway during the G1 phase, and this is especially important for the cells in tissues/organs that is resting in G0/G1 phase or non-dividing status. NHEJ is characterized by rejoining of broken ends without the use of extensive homology, and is frequently associated with the presence of small insertions/deletions at DSB site. In addition to DSB, ROS induced various modified bases, and more than 20 different types of oxidatively altered purines and pyrimidines have been detected [6]. These DNA damages also induce mutations and cell death, if not repaired.
Biomedical applications of plasma in either direct or indirect way are always associated with the generation of ROS that cause oxidation of cellular DNA in the treated tissues. If the plasma-induced DNA damages are not correctly repaired in the surviving cells, the accumulation of mutations occurs in the surviving cells, leading to the increase of a risk for carcinogenesis in the treated tissues. Therefore, biomedical applications of plasma need to be strictly evaluated in terms of safety implications. Here, we performed mutation analysis using transgenic mice exposed by non-thermal atmospheric air plasma. Direct exposure of air plasma jet induced the mutations, mainly deletions, in the in vitro cultured splenocytes from rpsL-transgenic mice in a dose-dependent manner. Direct exposures of air plasma jet induced DNA damages in the epidermis of mouse skin. These results suggested that further investigations are needed to establish safety protocols for the long-term applications of plasma such as wound-healing and cancer treatment.
[1] G. Fridman, G. Friedman, A. Gutsol, A. B. Shekhter, V. N. Vasilets and A. Fridman: Plasma Processes and Polymers. 5, 503 (2008).
[2] M. U. Rehman, P. Jawaid, H. Uchiyama and TKondo: Arch Biochem Biophys. 605,19 (2016).
[3] H. Tanaka, M. Mizuno, K. Ishikawa, K. Nakamura, H. Kajiyama, H. Kano, F. Kikkawa and M. Hori: Plasma Medicine. 1, 265 (2011).
[4] F. Utsumi, H. Kajiyama, K. Nakamura, H. Tanaka, M. Mizuno, K. Ishikawa, H. Kondo H. Kano, M. Hori and F. Kikkawa: PLoS One. 8, e81576 (2013).
[5] K. Rodgers and M. McVey: J Cell Physiol. 231, 15 (2016).
[6] E. Gajewski, G. Rao, Z. Nackerdien and M. Dizdaroglu: Biochemistry. 29, 7876 (1990)..
4. Tsuzuki Teruhisa, Ohno Mizuki, Takano Noriko, Taguchi Ken-ichi, Nakabeppu Yusaku, Nakatsu Yoshimichi, DNA repair system as a constituent of mechanism underlying practical threshold of oxidative stress-induced tumorigenesis, The 12th International Conference and the 5th Asian Congress on Environmental Mutagens, 2017.11.
5. Tsuzuki Teruhisa, Ohno Mizuki, Takano Noriko, Taguchi Ken-ichi, Nakabeppu Yusaku, Nakatsu Yoshimichi, Oxidative stress-induced tumorigenesis: Lesson from the experiments with DNA repair-deficient mice, 4th Transgenic Technology Meeting, 2017.10.
6. Tsuzuki Teruhisa, Ohno Mizuki, Takano Noriko, Taguchi Ken-ichi, Nakabeppu Yusaku, Aoki Yasunobu, Nohmi Takehiko, Nakatsu Yoshimichi, Oxidative stress-induced intestinal tumors in Mutyh-deficient mice treated with low doses of potassium bromate, 6th US-Japan DNA Repair Meeting, 2017.05.
7. Yoshimichi Nakatsu, Noriko Takano, Mizuki Ohno, Satoshi Kitazaki, Kazunori Koga, Akiyo Tanaka, Masaharu Shiratani, Teruhisa Tsuzuki, Analyses of oxidative mutagenesis and carcinogenesis using genetically modified mice: application to plasma medicine, The 4th International Workshop on Plasma for Cancer Treatment (IWPCT-2017), 2017.03, Oxygen radicals attack DNA, and induce various lesions into the DNA. Among such lesions, 8-oxo-7, 8-dihydroguanine (8-oxoG) is highly mutagenic because of its ambiguous pairing property. Three enzymes, MTH1, OGG1, and MUTYH, play important roles in avoiding the 8-oxoG-related mutagenesis in mammalian cells. We have established an experimental system for oxidative DNA damage-induced mutagenesis and tumorigenesis in the intestine of mice. Oral administration of oxidizing reagent, potassium bromate (KBrO3), effectively induced G:C to T:A mutations and epithelial tumors in the small intestines of Mutyh-deficient mice, implying the significance of Mutyh in the suppression of mutagenesis and tumorigenesis induced by oxidative stress. To elucidate the roles of other DNA repair genes in the suppression of oxidative stress-induced tumorigenesis, we performed KBrO3-induced tumorigenesis experiments using various types of DNA repair-deficient mice. We observed an enhanced tumor-formation in the small intestines of Msh2-deficient mice, as compared with the wild type. No such enhancement was observed in Xpa-deficient mice. These results indicate that mismatch repair, but not nucleotide excision repair, is involved in the suppression of oxidative stress-induced intestinal tumorigenesis in mice. The number of tumors was marginally increased in Ogg1- and Mth1-deficient mice, in comparison to the wild-type mice, suggesting that in contrast to Mutyh, Ogg1 and Mth1 may play a limited role in the suppression of intestinal tumorigenesis caused by oxidative stress. Our data indicate that among the repair factors examined, only Mutyh and Msh2 play a significant role in the suppression of oxidative stress-induced intestinal tumorigenesis in mice.
To establish the application of plasma to humans, it is important to evaluate the potential risk of plasma to organisms. Plasma is a gas composed of electrons, various ions, and reactive oxygen/nitrogen species. The plasma exposure produced RONS not only in external environment but also in cells, thus anticipated the inductions of DNA damages and mutations. Therefore, we performed the mutation analyses using mouse cells exposed by non-thermal atmospheric air plasma. Direct exposure of air plasma jet induced the mutations, mainly deletions, in the in vitro cultured splenocytes from rpsL-transgenic mice in a dose-dependent manner. On the other hand, direct exposures of air plasma jet induced DNA damages in the epidermis of mouse skin. However, the increases of mutation frequencies were not detected in the air plasma-exposed mouse skins. These results suggest that genetically modified mice are useful tool for the risk assessment of plasma..
, International Conference on Plasma Medical Science Innovation (ICPMSI) 2017, 2017.02.
9. Yoshimichi Nakatsu, Noriko Takano, Mizuki Ohno, Satoshi Kitazaki, Kazunori Koga, Akiyo Tanaka, Masaharu Shiratani, Teruhisa Tsuzuki, Analyses of oxidative mutagenesis and carcinogenesis using genetically modified mice: application to plasma medicine, 第26回日本MRS年次大会/Internatinal Symposium C-4, 2016.12.
10. Hayashida Genki, Nakatsu Yoshimichi, Hidaka Kyoko, Fujikane Ryosuke, Hidaka Masumi, Tsurimoto Toshiki, Tsuzuki Teruhisa, Development of assay systems to characterize the variants of mismatch repair factor MSH2 found in Lynch syndrome, The 10th International 3R (Replication, Recombination and Repair) Symposium, 2016.11.
11. Yoshimichi Nakatsu, Noriko Takano, Mizuki Ohno, Satoshi Kitazaki, Kazunori Koga, Takaaki Amano, Akiyo Tanaka, Miyuki Hirata, Masaharu Shiratani, Teruhisa Tsuzuki , Mutagenesis in Human and Mouse Cells Irradiated by Non-thermal Atmospheric Air Plasma
, 6th International Conference on Plasma Medicine (ICPM-6), 2016.09.
12. Mizuki Ohno, Noriko Takano, SAKUMI Kunihiko, Ryutaro Fukumura, Yuki Iwasaki, Toshimichi Ikemura, Yoichi Gondo, Yusaku Nakabeppu, Yoshimichi Nakatsu, Teruhisa Tsuzuki, Role of the oxidative DNA damage repair system in somatic and germline mutations in mice, Zing conference "Genome Integrity" , 2015.08.
13. Mizuki Ohno, Noriko Takano, Kunihiko Sakumi, Ryutaro Fukumura, Yuki Iwasaki, Toshimichi Ikemura, Yoichi Gondo, Yusaku Nakabeppu, Yoshimichi Nakatsu, Teruhisa Tsuzuki, Influence of Oxidative DNA Damage on the Rate of Somatic and Germiline Mutation, 15th International Congress of Radiation Research, 2015.05.
14. Teruhisa Tsuzuki, Mizuki Ohno, Noriko Takano, Kenichi Taguchi, Yusaku Nakabeppu, Yoshimichi Nakatsu, Oxidative Stress-induced Tumorigenesis: Lesson from the Experiments with DNA Repair-deficient Mice, Advances in Understanding the Biological Consequences by Environmental Stressors, 15th International Congress of Radiation Research, 2015.05.
15. Yoshimichi Nakatsu, Jingshu Piao, Takuya Hashizume, Mizuki Ohno, Kenichi Taguchi, Teruhisa Tsuzuki, Mismatch Repair Deficient Mice Show Susceptibility to Oxidative Stress-induced Intestinal Carcinogenesis, 15th International Congress of Radiation Research, 2015.05.
16. Teruhisa Tsuzuki, Mizuki Ohno, Noriko Takano, Kenichi Taguchi,, Yusaku Nakabeppu, Yasunobu Aoki, Takehiko Nohmi, Yoshimichi Nakatsu, Oxidative stress-induced intestinal tumors in Mutyh-deficient mice treated with low doses of potassium bromate, 4th Asian Conference on Environmental Mutagen, 2014.12.
17. Teruhisa Tsuzuki, Mizuki Ohno, Noriko Takano, Kenichi Taguchi, Yusaku Nakabeppu, Yasunobu Aoki, Takehiko Nohmi, Yoshimichi Nakatsu, Oxidative stress-induced intestinal tumors in Mutyh-deficient mice treated with low doses of potassium bromate, 5th US-Japan DNA Repair Meeting, 2014.10.
18. Teruhisa Tsuzuki, Jing Shu Piao, Noritaka Matsumoto, Yoshimichi Nakatsu, The roles of mismatch repair system and p53 in the suppression of oxidative stress-induced intestinal tumor-formation in mice., 4th US-Japan DNA Repair Meeting, 2012.04, In living cells, reactive oxygen species (ROS) is constantly generated by cellular metabolic reactions such as mitochondrial respiration, and also by external cause such as exposure to radiation and chemicals. DNA and its substrate, nucleotides, are continuously oxidized by ROS, and resulting oxidative DNA damages appear to cause mutagenesis and carcinogenesis in mammal. To counteract this hazardous effects of ROS, mammalian cells are equipped with several enzymatic systems. Following KBrO3 treatment in all mouse types, tumor-formation in the small intestines of Msh2-/- mice significantly increased as compared with Msh2+/+ and Msh2+/- mice. These results suggest that mismatch repair (MMR) is involved in the suppression of oxidative stress-induced intestinal tumorigenesis in mice. Following KBrO3-treatment, tumor-formation in the small intestine of Trp53-/- mice significantly increased as compared with Trp53+/+ and Trp53+/- mice and tumor-formation in the small intestine of Trp53+/- mice moderately increased as compared with Trp53+/+ mice. In addition to previous observation with Mutyh-deficient mice, our data suggest that MMR system as well as Trp53 is also involved in the suppression of oxidative stress-induced intestinal tumorigenesis in mice..
19. Oxidartive stress-induced intestinal tumorigensis in Trp53-deficient mice.
20. Oxidartive stress-induced intestinal tumorigensis in Trp53-deficient mice.
21. The roles of tumor-associated genes, Mutyh, Msh2 and Trp53, in the suppression of oxidative stress-induced tumorigenesis in mice.
22. A MAPO1 protein complex that involved in the induction of apoptosis triggered by O6-methylguanine.
23. Oxidative stress and intestinal carcinogenesis.
Membership in Academic Society
  • The Japanese Environmental Mutagen Society (JEMS)
  • Genetic Society of Japan
  • The Japan Radiation Research Society
  • Japanese Cancer Association
  • The Molecular Biology Society of Japan
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