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Masatoshi Fujita Last modified date:2023.11.28

Professor / Department of Pharmaceutical Health Care and Sciences
Department of Pharmaceutical Health Care and Sciences
Faculty of Pharmaceutical Sciences

Graduate School
Undergraduate School

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Academic Degree
Country of degree conferring institution (Overseas)
Field of Specialization
Molecular Biology, Biochemistry, Molecular Oncology
Total Priod of education and research career in the foreign country
Outline Activities
Chromosomal DNA Replication and Cancer Research
In human cells, genomic DNA, which carries genetic information, has to be replicated faithfully, completely, and only once during a single cell cycle to maintain integrity. If some errors occur during copying DNA (DNA replication), then it would sometimes lead to bad consequences. “Cancer” is one and serious example resulting from such replication errors (mutations).

Molecular mechanisms for cell cycle regulation of DNA replication initiation
Several molecular mechanisms contribute to the maintenance of genomic integrity. Replicative DNA polymerases synthesize new daughter strand using complementary parental strand. However, DNA polymerases very rarely incorporate incorrect nucleotides. The mismatch repair pathway that removes inappropriate nucleotides is a fail-safe mechanism for such errors and the disturbance is well known to cause genomic instability and eventual cancer. Chromosomal DNAs are often damaged, for example by ultraviolet, which should also be repaired. Disruption of such repair mechanism also leads to cancer.
We have been interested in elucidating molecular mechanisms for cell cycle regulation of DNA replication initiation, another crucial aspect of replication controls. In human cells, genomic DNA is fragmented into multiple chromosomes, which may allow genome size to expand during the evolution. As a result, DNA replication initiates from multiple replication origins. However, effective operation of the “multiple replication origin” system gives rise to an important problem: i.e. multiple replication origins should each be activated precisely only once during each S phase.
Recent research progress by us and other groups has uncovered the mechanisms. It is now clear that the “once and only once replication per single cell cycle” is achieved by the periodic assembly and disassembly of pre-replication complexes (pre-RCs) at replication origins. The pre-RC assembly reaction, known as “licensing”, involves the loading of a presumptive replicative helicase, the MCM2-7 complex, onto chromatin by the origin recognition complex (ORC), CDC6 and Cdt1. Two critical inhibitory factors for the pre-RC assembly are cyclin/Cdks (Cdk1 and Cdk2) and geminin. During late mitosis through the G1 phase, a cell cycle regulatory E3 ubiquitin ligase APC/C restrains cyclins and geminin by targeting them for proteolysis through polyubiquitination. Thus, pre-RC assembly only occurs during this period. Following APC/C inactivation at the onset of S phase, Cdks are activated, stimulating DNA unwinding by MCM. Then, DNA polymerases synthesize new DNA.
To prevent re-replication, the re-establishment of pre-RC, in other words re-binding of MCM, needs to be suppressed during the S, G2 and M phases of the cell cycle. Cdks play a central role also in this context by preventing re-establishment of pre-RC through multiple mechanisms. One is by phosphorylation of CDC6, leading to CDC6 nuclear export. ORC1 is degraded after S phase, presumably depending on phosphorylation by cyclin A/Cdks and binding to SCFSkp2 ubiquitin ligase.

Cdt1, a central factor for the cell cycle regulation of replication initiation: Elucidating the strict regulations by three ubiquitin ligases
It was originally suggested that inhibition of Cdt1 function after S phase is due to geminin binding. However, we have recently demonstrated that three ubiquitin ligases strictly control Cdt1 proteolysis, showing that Cdt1 is a central player in the cell cycle regulation of replication initiation. During S and G2 phases, Cdt1 is brought to proteolysis by Cdk phosphorylation-dependent SCFSkp2-mediated ubiquitination. Interestingly, Cdt1 is also regulated by replication-coupled, Cullin4-DDB1Cdt2 ubiquitin ligase-mediated ubiquitination, which is dependent on Cdt1 binding to PCNA, an eukaryotic sliding clamp stimulating DNA polymerases. In addition, when cells enter quiescence, Cdt1 is rapidly cleared by APC/CCdh1-mediated proteolysis.

Cdt1 deregulation induces chromosomal instability, a mechanism leading to cancer
As expected from the strict regulation, deregulation of Cdt1 is a deleterious insult, leading to re-replication and/or chromosomal damage. The induced chromosomal instability may eventually lead to carcinogenesis and Cdt1 overexpression is in fact often observed in human cancers. By other groups, it has been suggested that Cdt1 overexpression could endow cells with the transforming ability.

Cdt1-geminin system could be a novel molecular target for anti-cancer chemotherapeutic agents
We also think that tumor cells could be selectively eliminated by modulating the Cdt1-geminin interactions and have been seeking small molecule compounds that affect the interaction.

Research Interests
  • Function and regulation of DNA replication initiation proteins, ORC, CDC6, Cdt1 and MCM during the cell cycle
    keyword : DNA replication, cell cycle regulation, replication initiation, ORC, CDC6, Cdt1, MCM
  • Involvement of the replication initiation proteins in telomere homeostasis
    keyword : DNA replication initiation factors, telomere
  • Molecular mechanisms for cellular responses to chromosomal stress through ATM-Chk2 and ATR-Chk1 pathways and involvement of the replication initiation proteins in such process
    keyword : replication stress、ATM、ATR、DNA replication initiation factors
  • Search for Cdt1-geminin binding inhibitors that could selectively damage cancer cells by inducing re-replication
    keyword : Cdt1-geminin inhibitor、anti-neoplastic drug
Academic Activities
1. Nozomi Sugimoto, Masatoshi Fujita, Molecular mechanism for chromatin regulation during MCM loading in mammalian cells, Springer New York LLC, 10.1007/978-981-10-6955-0_3, 61-78, 2018.01, DNA replication is a fundamental process required for the accurate and timely duplication of chromosomes. During late mitosis to G1 phase, the MCM2-7 complex is loaded onto chromatin in a manner dependent on ORC, CDC6, and Cdt1, and chromatin becomes licensed for replication. Although every eukaryotic organism shares common features in replication control, there are also some differences among species. For example, in higher eukaryotic cells including human cells, no strict sequence specificity has been observed for replication origins, unlike budding yeast or bacterial replication origins. Therefore, elements other than beyond DNA sequences are important for regulating replication. For example, the stability and precise positioning of nucleosomes affects replication control. However, little is known about how nucleosome structure is regulated when replication licensing occurs. During the last decade, histone acetylation enzyme HBO1, chromatin remodeler SNF2H, and histone chaperone GRWD1 have been identified as chromatin-handling factors involved in the promotion of replication licensing. In this review, we discuss how the rearrangement of nucleosome formation by these factors affects replication licensing..
1. Yoshida, K. and Fujita, M. , DNA damage response that enhance resilience to replication stress., Cell. Mol. Life Sci. , 10.1007/s00018-021-03926-3, 2021.11.
2. Takuya Takafuji, Kota Kayama, Nozomi Sugimoto, Masatoshi Fujita, GRWD1, a new player among oncogenesis-related ribosomal/nucleolar proteins, Cell Cycle, 10.1080/15384101.2017.1338987, 2017.08, Increasing attention has been paid to certain ribosomal or ribosome biosynthesis-related proteins involved in oncogenesis. Members of one group are classified as “tumor suppressive factors” represented by RPL5 and RPL11; loss of their functions leads to cancer predisposition. RPL5 and RPL11 prevent tumorigenesis by binding to and inhibiting the MDM2 ubiquitin ligase and thereby up-regulating p53. Many other candidate tumor suppressive ribosomal/nucleolar proteins have been suggested. However, it remains to be experimentally clarified whether many of these factors can actually prevent tumorigenesis and if so, how they do so. Conversely, some ribosomal/nucleolar proteins promote tumorigenesis. For example, PICT1 binds to and anchors RPL11 in nucleoli, down-regulating p53 and promoting tumorigenesis. GRWD1 was recently identified as another such factor. When overexpressed, GRWD1 suppresses p53 and transforms normal human cells, probably by binding to RPL11 and sequestrating it from MDM2. However, other pathways may also be involved..
3. Mitsunori Higa, Masatoshi Fujita, Kazumasa Yoshida, DNA Replication Origins and Fork Progression at Mammalian Telomeres, Genes (Basel). 2017 Mar 28;8(4). pii: E112. doi: 10.3390/genes8040112., 2017.03.
4. Fujita M, Cdt1 revisited: complex and tight regulation during the cell cycle and consequences of deregulation in mammalian cells, Cell Div. 1: 22, 2006.
5. Fujita M., DNA Replication Initiation., Encyclopedic Reference of Genomics and Proteomics in Molecular Medicine (edited by Ganten, D. and Ruckpaul, K.). Springer, Berlin Heidelberg & New York, pp446-449, 2006.
1. Higa, Mitsunori; Matsuda, Yukihiro; Fujii, Jumpei; Sugimoto, Nozomi; Yoshida, Kazumasa; Fujita, Masatoshi, TRF2-mediated ORC recruitment underlies telomere stability upon DNA replication stress, NUCLEIC ACIDS RESEARCH, 10.1093/nar/gkab1004, 49, 21, 12234-12251, 2021.12, テロメアは内因性の「複製が困難な領域」であることが知られている。テロメアリピート配列結合タンパク質TRF2は必須のテロメア構造保護因子であるが、一方でORCと言われる複製開始点認識複合体と結合し、この結合を介して効率よく複製開始複合体がテロメアに形成されることを我々は報告していた。しかし、この分子機構が実際にテロメアの安定性保持に必要なのかは未だ不明確なままであった。今回、ORC1結合欠損変異型TRF2を同定すること等により、このTRF2とORCの結合を介した効率よい複製開始複合体のテロメアでの形成がテロメア安定性保持に重要であることを明らかにした。例えば、ORC1結合欠損TRF2 EE変異体を持つ細胞では、テロメアが切断されてしまうことで形成されるテロメア含有微小核の形成頻度が複製ストレス時に大きく上昇する。以上のことから、TRF2-ORC結合を介して染色体末端に効率よく複製開始複合体を形成することにより、染色体内側からのDNA複製の進行が阻害された場合に末端側から複製を開始することができ、テロメアの安定性が保たれることが明らかになった。本成果は、ゲノム安定性維持機構の全容解明に向けた重要な成果であると考えられる。.
2. Ishimoto R, Tsuzuki Y, Matsumura T, Kurashige S, Enokitani K, Narimatsu K, Higa M, Sugimoto N, Yoshida K, Fujita M., SLX4-XPF mediates DNA damage responses to replication stress induced by DNA-protein interactions, JOURNAL OF CELL BIOLOGY, 10.1083/jcb.202003148, 220, 1, 2021.01, DNA複製の進行を妨げゲノム不安定性を引き起こす複製ストレスに対するDNA損傷応答経路の研究は、これまで薬剤や紫外線等による外因性の複製ストレスに対するものを中心に進められてきた。一方、ヒトのゲノムにおいて内因性の複製ストレスとして「複製が困難なゲノム領域 (テロメア、セントロメア、リボソームDNA領域等)」の存在が知られているが、これら領域の複製ストレスに対する応答の詳細は不明であった。本研究では、内因性の複製ストレスの一因である「強固なDNA-タンパク質複合体」にDNA複製装置が衝突した時にどのようなDNA損傷応答が起きるのかを、ヒト染色体上での複製障害物として大腸菌由来のlacO-LacI相互作用を利用したユニークな実験モデル系を用いて明らかにした。すなわち、まずDNA損傷応答の足場タンパク質であるSLX4が構造特異的DNAエンドヌクレアーゼXPFをストレス部位へと呼び込み、その下流でATR、FANCD2、RAD52が働くという新規経路が明らかとなった。さらに、lacO配列の複製完了にSLX4-XPF-ATR経路が大きく寄与することもわかった。本研究は、DNA複製やゲノム修復維持機構、さらにがん遺伝子活性化による複製ストレスによって引き起こされるゲノム不安定性誘導機構等の全容解明に向けた重要な基盤になると考えられる。.
3. Takuya Yokoyama, Masaki Yukuhiro, Yuka Iwasaki, Chika Tanaka, Kazunari Sankoda, Risa Fujiwara, Atsushi Shibuta, Taishi Higashi, Keiichi Motoyama, Hidetoshi Arima, Kazumasa Yoshida, Nozomi Sugimoto, Hiroyuki Morimoto, Hidetaka Kosako, Takashi Ohshima, Masatoshi Fujita, Identification of candidate molecular targets of the novel antineoplastic antimitotic NP-10, Scientific reports, 10.1038/s41598-019-53259-2, 9, 1, 2019.12, We previously reported the identification of a novel antimitotic agent with carbazole and benzohydrazide structures: N′-[(9-ethyl-9H-carbazol-3-yl)methylene]-2-iodobenzohydrazide (code number NP-10). However, the mechanism(s) underlying the cancer cell-selective inhibition of mitotic progression by NP-10 remains unclear. Here, we identified NP-10-interacting proteins by affinity purification from HeLa cell lysates using NP-10-immobilized beads followed by mass spectrometry. The results showed that several mitosis-associated factors specifically bind to active NP-10, but not to an inactive NP-10 derivative. Among them, NUP155 and importin β may be involved in NP-10-mediated mitotic arrest. Because NP-10 did not show antitumor activity in vivo in a previous study, we synthesized 19 NP-10 derivatives to identify more effective NP-10-related compounds. HMI83-2, an NP-10-related compound with a Cl moiety, inhibited HCT116 cell tumor formation in nude mice without significant loss of body weight, suggesting that HMI83-2 is a promising lead compound for the development of novel antimitotic agents..
4. Hiroki Fujiyama, Takahiro Tsuji, Kensuke Hironaka, Kazumasa Yoshida, Nozomi Sugimoto, and Masatoshi Fujita, GRWD1 directly interacts with p53 and negatively regulates p53 transcriptional activity, Journal of Biochemistry, 10.1093/jb/mvz075, 2019.09.
5. Issay Morii, Yukiko Iwabuchi, Sumiko Mori, Masaki Suekuni, Toyoaki Natsume, Kazumasa Yoshida, Nozomi Sugimoto, Masato T. Kanemaki, Masatoshi Fujita, Inhibiting the MCM8-9 complex selectively sensitizes cancer cells to cisplatin and olaparib, Cancer Science, 10.1111/cas.13941, 110, 3, 1044-1053, 2019.03, MCM8 and MCM9 are paralogues of the MCM2-7 eukaryotic DNA replication helicase proteins and play a crucial role in a homologous recombination-mediated repair process to resolve replication stress by fork stalling. Thus, deficiency of MCM8-9 sensitizes cells to replication stress caused, for example, by platinum compounds that induce interstrand cross-links. It is suggested that cancer cells undergo more replication stress than normal cells due to hyperstimulation of growth. Therefore, it is possible that inhibiting MCM8-9 selectively hypersensitizes cancer cells to platinum compounds and poly(ADP-ribose) polymerase inhibitors, both of which hamper replication fork progression. Here, we inhibited MCM8-9 in transformed and nontransformed cells and examined their sensitivity to cisplatin and olaparib. We found that knockout of MCM9 or knockdown of MCM8 selectively hypersensitized transformed cells to cisplatin and olaparib. In agreement with reported findings, RAS- and human papilloma virus type 16 E7-mediated transformation of human fibroblasts increased replication stress, as indicated by induction of multiple DNA damage responses (including formation of Rad51 foci). Such replication stress induced by oncogenes was further increased by knockdown of MCM8, providing a rationale for cancer-specific hypersensitization to cisplatin and olaparib. Finally, we showed that knocking out MCM9 increased the sensitivity of HCT116 xenograft tumors to cisplatin. Taken together, the data suggest that conceptual MCM8-9 inhibitors will be powerful cancer-specific chemosensitizers for platinum compounds and poly(ADP-ribose) polymerase inhibitors, thereby opening new avenues to the design of novel cancer chemotherapeutic strategies..
6. Shinya Watanabe, Hiroki Fujiyama, Takuya Takafuji, Kota Kayama, Masaki Matsumoto, Keiichi I. Nakayama, Kazumasa Yoshida, Nozomi Sugimoto, Masatoshi Fujita, GRWD1 regulates ribosomal protein L23 levels via the ubiquitin-proteasome system, Journal of Cell Science, 10.1242/jcs.213009, 131, 15, 2018.08, Glutamate-rich WD40 repeat-containing 1 (GRWD1) is a Cdt1- binding protein that promotes mini-chromosome maintenance (MCM) loading through its histone chaperone activity. GRWD1 acts as a tumor-promoting factor by downregulating p53 (also known as TP53) via the RPL11-MDM2-p53 axis. Here, we identified GRWD1- interacting proteins using a proteomics approach and showed that GRWD1 interacts with various proteins involved in transcription, translation, DNA replication and repair, chromatin organization, and ubiquitin-mediated proteolysis. We focused on the ribosomal protein ribosomal protein L23 (RPL23), which positively regulates nucleolar stress responses through MDM2 binding and inhibition, thereby functioning as a tumor suppressor. Overexpression of GRWD1 decreased RPL23 protein levels and stability; this effect was restored upon treatment with the proteasome inhibitor MG132. EDD (also known as UBR5), an E3 ubiquitin ligase that interacts with GRWD1, also downregulated RPL23, and the decrease was further enhanced by co-expression of GRWD1. Conversely, siRNA-mediated GRWD1 knockdown upregulated RPL23. Co-expression of GRWD1 and EDD promoted RPL23 ubiquitylation. These data suggest that GRWD1 acts together with EDD to negatively regulate RPL23 via the ubiquitinproteasome system. GRWD1 expression reversed the RPL23- mediated inhibition of anchorage-independent growth in cancer cells. Our data suggest that GRWD1-induced RPL23 proteolysis plays a role in downregulation of p53 and tumorigenesis..
7. Nozomi Sugimoto, Kazumitsu Maehara, Kazumasa Yoshida, Yasuyuki Ohkawa, Masatoshi Fujita, Genome-wide analysis of the spatiotemporal regulation of firing and dormant replication origins in human cells, Nucleic Acids Research, 10.1093/nar/gky476, 46, 13, 6683-6696, 2018.07, In metazoan cells, only a limited number of mini chromosome maintenance (MCM) complexes are fired during S phase, while the majority remain dormant. Several methods have been used to map replication origins, but such methods cannot identify dormant origins. Herein, we determined MCM7-binding sites in human cells using ChIP-Seq, classified them into firing and dormant origins using origin data and analysed their association with various chromatin signatures. Firing origins, but not dormant origins, were well correlated with open chromatin regions and were enriched upstream of transcription start sites (TSSs) of transcribed genes. Aggregation plots of MCM7 signals revealed minimal difference in the efficacy of MCM loading between firing and dormant origins. We also analysed common fragile sites (CFSs) and found a low density of origins at these sites. Nevertheless, firing origins were enriched upstream of the TSSs. Based on the results, we propose a model in which excessive MCMs are actively loaded in a genome-wide manner, irrespective of chromatin status, but only a fraction are passively fired in chromatin areas with an accessible open structure, such as regions upstream of TSSs of transcribed genes. This plasticity in the specification of replication origins may minimize collisions between replication and transcription..
8. Mitsunori Higa, Seiichiro Kurashige, Daisuke Kohmon, Kouki Enokitani, Satoko Iwahori, Nozomi Sugimoto, Kazumasa Yoshida, Masatoshi Fujita, TRF2 recruits ORC through TRFH domain dimerization, BIOCHIMICA ET BIOPHYSICA ACTA-MOLECULAR CELL RESEARCH, 10.1016/j.bbamcr.2016.11.004, 1864, 1, 191-201, 2017.01.
9. Kota Kayama, Shinya Watanabe, Takuya Takafuji, Takahiro Tsuji, Kensuke Hironaka, Masaki Matsumoto, Keiichi Nakayama, Masato Enari, Takashi Kohno, Kouya Shiraishi, Tohru Kiyono, Kazumasa Yoshida, Nozomi Sugimoto, Masatoshi Fujita, GRWD1 negatively regulates p53 via the RPL11-MDM2 pathway and promotes tumorigenesis, EMBO REPORTS, 10.15252/embr.201642444, 18, 1, 123-137, 2017.01, リボソームタンパク質RPL11は、MDM2ユビキチンリガーゼに結合することによりその活性を阻害し、結果としてp53を誘導する。すなわちRPL11はがん抑制因子として機能する。我々は、GRWD1がRPL11と結合しその機能を抑制することにより、がん遺伝子として機能することを発見した。GRWD1のsiRNAによる抑制は、アクチノマイシンDによる核小体ストレスやブレオマイシンによるDSBによって誘導されるp53活性化を促進する。逆に、GRWD1の過剰発現はp53を不安定化し抑制する。また、GRWD1はがん抑制因子RPL11と直接結合することでRPL11によるMDM2抑制を阻害し、結果としてp53誘導に対して抑制的に働く。更に、 GRWD1をHPV16 E7(RBを不活化)、活性化Ras G12Vと共にヒト正常細胞に発現させると、細胞をtransformすることもわかった。 このGRWD1のtransform活性は、RPL11との結合に依存していた。GRWD1のtransforming活性と一致して、p53が野生型のある種のがん患者において、GRWD1の過剰発現は予後不良と強く相関していた。以上から、GRWD1は新たながん(促進)遺伝子として機能していることが明らかとなった。.
10. Masahiro Aizawa, Nozomi Sugimoto, Shinya Watanabe, Kazumasa Yoshida, Masatoshi Fujita, Nucleosome assembly and disassembly activity of GRWD1, a novel Cdt1-binding protein that promotes pre-replication complex formation, BIOCHIMICA ET BIOPHYSICA ACTA-MOLECULAR CELL RESEARCH, 10.1016/j.bbamcr.2016.08.008, 1863, 11, 2739-2748, 2016.11.
11. Nozomi Sugimoto, Kazumitsu Maehara, Kazumasa Yoshida, Shuhei Yasukouchi, Satoko Osano, Shinya Watanabe, Masahiro Aizawa, Takashi Yugawa, Tohru Kiyono, Hitoshi Kurumizaka, Yasuyuki Ohkawa, Masatoshi Fujita, Cdt1-binding protein GRWD1 is a novel histone-binding protein that facilitates MCM loading through its influence on chromatin architecture, NUCLEIC ACIDS RESEARCH, 10.1093/nar/gkv509, 43, 12, 5898-5911, 2015.07, 真核細胞染色体DNAはクロマチン構造をとっており、これは一般的に複製や転写に対して阻害的であり、反応にあわせてクロマチン動態を制御する必要がある。一方、DNA複製の最初のステップは、複製開始複合体(pre-RC)形成(ORC/CDC6/Cdt1によるMCM helicaseの装着反応)である。このMCMの装着反応はクロマチン上で高効率で起きる必要があり、その効率の低下はゲノムの不安定性を惹起することが知られている。しかしながら、高効率なMCM装着に必要なクロマチン動態を制御するヒストンシャペロンは不明なままであった。本論文では、我々が新規Cdt1結合蛋白として同定したGRWD1が、CDC6/Cdt1依存性に複製開始点に結合し、MCM装着の促進に働いている事を明らかにした。さらに、GRWD1は、その酸性アミノ酸ドメインを介してヒストンと結合し、ヒストンシャペロン活性を示すことが分かった。以上から、GRWD1はCDC6/Cdt1依存性に複製開始点に結合し、ヌクレオソーム構造を緩めることにより効率よいMCMローディングを促進していると言うモデルが考えられる。.
12. Makoto Ohira, Yuka Iwasaki, Chika Tanaka, Michitaka Kuroki, Naoki Matsuo, Tatsuhiko Kitamura, Masaki Yukuhiro, Hiroyuki Morimoto, Nisha Pang, Tohru Kiyono, Masahide Amemiya, Kozo Tanaka, Kazumasa Yoshida, Nozomi Sugimoto, Takashi Ohshima, Masatoshi Fujita, A novel anti-microtubule agent with carbazole and benzohydrazide structures suppresses tumor cell growth in vivo, Biochimica et Biophysica Acta (BBA) - General Subjects, 10.1016/j.bbagen.2015.04.013, 1850, 1676-1684, 2015.05.
13. Satoko Iwahori, Daisuke Kohmon, Junya Kobayashi, Yuhei Tani, Takashi Yugawa, Kenshi Komatsu, Tohru Kiyono, Nozomi Sugimoto, Masatoshi Fujita, ATM regulates Cdt1 stability during the unperturbed S phase to prevent re- replication, CELL CYCLE, 10.4161/cc.27274, 13, 3, 471-481, 2014.02, ATMはDNA損傷応答、特にDNA二重鎖切断(DSB)応答において中心的な役割を持っているPI3キナーゼ関連キナーゼである。その変異は、毛細血管拡張性小脳失調症(AT)の原因となる。しかし、ATMはDSB誘導時のみならず、通常の細胞周期においても何らかの機能を持っている可能性が考えられていた。Cdt1はDNA複製開始に必須の因子であり、ORC/CDC6と協調してMCMヘリカーゼをクロマチンにloadすることにより、複製開始複合体を形成する。一方、Cdt1は細胞周期S期になると分解抑制され、この制御は複製を1回のみ正確に行うために極めて重要である。本研究で我々は、ATMがこのCdt1のS期の分解制御に関与していることを明らかにした。ATMは、恐らくAkt-SCF-Skp2の経路を介して、Cdt1の適切な分解を促進している。本研究はATMの新規機能を明らかにすると共に、ATの病態理解を進める上でも重要であると考えられる。.
14. Naoki Nishimoto, Masanori Watanabe, Shinya Watanabe, Nozomi Sugimoto, Takashi Yugawa, Tsuyoshi Ikura, Osamu Koiwai, Tohru Kiyono, Masatoshi Fujita, Heterocomplex formation by Arp4 and beta-actin is involved in the integrity of the Brg1 chromatin remodeling complex, JOURNAL OF CELL SCIENCE, 10.1242/jcs.104349, 125, 16, 3870-3882, 2012.08, 筋肉の主要な蛋白質として知られているアクチンは、細胞運動や細胞増殖においても重要な役割を演じている。最近、このアクチンが細胞核内にも存在し、転写制御や細胞周期制御を通して細胞増殖制御に必須の役割を演じていることが示唆されつつあるが、その詳細は不明である。また、核内にはアクチンと相同性を持つアクチン関連蛋白質Arp4も存在するが、同様に機能や制御の詳細は不明なままであった。本研究では、アクチンとArp4が複合体(おそらく二量体)を形成することにより、幾つかの重要なクロマチンリモデリング因子複合体、HAT(ヒストンアセチル化酵素)複合体およびc-myc転写因子複合体(主要ながん遺伝子の一つ)の構成成分となり、それらの複合体の機能制御に関与していることを明らかにした。.
15. Sugimoto N, Yugawa T, Iizuka M, Kiyono T and Fujita M., Chromatin remodeler Sucrose Non-Fermenting 2 Homolog (SNF2H) is recruited onto DNA replication origins through interaction with Cdc10 protein-dependent transcript 1 (Cdt1) and promotes pre-replication complex formation., J. Biol. Chem. , 286: 39200-39210, 2011.11, DNA複製開始反応制御機構の解明は、細胞増殖制御の理解と言う点のみならず、新規抗がん剤開発を考える上でも重要である。Cdt1はORC/CDC6と協調して、複製ヘリカーゼMCM複合体を染色体へloadingする。Cdt1は強くこの反応を促進する。このCdt1によるMCM loading反応の促進において、ATP依存性クロマチンリモデリング因子SNF2Hが重要な役割を演じている事を明らかにし報告した。すなわち、(1)SNF2HはCdt1と物理的に相互作用し、これらに依存して複製開始点に結合する。(2)SNF2Hの複製開始点への結合はG1期で増加し、G2/M期で減少する。(3)SNF2HをsiRNAで抑制すると複製開始点でのMCM loadingが抑制される。以上から、SNF2HはCdt1との結合を介して複製開始点に集積し、ヌクレオソームの構造を制御することでMCMのクロマチン結合を制御しているとの仮説が考えられる。.
16. Yoshida K, Sugimoto N, Iwahori S, Yugawa T, Narisawa-Saito M, Kiyono T, Fujita M., CDC6 interaction with ATR regulates activation of a replication checkpoint in higher eukaryotic cells., J. Cell Sci., 123: 225-235, 2010.01.
17. Sugimoto N, Yoshida K, Tatsumi Y, Yugawa T, Narisawa-Saito M, Waga S, Kiyono T, Fujita M., Redundant and differential regulation of multiple licensing factors ensures prevention of rereplication in normal human cells. , J. Cell Sci. , 122: 1184-1191, 2009.04.
18. Sugimoto N, Kitabayashi I, Osano S, Tatsumi Y, Yugawa T, Narisawa-Saito M, Matsukage A, Kiyono T, Fujita, M., Identification of novel human Cdt1-binding proteins by a proteomics approach: Proteolytic regulation by APC/C-Cdh1., Mol. Biol. Cell , 19: 1007-1021, 2008.03.
19. Tatsumi Y, Ezura K, Yoshida K, Yugawa T, Narisawa-Saito M, Kiyono T, Ohta S, Obuse C, Fujita M., Involvement of human ORC and TRF2 in pre-replication complex assembly at telomeres, Genes Cells , 13: 1045-1059, 2008.10.
20. Mizushina Y, Takeuchi T, Hada T, Maeda N, Sugawara F, Yoshida H, Fujita M., The inhibitory action of SQDG (sulfoquinovosyl diacylglycerol) from spinach on Cdt1-geminin interaction., Biochimie , 90: 947-956, 2008.06.
21. Tatsumi Y, Sugimoto N, Yugawa T, Narisawa-Saito M, Kiyono T, Fujita M., Deregulation of Cdt1 induces chromosomal damage without rereplication and leads to chromosomal instability., J. Cell Sci. , 119: 3128-3140, 2006.08.
22. Nishitani H, Sugimoto N, Roukos V, Nakanishi Y, Saijo M, Obuse C, Tsurimoto T, Nakayama K-I, Nakayama K, Fujita M, Lygerous Z, Nishimoto T., Two E3 ubiquitin ligases, SCF-Skp2 and DDB1-Cul4, target human Cdt1 for proteolysis., EMBO J. , 25: 1126-1136, 2006.03.
23. Sugimoto N, Tatsumi Y, Tsurumi T, Matsukage A, Kiyono T, Nishitani H, Fujita M. , Cdt1 phosphorylation by cyclin A-dependent kinases negatively regulates its function without affecting geminin binding., J Biol. Chem. , 10.1074/jbc.M313175200, 279, 19, 19691-19697, 279: 19691-19697, 2004.05.
1. 杉本 のぞみ、前原 一満、大川 恭行、藤田 雅俊, Genome-wide analysis of the spatiotemporal regulation of firing and dormant replication origins in human cells, 第45回日本分子生物学会年会ワークショップ, 2022.12.
2. Hiroki Fujiyama, Takuya Takafuji, Tohru Kiyono, Kazumasa Yoshida, Nozomi Sugimoto, Masatoshi Fujita, Elucidation of tumor-suppressive mechanism of Ribosomal Protein S19 associated with Diamond-Blackfan anemia, 第79回日本癌学会学術総会 International Sessions, 2021.09.
3. 藤田雅俊, Ribosomal/nucleolar proteins that act as oncoproteins or antioncoproteins
, 熊本大学HIGOセミナー, 2020.11.
4. 杉本 のぞみ、藤田 成、辻田 沙伎、岩村 拓人、前原 一満、吉田 和真、大川 恭行、藤田 雅俊, Systematic elucidation of mechanisms underlying formation of licensed chromatin in human cells, 第42回日本分子生物学会年会ワークショップ, 2019.12.
5. Mitsunori Higa, Yukihiro Matsuda, Nozomi Sugimoto, Kazumasa Yoshida, Masatoshi Fujita, ORC recruitment by TRF2 contributes to telomere stability upon DNA replication stress, Cold Spring Harbor Laboratory Meeting on "Eukaryotic DNA replication and genome maintenance", 2019.09.
6. Masatoshi Fujita, Ribosomal/nucleolar proteins that act as oncogenes or ationcogenes, The 4th Japan-Taiwan Joint Symposium for Pharmaceutical Sciences, 2018.08.
7. Masatoshi Fujita, Nozomi Sugimoto, Kota Kayama, Shinya Watanabe, Masahiro Aizawa, Kazumasa Yoshida, GRWD1 counteracts p53 via the RPL11-MDM2 pathway and promotes tumorigenesis, 10th 3R International Symposium, 2016.11.
8. Kota kayama, Nozomi Sugimoto, Kazumasa Yoshida, Keiichi Nakayama, Tohru Kiyono, Masatoshi Fujita, GRWD1 negatively regulates p53 via the RP (ribosomal protein)-MDM2 pathway and promotes anchorage-independent growth, 第74回日本癌学会学術総会, 2015.10.
9. Nozomi Sugimoto, Kazumasa Yoshida, Kazumitsu Maehara, Yasuyuki Ohkawa, Masatoshi Fujita, Genome-wide analysis for spatiotemporal regulation of the pre-RC formation and firing in human cells, Cold Spring Harbor Laboratory Meeting on "Eukaryotic DNA replication and genome maintenance", 2015.09.
10. Nozomi Sugimoto, Masahiro Aizawa, Kazumasa Yoshida, Kazumitsu Maehara, Yasuyuki Ohkawa, Masatoshi Fujita, Genome-scale analysis for spatiotemporal regulation of the pre-RC formation and firing in human cells, International Symposium on Non-coding DNA and Chromosomal Integrity, 2015.08.
11. Nozomi Sugimoto, Kazumitsu Maehara, Kazumasa Yoshida, Shuhei Yasukouchi, Shinya Watanabe, Masahiro Aizawa, Tohru Kiyono, Hitoshi Kurumizaka, Yasuyuki Ohkawa, Masatoshi Fujita, Genome-wide relationship between pre-replication complex formation and chromatin status, The 9th 3R Symposium, 2014.11.
12. Nozomi Sugimoto, Kazumitsu Maehara, Shuhei Yasukouchi, Shinya Watanabe, Tohru Kiyono, Hitoshi Kurumizaka, Yasuyuki Ohkawa, Masatoshi Fujita, Cdt1-Binding Protein GRWD1 Is a Novel Histone Chaperone That Regulates Chromatin Structure and MCM Loading, Cold Spring Harbor Laboratory Meeting on "Eukaryotic DNA replication and genome maintenance", 2013.09.
13. Nozomi Sugimoto, 安河内 周平, 前原 一満, 清野 透, 胡桃坂仁志, Yasuyuki Ohkawa, Masatoshi Fujita, The Cdt1-binding protein GRWD1 is a novel histone chaperon involved in replication licensing and cell growth, 第35回日本分子生物学会年会 ワークショップ, 2012.12.
14. Nozomi Sugimoto, 安河内 周平, Shinya Watanabe, 清野 透, 胡桃坂仁志, Masatoshi Fujita, The Cdt1-binding protein GRWD1 is a novel histone chaperon involved in replication licensing, 第34回日本分子生物学会年会 ワークショップ, 2011.12.
Membership in Academic Society
  • The Pharmaceutical Society of Japan
  • The Japanese Society of Internal Medicine
  • Japanese Cancer Association
  • The Molecular Biology Society of Japan
Other Educational Activities
  • 2015.04.