1996 Fiscal Year Annual Research Report

染色体複製開始の制御機構

Research Project

Project/Area Number	08277101
Research Category	Grant-in-Aid for Scientific Research on Priority Areas
Research Institution	Osaka University
Principal Investigator	滝澤温彦大阪大学, 大学院・理学研究科, 助手 (60154944)
Co-Investigator(Kenkyū-buntansha)	白髭克彦奈良先端科学技術大学院大学, バイオサイエンス研究科, 助手 (90273854) 正井久雄東京大学, 医科学研究所・分子生物学研究部, 助教授 (40229349) 升方久夫大阪大学, 大学院・理学研究科, 助教授 (00199689) 四宮知行三菱化学生命科学研究所, 核機能プロジェクト, 主任研究員松影昭夫愛知県がんセンター研究所, 生物学部, 部長 (90019571)
Keywords	DNA複製開始領域 / DNA複製開始蛋白質 / MCM / P1ファミリー蛋白質 / CDC7類似キナーゼ / 複製関連遺伝子 / 転写調節因子 / DREF
Research Abstract	複製開始領域の同定・単離とその特性分裂酵母第2染色体上に同定した自立複製配列を用いて複製開始必須域を解析し、自律複製能を持つ約1kbのDNA配列のなかで100bp前後の3つの必須領域を同定し、アデニストレッチが複製に必須であることを明らかにした。ショウジョウバエの培養細胞を用いて同定したDNAポリメラーゼα遺伝子の下流にある複製開始領域でもATに富む配列が多数存在することから複製開始に必要な領域に共通性があることが示唆される。出芽酵母の第6染色体に同定された複製開始領域ではS期における複製開始の時期が異なること、複製開始のタイミングが複製のチェック機構によって制御されていることが示された。ショウジョウバエでは発生の進行によって複製開始領域が変化することと合わせて考えると染色体の構造が複製制御に重要な働きをしていることが示唆される。複製開始蛋白質の同定とその作用機構アフリカツメガエル卵無細胞DNA複製系を用いてXMCM3を含む6種類のMCM/P1ファミリーに属する蛋白質複合体が複製のライセンス化に必須であることを明らかにした。また特定のMCM蛋白質が複製反応の開始過程に関わっていることを見い出した。MCM2,3,7蛋白質は複製開始に必須なCDC7のヒト類似キナーゼでリン酸化されることが明らかにされ、さらにCDC7の活性が細胞周期のエンジンであるcdkキナーゼによって調節されている可能性が示され、細胞周期の制御と複製開始蛋白質の研究に新たな接点が見い出された。複製関連遺伝子群の発現制御機構 DNA複製関連遺伝子の転写制御に重要な機能を果たしていると考えられる転写調節因子DREFが細胞増殖に関わる癌遺伝子や細胞周期関連遺伝子の転写調節にも関わっていること、複製関連遺伝子の転写調節にはE2FやDREF以外の転写因子が関与すること、DREFが細胞増殖や分化に必須な因子であることを明らかにした。

Research Products
(15 results)

All Other

All Publications (15 results)

[Publications] Kubota,Y.: "Licensing of DNA replication by a multi-protein complex of MCM/P1 proteins in Xenopus eggs." EMBO J.in press.
[Publications] Thommes,P.: "The RLF-M component of the replication licensing system forms complexes containing all six MCM/Pl polypeptides." EMBO J.in press.
[Publications] Yamaguchi,M.: "Roles of muliple promoter elements of the proliferating cell unclear antigen gene during Drosophila development." Genes to Cells. 1. 47-58 (1996)
[Publications] Hirose,F.: "Isolation and characterization of cDNA for DREF,a promoter-activating factor for Drosophila DNA replication-related genes.," J.Biol.Chem.271. 3930-3937 (1996)
[Publications] Takahashi,Y.: "DNA replication-related elements cooperate to enhance promoter activity of the Drosophila DNA polymerase a 73-kDa subunit gene." J.Biol.Chem.271. 14541-14547 (1996)
[Publications] Ohno,K.: "Transcriptional regulation of the Drosophila CycA gene by the DNA replication-related element(DRE) and DRE-binding factor(DREF)." Nucleic Acids Res.24. 3942-3946 (1996)
[Publications] Ryu,J.-R.: "Trancriptional regulation of the Drosophila raf protooncogene by the DNA replication-related element (DRE)/DRE-binding factor(DREF) system." Nucleic Acids Res.,in press.
[Publications] Masai,H.: "Mecahnisms of primer RNA synthesis and D-loop/R-loop-dependent DNA replication of Escherichia coli" Biochimie in press.
[Publications] Ohotoshi,A.: "Analyses of S.cerevisiae Cdc7 kinase point mutants:dominant negative inhibition of DNA replication by overexpression of non-functional Cdc7 protins" Mol.Gen.Genet.in press.
[Publications] Ohotoshi,A.: "Genetic interactions between CDC7 and CDC28:growh inhibition of cdc28-1N by cdc7 point mutants" Genes to Cells. 1. 895-904 (1996)
[Publications] Ikeda,M.: "A rapid and convenient method for screening of DNA binding and protein-interacting molecules by the use of a GST-fusion protein library" Gene. 181. 167-171 (1996)
[Publications] Ikeda,M.: "CTBP-1/RBP-1 of Saccharomyces cerevisiae protein which binds to T-rich single-stranded DNA containing the 11 base-pair core seqeunce of autonomously replicating seqence,is apolydeoxypyrimidine binding protein" Eur.J.Biochem.238. 38-47 (1996)
[Publications] Masai,H.: "DnaA-and PriA-dependent primosomes:two distinct replication complexes for replication of Escherichia coli chromosome" Frontiers in Bioscience. 1. 49-59 (1996)
[Publications] Obuse,C.: "A replication-enhancing element with transcriptional silencing activity in autonomously replicating human chromosomal fragments" Mol.Biol.Cell. 7. 43-55 (1996)
[Publications] Okuno,Y.: "Identification of a predominant replication origin in fission yeast" Nucleic Acids Res.25. 530-536 (1997)

1996 Fiscal Year Annual Research Report

染色体複製開始の制御機構

Principal Investigator

滝澤 温彦 大阪大学, 大学院・理学研究科, 助手 (60154944)

Research Products

[Publications] Kubota,Y.: "Licensing of DNA replication by a multi-protein complex of MCM/P1 proteins in Xenopus eggs." EMBO J.in press.

[Publications] Thommes,P.: "The RLF-M component of the replication licensing system forms complexes containing all six MCM/Pl polypeptides." EMBO J.in press.

[Publications] Yamaguchi,M.: "Roles of muliple promoter elements of the proliferating cell unclear antigen gene during Drosophila development." Genes to Cells. 1. 47-58 (1996)

[Publications] Hirose,F.: "Isolation and characterization of cDNA for DREF,a promoter-activating factor for Drosophila DNA replication-related genes.," J.Biol.Chem.271. 3930-3937 (1996)

[Publications] Takahashi,Y.: "DNA replication-related elements cooperate to enhance promoter activity of the Drosophila DNA polymerase a 73-kDa subunit gene." J.Biol.Chem.271. 14541-14547 (1996)

[Publications] Ohno,K.: "Transcriptional regulation of the Drosophila CycA gene by the DNA replication-related element(DRE) and DRE-binding factor(DREF)." Nucleic Acids Res.24. 3942-3946 (1996)

[Publications] Ryu,J.-R.: "Trancriptional regulation of the Drosophila raf protooncogene by the DNA replication-related element (DRE)/DRE-binding factor(DREF) system." Nucleic Acids Res.,in press.

[Publications] Masai,H.: "Mecahnisms of primer RNA synthesis and D-loop/R-loop-dependent DNA replication of Escherichia coli" Biochimie in press.

[Publications] Ohotoshi,A.: "Analyses of S.cerevisiae Cdc7 kinase point mutants:dominant negative inhibition of DNA replication by overexpression of non-functional Cdc7 protins" Mol.Gen.Genet.in press.

[Publications] Ohotoshi,A.: "Genetic interactions between CDC7 and CDC28:growh inhibition of cdc28-1N by cdc7 point mutants" Genes to Cells. 1. 895-904 (1996)

[Publications] Ikeda,M.: "A rapid and convenient method for screening of DNA binding and protein-interacting molecules by the use of a GST-fusion protein library" Gene. 181. 167-171 (1996)

[Publications] Ikeda,M.: "CTBP-1/RBP-1 of Saccharomyces cerevisiae protein which binds to T-rich single-stranded DNA containing the 11 base-pair core seqeunce of autonomously replicating seqence,is apolydeoxypyrimidine binding protein" Eur.J.Biochem.238. 38-47 (1996)

[Publications] Masai,H.: "DnaA-and PriA-dependent primosomes:two distinct replication complexes for replication of Escherichia coli chromosome" Frontiers in Bioscience. 1. 49-59 (1996)

[Publications] Obuse,C.: "A replication-enhancing element with transcriptional silencing activity in autonomously replicating human chromosomal fragments" Mol.Biol.Cell. 7. 43-55 (1996)

[Publications] Okuno,Y.: "Identification of a predominant replication origin in fission yeast" Nucleic Acids Res.25. 530-536 (1997)

滝澤温彦大阪大学, 大学院・理学研究科, 助手 (60154944)