1991 年度実績報告書

ゲノム複製開始の制御

研究課題

研究課題/領域番号	03261102
研究機関	大阪大学
研究代表者	吉川寛大阪大学, 医学部, 教授 (70019876)
研究分担者	山口和男金沢大学, 遺伝子実験施設, 教授 (00019879) 堀内嵩基礎生物学研究所, 教授 (60108644) 堀内賢介国立遺伝研, 細胞遺伝, 教授 (70219210) 今本文男京都薬科大, 生命薬学研, 教授 (00029761) 小笠原直毅大阪大学, 医学部, 講師 (10110553)
キーワード	ゲノム複製 / 複製開始点 / ARS配列 / 複製遺伝子 / 複製酵素
研究概要	プラスミド、ウイルス、細菌、動植物細胞を対象にゲノム複製の開始とその制御の分子機構の解析を行った(計画研究13、公募研究6)。対象にとらわれず、研究のレベル毎に成果をまとめた。 1.開始調節の分子機構の解明:開始蛋白とその結合配列との相互作用が開始の頻度と時間決定に重要であることが、プラスミド(山口、犬塚)と枯草菌(小笠原)で明らかになり相互作用の詳細がin vitroで解析された。また開始の微調節に働く、転写(小川)、DNA高次構造(堀内、今本)、クロマチン(石見)の解析を進めた。 2.複製開始点の分離同定:自己複製能を持つ配列(ARS)の分離を系統的に行い、新しくマイコプラズマ(宮田)とタバコプラスチドのoriC(武田)、酵母第VI染色体全ARS(吉川)、ヒト染色体のARS(升方)を分離した。またin vitro合成系によるヒト開始点の同定(釣本)、Xenopus染色体上の開始点の分布解明(四宮)のための方法を開発した。 3.開始の遺伝因子の探索:真核生物では開始に働く因子は未知である。酵母を材料に変異細胞の解析によって開始に関与する遺伝子を分離しその作用点の解析を進めた(菊地)。 4.複製開始の装置の解析:in vitro合成が開発されている大腸菌について複製開始に必須な蛋白複合体の構成と機能を解析した(正井)。研究が遅れている真核生物については前段階として複製の必須構成酵素の解析を酵母(杉野)、ヒト細胞(吉田)で格段に進めた。 5.各々の研究は順調に進展し、原核生物については複製開始と調節に関与する分子的実体がほぼ網羅された。今後これらの因子間の相互作用、超複合体の形成と細胞内局在を系統的に研究する道が開けた。真核生物では分離された酵母ARSの染色体複製における役割を中心に哺乳類細胞での開始点の実体を明かにすることが期待できる。

研究成果

(23件)

すべてその他

すべて文献書誌 (23件)

[文献書誌] Iwasaki,T.: "The direct cloning of the yeast genome using the gap-filling method and the complete physical mapping of Saccharomyces cerevisiae chromosome VI" Gene. 109. 81-87 (1991)
[文献書誌] Ishimi,Y.: "Identification and molecular cloning of yeast homolog of nucleosome assembly protein I which facilitates nucleosome assembly in vitro" J.Biol.Chem.266. 7025-7029 (1991)
[文献書誌] Ishimi,Y.: "Replication of the simian virus 40 chromosome with purified proteins" J.Biol.Chem.266. 16141-16418 (1991)
[文献書誌] Sugasawa,K.: "Non-conservative segregation of parental nucleosome during simian virus 40 chromosome replication in vitro" Proc.Natl.Acad.Sci.USA. 89. (1992)
[文献書誌] Kano,Y.: "Participation of the histone-like protein HU and of IHF in minichromosomal maintenance in Escherichia coli" Gene. 103. 25-30 (1991)
[文献書誌] Yoshikawa,H.: "Structure and function of DnaA and DnaA-box in eubacteria:evolutionary relationship of bacterial replication origins" Mol.Microbiol.5. 2589-2597 (1991)
[文献書誌] Moriya,S.: "Cloning of autonomously replicating sequence (ars) from the Bacillus subtilis chromosome" Mol.Microbiol.6. 309-315 (1992)
[文献書誌] Ogasawara,N.: "Genes and their organization in replication origin region of bacterial chromosome" Mol.Microbiol.6. (1992)
[文献書誌] Shinomiya,T.: "Analysis of chromosomal replicons in early embryos of Drosophila melanogaster by two-dimensional gel electrophoresis" Nucleic Acids Res.14. 3935-3941 (1991)
[文献書誌] Araki,H.: "Cloning DPB3,the gene encoding the third subunit of DNA polymerase II of Saccharomyces cerevisiae" Nucleic Acids Res.19. 4867-4872 (1991)
[文献書誌] Morrison,A.: "Eukaryotic DNA polymerase amino acid sequence required for 3'-5' exonuclease activity" Proc.Natl.Acad.Sci.USA. 88. 9473-9477 (1991)
[文献書誌] Araki,H.: "DNA polymerase II,the probable homolog of mammalian DNA polymerase ε,replicates chromosomal DNA in yeast Saccharomyces cerevisiae" EMBOJ.11. (1992)
[文献書誌] Higashitani,N.: "SOS induction in Escherichia coli by infection with mutant filamentous phage that are defective in initiation of complementary strand DNA synthesis" J.Bacteriol.174. (1992)
[文献書誌] Hidaka,M.: "A new identified DNA replication terminus site,TerE,on the Escherichia coli chromosome" J.Bacteriol.173. 391-393 (1991)
[文献書誌] Hidaka,M.: "Termination complex in Escherichia coli inhibits SV40 DNA replication in vitro by impeding the action of T antigen helicase" J.Biol.Chem.267. (1992)
[文献書誌] Kobayashi,T.: "Identification of a site required for DNA replication fork blocking activity in the rRNA gene cluster in Saccharomyces cerevisiae" Mol.Gen.Genet.(1992)
[文献書誌] Maki,H.: "A strong mutator effect caused by an amino acid change in the α subunit of DNA polymerase III of Escherichia coli" J.Biol.Chem.266. 5055-5061 (1991)
[文献書誌] Mo,J.-Y.: "Mutational specificity of the dnaE173 mutator associated with a defect in the catalytic subunit of DNA polymerse III of Escherichia coli" J.Mol.Biol.222. 925-936 (1991)
[文献書誌] Maki,H.: "MutT protein specifically hydrolyzes 8-oxod GTP,a potent mutagenic substrate for DNA synthesis" Nature. 355. 273-275 (1992)
[文献書誌] Nomura,N.: "Identification of eleven single-strand initiation sequences (ssi) for priming of DNA replication in the F,R6K,R100 and ColE2 plasmids" Gene. 108. 15-22 (1991)
[文献書誌] Fueki,T.: "Exonuclease III promotes in vitro binding of the replication initiator protein of pSC101 to the repeated sequences in the ori region" Biochem.Biophys.Acta. 1089. 325-330 (1991)
[文献書誌] Fueki,T.: "A single-stranded region located downstream of the repeated sequence in the ori region of pSC101 is required for binding of the Rep protein in vitro" J.Biochem.110. 775-779 (1991)
[文献書誌] Shioda,M.: "Stimulation of DNA polymerase α activity by microtuble-associated proteins" Biochemistry. 30. 11403-11412 (1991)

1991 年度 実績報告書

ゲノム複製開始の制御

研究代表者

吉川 寛 大阪大学, 医学部, 教授 (70019876)

研究成果

[文献書誌] Iwasaki,T.: "The direct cloning of the yeast genome using the gap-filling method and the complete physical mapping of Saccharomyces cerevisiae chromosome VI" Gene. 109. 81-87 (1991)

[文献書誌] Ishimi,Y.: "Identification and molecular cloning of yeast homolog of nucleosome assembly protein I which facilitates nucleosome assembly in vitro" J.Biol.Chem.266. 7025-7029 (1991)

[文献書誌] Ishimi,Y.: "Replication of the simian virus 40 chromosome with purified proteins" J.Biol.Chem.266. 16141-16418 (1991)

[文献書誌] Sugasawa,K.: "Non-conservative segregation of parental nucleosome during simian virus 40 chromosome replication in vitro" Proc.Natl.Acad.Sci.USA. 89. (1992)

[文献書誌] Kano,Y.: "Participation of the histone-like protein HU and of IHF in minichromosomal maintenance in Escherichia coli" Gene. 103. 25-30 (1991)

[文献書誌] Yoshikawa,H.: "Structure and function of DnaA and DnaA-box in eubacteria:evolutionary relationship of bacterial replication origins" Mol.Microbiol.5. 2589-2597 (1991)

[文献書誌] Moriya,S.: "Cloning of autonomously replicating sequence (ars) from the Bacillus subtilis chromosome" Mol.Microbiol.6. 309-315 (1992)

[文献書誌] Ogasawara,N.: "Genes and their organization in replication origin region of bacterial chromosome" Mol.Microbiol.6. (1992)

[文献書誌] Shinomiya,T.: "Analysis of chromosomal replicons in early embryos of Drosophila melanogaster by two-dimensional gel electrophoresis" Nucleic Acids Res.14. 3935-3941 (1991)

[文献書誌] Araki,H.: "Cloning DPB3,the gene encoding the third subunit of DNA polymerase II of Saccharomyces cerevisiae" Nucleic Acids Res.19. 4867-4872 (1991)

[文献書誌] Morrison,A.: "Eukaryotic DNA polymerase amino acid sequence required for 3'-5' exonuclease activity" Proc.Natl.Acad.Sci.USA. 88. 9473-9477 (1991)

[文献書誌] Araki,H.: "DNA polymerase II,the probable homolog of mammalian DNA polymerase ε,replicates chromosomal DNA in yeast Saccharomyces cerevisiae" EMBOJ.11. (1992)

[文献書誌] Higashitani,N.: "SOS induction in Escherichia coli by infection with mutant filamentous phage that are defective in initiation of complementary strand DNA synthesis" J.Bacteriol.174. (1992)

[文献書誌] Hidaka,M.: "A new identified DNA replication terminus site,TerE,on the Escherichia coli chromosome" J.Bacteriol.173. 391-393 (1991)

[文献書誌] Hidaka,M.: "Termination complex in Escherichia coli inhibits SV40 DNA replication in vitro by impeding the action of T antigen helicase" J.Biol.Chem.267. (1992)

[文献書誌] Kobayashi,T.: "Identification of a site required for DNA replication fork blocking activity in the rRNA gene cluster in Saccharomyces cerevisiae" Mol.Gen.Genet.(1992)

[文献書誌] Maki,H.: "A strong mutator effect caused by an amino acid change in the α subunit of DNA polymerase III of Escherichia coli" J.Biol.Chem.266. 5055-5061 (1991)

[文献書誌] Mo,J.-Y.: "Mutational specificity of the dnaE173 mutator associated with a defect in the catalytic subunit of DNA polymerse III of Escherichia coli" J.Mol.Biol.222. 925-936 (1991)

[文献書誌] Maki,H.: "MutT protein specifically hydrolyzes 8-oxod GTP,a potent mutagenic substrate for DNA synthesis" Nature. 355. 273-275 (1992)

[文献書誌] Nomura,N.: "Identification of eleven single-strand initiation sequences (ssi) for priming of DNA replication in the F,R6K,R100 and ColE2 plasmids" Gene. 108. 15-22 (1991)

[文献書誌] Fueki,T.: "Exonuclease III promotes in vitro binding of the replication initiator protein of pSC101 to the repeated sequences in the ori region" Biochem.Biophys.Acta. 1089. 325-330 (1991)

[文献書誌] Fueki,T.: "A single-stranded region located downstream of the repeated sequence in the ori region of pSC101 is required for binding of the Rep protein in vitro" J.Biochem.110. 775-779 (1991)

[文献書誌] Shioda,M.: "Stimulation of DNA polymerase α activity by microtuble-associated proteins" Biochemistry. 30. 11403-11412 (1991)

1991 年度実績報告書

吉川寛大阪大学, 医学部, 教授 (70019876)