1997 Fiscal Year Annual Research Report

真核生物の遺伝的組換えシステムの研究

Research Project

Project/Area Number	06101003
Research Institution	Osaka University
Principal Investigator	小川英行大阪大学, 大学院・理学研究科, 教授 (70028207)
Co-Investigator(Kenkyū-buntansha)	柴田武彦理化学研究所, バイオデザイン研究グループ, 研究者 (70087550) 小川智子国立遺伝学研究所, 細胞遺伝研究系, 教授 (80028208) 坪内英生大阪大学, 大学院・理学研究科, 助手 (20283822)
Keywords	出芽酵母 / 減数分裂期組換え / DNA二重鎖切断修復 / MRE11,RAD50 & XRS2遺伝子 / MER3遺伝子 / MRE2とスプライシング / Rad51とRad52の相互作用 / RPA
Research Abstract	今年度の主要な成果は次の通である。 (1)Mre11蛋白質の機能解析:小川英行、小川智子減数分裂期組換えの開始とDNA二重鎖切断の修復に、Mre11、Xrs2、Rad50蛋白質が関与する。精製したMre11蛋白質はssDNAエンドヌクレアーゼ活性を持ち、Rad50及びXrs2と結合する。またMre11の各種欠失蛋白質の解析から、ヌクレアーゼ活性、Rad50との相互作用部位、DNA結合領域、減数分裂期特異的蛋白質との相互作用部位を決定した。別のドメインのmre11変異同士は、互いに相補できることが明らかになった。 (2)MER3の機能:小川英行 MRE2遺伝子は、MER2遺伝子pre-mRNAのスプライシングを可能にして、減数分裂期組換えの開始に関与することを明らかにした。しかも新しい遺伝子MER3 pre-mRNAのスプライシングにも関与して居ることがわかった。Mer3蛋白質はヘリカーゼに共通のモチーフを持ち、交差型組換え体形成に密接に関与する。 (3)Rad51組換え反応に於けるRad52の機能:小川智子。 Rad51蛋白質の相補鎖移行反応とssDNA依存ATPase活性の強さは、Rad51がssDNAに結合して作る特殊なラセン構造体の形成能に関わっている。ssDNA結合蛋白質RPAはssDNAの2次構造を解消してRad51の構造体形成を促進する。しかし先にRPAがssDNAに結合するとRad51のDNA結合活性が抑えられる。ところがこの阻害をRad52が解消する。その機構は、ssDNAに結合したRPAはRad52と結合するが、Rad52はRad51と結合して、Rad51の構造体形成を促進する。Rad52とRPA、またはRad51との相互作用には種特異性があり、同種のものでしかRad51の活性の促進は起こらない。また、3種の相互作用部位を欠失した変異蛋白質では促進が観察できない。

Research Products

(18 results)

All Other

All Publications (18 results)

[Publications] Tsubouchi,H.: "A novel mre11 mutation impairs processing of double-strand breaks of DNA during both mitosis and meiosis." Mol.Cell.Biol.18. 260-268 (1998)
[Publications] Ohta,K.: "Mutations in the MRE11,RAD50,XRS2,and MRE2 genes alter chromatin configuration at meiotic DNA double-stranded break sites in premeiotic and meiotic cells." Proc.Natl.Acad.Sci.USA. 95. 646-651 (1998)
[Publications] Sonoda,E.: "Rad51-deficient vertebrate cells accumulate chromosomal breaks prior to cell death." EMBOJ.17. 598-608 (1998)
[Publications] Namsaraev,E.A.: "Biochemical basis of hyper-recombinogenic activity of Pseudomonas aeruginosa RecA protein in Escherichia coli cells." Mol.Microbiol.27. 727-738 (1998)
[Publications] Shinohara A.: "Stimulation of by Rad52 of yeast Rad51-mediated Recombination." Nature. 391. 404-407 (1998)
[Publications] Shinohara A.: "Rad51-independet Homologous Recombination Mediated by Rad52." Genes to Cells. 3 (in press). (1998)
[Publications] Shinohara A.: "Rad51-independet Homologous Recombination Mediated by Rad52." Genes to Cells. (in press). (1998)
[Publications] Ohta,K.: "Mutations in the MRE11,RAD50,XRS2,and MRE2 genes alter chromatin configurations at meiotic DNA double-stranded break sites in premeiotic and meiotic cells." Proc.Natl.Acad.Sci.USA. 95. 646-651 (1998)
[Publications] Nakagawa,T.: "Involvement of the MRE2 gene of yeast in formation of meiosis-specific double-strand breaks and crossover recombination through RNA splicing." Genes Cells. 2. 65-79 (1997)
[Publications] Alexseyev,A.A.: "A recombinational defect in the C-terminal domain of Escherichia coli RecA2278-5 protein is compensated by protein binding to ATP." Mol.Microbiol.23. 255-265 (1997)
[Publications] Mizuta,R.: "RAB22 and RAB163/mouse BRCA2 : Proteins that specifically interact with the Rad51 protein." Proc.Natl.Acad.Sci.USA. 94. 6927-6932 (1997)
[Publications] Shinohara,M.: "Characterization of the roles of the Saccharomyces cerevisiae RAD54 gene and a homologue of RAD54,RDH/TID1,in mitosis and meiosis." Genetics. 147. 1545-1556 (1997)
[Publications] Shinohara A.: "S.cerevisiae recA homologues RAD51 and DMC1 have both distinct and overlapping roles in meiotic recombination." Genes to Cells. 2. 615-629 (1997)
[Publications] Mizuno,K.-i.: "The meiotic recombination hotspot created by the single base substitution,ade6-M26,results in remodeling of chromatin structure in fission yeast." Genes Dev.11. 876-886 (1997)
[Publications] Aihara,H.: "An interaction between a specified surface of the C-terminal domain of RecA protein and double-stranded DNA for homologous pairing." Mol.Biol.274. 213-221 (1997)
[Publications] Nishinaka,T.: "An extended DNA structure through deoxyribose-base stacking induced by RecA protein." Proc.Natl.Acad.Sci.USA. 94. 6623-6628 (1997)
[Publications] 小川智子: "DNAとその結合タンパク質複合対の観察法." 細胞工学. 16. 1054-1069 (1997)
[Publications] 小川智子: "組換え機構、シリーズ・ニューバイオフィジックス(日本生物物理学会編)第2巻遺伝子の構造生物学,嶋本伸雄,郷通子編集" 共立出版., 16 (1997)

1997 Fiscal Year Annual Research Report

真核生物の遺伝的組換えシステムの研究

Principal Investigator

小川 英行 大阪大学, 大学院・理学研究科, 教授 (70028207)

Research Products

[Publications] Tsubouchi,H.: "A novel mre11 mutation impairs processing of double-strand breaks of DNA during both mitosis and meiosis." Mol.Cell.Biol.18. 260-268 (1998)

[Publications] Ohta,K.: "Mutations in the MRE11,RAD50,XRS2,and MRE2 genes alter chromatin configuration at meiotic DNA double-stranded break sites in premeiotic and meiotic cells." Proc.Natl.Acad.Sci.USA. 95. 646-651 (1998)

[Publications] Sonoda,E.: "Rad51-deficient vertebrate cells accumulate chromosomal breaks prior to cell death." EMBOJ.17. 598-608 (1998)

[Publications] Namsaraev,E.A.: "Biochemical basis of hyper-recombinogenic activity of Pseudomonas aeruginosa RecA protein in Escherichia coli cells." Mol.Microbiol.27. 727-738 (1998)

[Publications] Shinohara A.: "Stimulation of by Rad52 of yeast Rad51-mediated Recombination." Nature. 391. 404-407 (1998)

[Publications] Shinohara A.: "Rad51-independet Homologous Recombination Mediated by Rad52." Genes to Cells. 3 (in press). (1998)

[Publications] Shinohara A.: "Rad51-independet Homologous Recombination Mediated by Rad52." Genes to Cells. (in press). (1998)

[Publications] Ohta,K.: "Mutations in the MRE11,RAD50,XRS2,and MRE2 genes alter chromatin configurations at meiotic DNA double-stranded break sites in premeiotic and meiotic cells." Proc.Natl.Acad.Sci.USA. 95. 646-651 (1998)

[Publications] Nakagawa,T.: "Involvement of the MRE2 gene of yeast in formation of meiosis-specific double-strand breaks and crossover recombination through RNA splicing." Genes Cells. 2. 65-79 (1997)

[Publications] Alexseyev,A.A.: "A recombinational defect in the C-terminal domain of Escherichia coli RecA2278-5 protein is compensated by protein binding to ATP." Mol.Microbiol.23. 255-265 (1997)

[Publications] Mizuta,R.: "RAB22 and RAB163/mouse BRCA2 : Proteins that specifically interact with the Rad51 protein." Proc.Natl.Acad.Sci.USA. 94. 6927-6932 (1997)

[Publications] Shinohara,M.: "Characterization of the roles of the Saccharomyces cerevisiae RAD54 gene and a homologue of RAD54,RDH/TID1,in mitosis and meiosis." Genetics. 147. 1545-1556 (1997)

[Publications] Shinohara A.: "S.cerevisiae recA homologues RAD51 and DMC1 have both distinct and overlapping roles in meiotic recombination." Genes to Cells. 2. 615-629 (1997)

[Publications] Mizuno,K.-i.: "The meiotic recombination hotspot created by the single base substitution,ade6-M26,results in remodeling of chromatin structure in fission yeast." Genes Dev.11. 876-886 (1997)

[Publications] Aihara,H.: "An interaction between a specified surface of the C-terminal domain of RecA protein and double-stranded DNA for homologous pairing." Mol.Biol.274. 213-221 (1997)

[Publications] Nishinaka,T.: "An extended DNA structure through deoxyribose-base stacking induced by RecA protein." Proc.Natl.Acad.Sci.USA. 94. 6623-6628 (1997)

[Publications] 小川 智子: "DNAとその結合タンパク質複合対の観察法." 細胞工学. 16. 1054-1069 (1997)

[Publications] 小川智子: "組換え機構、シリーズ・ニューバイオフィジックス(日本生物物理学会編)第2巻 遺伝子の構造生物学,嶋本伸雄,郷通子編集" 共立出版., 16 (1997)

小川英行大阪大学, 大学院・理学研究科, 教授 (70028207)

[Publications] 小川智子: "DNAとその結合タンパク質複合対の観察法." 細胞工学. 16. 1054-1069 (1997)

[Publications] 小川智子: "組換え機構、シリーズ・ニューバイオフィジックス(日本生物物理学会編)第2巻遺伝子の構造生物学,嶋本伸雄,郷通子編集" 共立出版., 16 (1997)