1990 Fiscal Year Annual Research Report

増殖因子による遺伝子発現とDNA複製開始に至るシグナル伝達機構の研究

Research Project

Project/Area Number	02404086
Research Institution	The University of Tokyo
Principal Investigator	新井賢一東京大学, 医科学研究所, 教授 (00012782)
Co-Investigator(Kenkyū-buntansha)	村松正明東京大学医科学研究所, 教務職員 (50230008) 正井久雄東京大学医科学研究所, 助手 (40229349) 中山直樹東京大学医科学研究所, 助手 (80227967) 横田崇東京大学医科学研究所, 助教授 (50134622)
Keywords	増殖因子 / 遺伝子発現 / DNA複製 / 細胞周期 / シグナル伝達 / 発現クロ-ニング / 分子生物学
Research Abstract	活性化T細胞におけるGMーCSF遺伝子発現はCLE2/GC boxおよびCLE0を介して制御されている。前者にはNFーGM2とA1,A2,Bが結合するが、Jurkat細胞より精製されたNFーGM2はNFーκBに、A1はSP1に類似している。またCLE0に結合するタンパク質はetsファミリ-である可能性がある。ヒトILー3遺伝子のGCに富む領域はin vitroでTATA依存性転写を促進し、in vivo共形質導入法でp40^<tax>に著しく応答する。このGC領域をプロ-ブとして、その結合特異性と転写活性が相関するタンパク質複合体αを同定し、上流シグナルとの関連や基本的転写機構を活性化する分子機構について検討を加えた。また偽基質領域に変異を導入した構成的に活性なPKCではキナ-ゼ活性と生物学的活性に解離を認めたが、これらの変異PKCの遺伝子発現への効果についても検討した。造血因子の作用を研究するためにマウスILー3レセプタ-及びヒトGMーCSFレセプタ-のβ鎖の遺伝子の単離を行なった。さらにその下流のシグナル伝達について検索するため酵母のG1サイクリン遺伝子を動物細胞発現ベクタ-によりサイトカイン依存性プレB細胞および胚幹細胞中で発現する実験を進めている。プレB細胞についてはすでに完了しG1サイクリンの細胞周期、サイトカイン要求性、および細胞死に与える影響を検討し細胞サイズへの効果は比較的小さいが変化を観察した。酵母においては同様の強制発現によりG1→S期遷移が非制御的に促進されるため、G1期が実質的に無くなり細胞のサイズが小さくなることが観察されている。大腸菌におけるDNA複製においてはAサイトヘアピン上におけるdnaA,dnaB,dnaCタンパク質に依存したABCプライモソ-ムの形成機構を詳細に研究し、Aの結合、BCの導入、プライモソ-ムの移動の3段階を経て進行することを示した。またφX174型プライモソ-ムの構成要素であるn'タンパク質遺伝子を単離し、その構造決定と量産系を確立した。さらにpriA機能は染色体複製には必須ではないが、n'サイトを持つpBR322の複製には必要とされることを示した。これらの各種プライモソ-ムのR1プラスミド複製における役割についても検討を行なった。

Research Products
(19 results)

All Other

All Publications (19 results)

[Publications] Nishida,J.: "Deinition of GC rich motif as regulatory sequence of the human ILー3 gene:Coordinate regulation of the ILー3 gene by CLE2/GC box of the GMーCSF gene in T cell activation." Int Immunol.(1991)
[Publications] Hayashida,K.: "Molecular cloning of a second subunit of the human GMーCSF receptor:Resonsitution of a high affinity GMーCSF receptor by coーexpression of the low affinity GMーCSF receptor and an ILー3 receptorーlike molecule." Proc Natl Acad Sci U S A.87. 9655-9659 (1990)
[Publications] Itoh,N.: "Molecular cloning of a cDNA encoding the murine interleukinー3 receptor reveals a consensus motif for a new cytokine receptor gene family." Science. 247. 324-327 (1990)
[Publications] Gorman,D.M.: "Cloning and expression of a gene encoding an interleukin 3 receptorーlike protein:Identification of another member of the cytokine receptor gene family." Proc Natl Acad Sci U S A.87. 5459-5463 (1990)
[Publications] Sugimoto,K.: "Inducible and nonーinducible factors coーoperatively active the GMーCSF promoter by interacting with two adjacent DNA motifs." Int Immunol.2. 787-794 (1990)
[Publications] Arai,K.: "Cytokines;Cordinators of immune and inflammatory responses." Ann Rev Biochem. 59. 783-836 (1990)
[Publications] Tsuji,K.: "Effects of interleukinー3 and interleukinー4 on the development of connective tissueーtype mast cells:interleukinー3 supports their survival and interleukinー4 triggers and supports their proliferation synergistically with interleukinー3." Blood. 75. 421-427 (1990)
[Publications] Tsuji,K.: "Synergistic action of phorbol ester and interleukin 3 in the induction of connective tissue type mast cell proliferation." J Immunol. 144. 678-684 (1990)
[Publications] Xie,G.: "Chimeric opioid peptides:tools for identifying opioid receptor types." Proc Natl Acad Sci USA. 87. 3180-3184 (1990)
[Publications] Schreurs,J.: "Evidence for a low affinity interleukin 3 receptor." Growth Factors. 2. 221-234 (1990)
[Publications] Watanabe,S.: "Colony stimulating factors and cytokine receptor network." Current opinion in Biotechnology. (1991)
[Publications] Masai,H.: "Roles of φX174 type primosome and G4 type primaseーdependent primings in initiation of lagging and leading strand synthesis of DNA replication" The Journal of Biological Chemistry. 265. 15124-15133 (1990)
[Publications] Masai,H.: "The ABC primosome:A novel priming system employing dnaA,dnaB,dnaC and primase on a hairpin containing a dnaA box sequence" The Journal of Biological Chemistry. 265. 15134-15144 (1990)
[Publications] Lee,E.H.: "The priA gene encoding the primosomal replicativen' protein of Escherichia coli." Proc Natl Acad Sci USA. 87. 4620-4624 (1990)
[Publications] Nomura,N.: "Identification of eleven singleーstranded sequences (ssi) for priming of DNA replication in the F,R6K,R100 and ColE2 plasmids" Gene. (1991)
[Publications] Nomoto,S.: "Regulation of the yeast pheromone response pathway by G protein subunits." EMBO J. 9. 691-696 (1990)
[Publications] Matsumoto,K.: "Mating pheromone signal transduction in yeast." “Molecular and cellular biology of yeast and filamentous funji"UCLA Symposia. (1990)
[Publications] Yokota,T.: "Handbook of Experimental Pharmacology,Vol.95/I Peptide Growth.Factors and Their Receptors I." SpringerーVerlag, 31 (1990)
[Publications] Miyatake,S.: "Molecular biology of cytokines." Verlag Chemie International, (1991)

1990 Fiscal Year Annual Research Report

増殖因子による遺伝子発現とDNA複製開始に至るシグナル伝達機構の研究

Principal Investigator

新井 賢一 東京大学, 医科学研究所, 教授 (00012782)

Research Products

[Publications] Nishida,J.: "Deinition of GC rich motif as regulatory sequence of the human ILー3 gene:Coordinate regulation of the ILー3 gene by CLE2/GC box of the GMーCSF gene in T cell activation." Int Immunol.(1991)

[Publications] Hayashida,K.: "Molecular cloning of a second subunit of the human GMーCSF receptor:Resonsitution of a high affinity GMーCSF receptor by coーexpression of the low affinity GMーCSF receptor and an ILー3 receptorーlike molecule." Proc Natl Acad Sci U S A.87. 9655-9659 (1990)

[Publications] Itoh,N.: "Molecular cloning of a cDNA encoding the murine interleukinー3 receptor reveals a consensus motif for a new cytokine receptor gene family." Science. 247. 324-327 (1990)

[Publications] Gorman,D.M.: "Cloning and expression of a gene encoding an interleukin 3 receptorーlike protein:Identification of another member of the cytokine receptor gene family." Proc Natl Acad Sci U S A.87. 5459-5463 (1990)

[Publications] Sugimoto,K.: "Inducible and nonーinducible factors coーoperatively active the GMーCSF promoter by interacting with two adjacent DNA motifs." Int Immunol.2. 787-794 (1990)

[Publications] Arai,K.: "Cytokines;Cordinators of immune and inflammatory responses." Ann Rev Biochem. 59. 783-836 (1990)

[Publications] Tsuji,K.: "Effects of interleukinー3 and interleukinー4 on the development of connective tissueーtype mast cells:interleukinー3 supports their survival and interleukinー4 triggers and supports their proliferation synergistically with interleukinー3." Blood. 75. 421-427 (1990)

[Publications] Tsuji,K.: "Synergistic action of phorbol ester and interleukin 3 in the induction of connective tissue type mast cell proliferation." J Immunol. 144. 678-684 (1990)

[Publications] Xie,G.: "Chimeric opioid peptides:tools for identifying opioid receptor types." Proc Natl Acad Sci USA. 87. 3180-3184 (1990)

[Publications] Schreurs,J.: "Evidence for a low affinity interleukin 3 receptor." Growth Factors. 2. 221-234 (1990)

[Publications] Watanabe,S.: "Colony stimulating factors and cytokine receptor network." Current opinion in Biotechnology. (1991)

[Publications] Masai,H.: "Roles of φX174 type primosome and G4 type primaseーdependent primings in initiation of lagging and leading strand synthesis of DNA replication" The Journal of Biological Chemistry. 265. 15124-15133 (1990)

[Publications] Masai,H.: "The ABC primosome:A novel priming system employing dnaA,dnaB,dnaC and primase on a hairpin containing a dnaA box sequence" The Journal of Biological Chemistry. 265. 15134-15144 (1990)

[Publications] Lee,E.H.: "The priA gene encoding the primosomal replicativen' protein of Escherichia coli." Proc Natl Acad Sci USA. 87. 4620-4624 (1990)

[Publications] Nomura,N.: "Identification of eleven singleーstranded sequences (ssi) for priming of DNA replication in the F,R6K,R100 and ColE2 plasmids" Gene. (1991)

[Publications] Nomoto,S.: "Regulation of the yeast pheromone response pathway by G protein subunits." EMBO J. 9. 691-696 (1990)

[Publications] Matsumoto,K.: "Mating pheromone signal transduction in yeast." “Molecular and cellular biology of yeast and filamentous funji"UCLA Symposia. (1990)

[Publications] Yokota,T.: "Handbook of Experimental Pharmacology,Vol.95/I Peptide Growth.Factors and Their Receptors I." SpringerーVerlag, 31 (1990)

[Publications] Miyatake,S.: "Molecular biology of cytokines." Verlag Chemie International, (1991)

新井賢一東京大学, 医科学研究所, 教授 (00012782)