Regulation of cell cycle in the somatic cells

体细胞细胞周期的调节

• 批准号: 08102008
• 负责人: NISHIMOTO Takeharu
• 金额: $ 197.12万
• 依托单位: KYUSHU UNIVERSITY
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Specially Promoted Research
• 财政年份: 1996
• 资助国家: 日本
• 起止时间: 1996 至 1999
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-08102008/
• 关键词: DAD1; HCF; RCC1; Ran; RanBP1; RanBP2; Mog1; cell cycle; Yrb2; アポトーシス; GTR1; DIS3; RanBPM; YRB2; MOG1; Rcclp; Gtrlp; RagA; Yrb2p; Crmlp; Cdc25B; アポプトーシス

This project is comprised of three parts, (1) the isolation of temperature-sensitive (ts) mutants for cell growth from hamster BHK21 cell line, (2) the identification of mutated genes in those mutants, and (3) the characterization of cloned mutated genes. I will review for each part. Part 1. Previously, I have established a method to isolate ts mutants of hamster BHK21 cell line. However, a majority of those mutants have a mutation in the X chromosome where a haploid dose of gene is expressed. In this project, I tried to isolate ts mutants of the autosome using a drug, benomyl. Finally, we isolated ts mutants belonging to a different complementation group. Part 2. We have identified mutated genes of ts mutants of hamster BHK21 cell line, which are genes encoding HCF (host cell factor for herpes virus proliferation), histidyl-tRNA synthetase and lysyl-tRNA synthetase, respectively. Part 3. I will review for each genes which have been cloned using ts mutants of hamster BHK21 cell line. H … More CF : Based on our finding, HCF was approved to be essential for host cell proliferation as well. The mutant defective in HCF gene, tsBN67, arrest in G0 phase after completion of the G1, S, G2 and M phase at the nonpermissive temperature. Thus, this mutant will be useful for characterizing the G0 phase at the molecular level. Aminoacyl-tRNA synthetase : mutants defective in histidyl- and lysyl-tRNA synthetases rapidly enter apoptosis under nonpermissive condition, indicating that there is a quality control for aminoacyl-tRNA synthesis. DAD1 : The mutant defective in this gene, tsBN7, has a defect in N-linked glycosylation, and mice defective in this gene die due to apoptosis. Our results indicate a tight relationship between apoptosis and cycle : 1) We have identified Ran-binding proteins, RanRP1/Yrb1p, RanBP3/Yrb2, RanBP2/Yrb2, Dis3p, RanBPM and Mog1. The finding of RanBPM introduced that Ran is directly involved in microtubule assembly. 2) We found that a dual phosphates, Cdc25B triggers mitosis. Due to the instability of Cdc25B, the entry of mitosis is blodked by cycloheximide, a protein synthesis inhibitor. 3) As a suppressor gene of prp20 and rna1-1, ts mutants of S. cerevisiae RCC1 and RanGAP homologue, we isolated GTR1 and GTR2. These are new G proteins which seems to function antagonistically to the Ran cycle. Less

本研究主要包括三个部分：（1）从仓鼠BHK 21细胞系中分离细胞生长温度敏感（ts）突变体，（2）鉴定这些突变体中的突变基因，（3）克隆突变基因的特征。我会对每一部分进行复习。部分1.在此之前，我已经建立了一种方法来分离仓鼠BHK 21细胞系的ts突变体。然而，这些突变体中的大多数在X染色体中具有突变，其中表达单倍体剂量的基因。在这个项目中，我试图用一种药物苯菌灵分离常染色体的ts突变体。最后，我们分离出属于不同互补组的ts突变体。部分2.我们已经确定了仓鼠BHK 21细胞株ts突变体的突变基因，它们分别是编码HCF（疱疹病毒增殖宿主细胞因子）、组氨酰-tRNA合成酶和赖氨酰-tRNA合成酶的基因。部分3.我将审查已使用仓鼠BHK 21细胞系的ts突变体克隆的每个基因。H ...更多信息 CF：根据我们的发现，HCF被证实对宿主细胞增殖也是必不可少的。突变株tsBN 67在非允许温度下，G1、S、G2和M期完成后，仍停滞在G 0期。因此，该突变体将用于在分子水平上表征G 0期。氨酰-tRNA合成酶：组氨酰-和赖氨酰-tRNA合成酶缺陷的突变体在非允许条件下迅速进入细胞凋亡，表明氨酰-tRNA合成存在质量控制。DAD 1：该基因缺陷突变体tsBN 7在N-连接糖基化方面存在缺陷，该基因缺陷的小鼠因细胞凋亡而死亡。我们的研究结果表明细胞凋亡与细胞周期的密切关系：1）我们鉴定了RanRP 1/Yrb 1 p，RanBP 3/Yrb 2，RanBP 2/Yrb 2，Dis 3 p，RanBPM和Mog 1。RanBPM的发现说明Ran直接参与微管的组装。2)我们发现，一个双磷酸，Cdc 25 B触发有丝分裂。由于Cdc 25 B的不稳定性，有丝分裂的进入被蛋白质合成抑制剂放线菌酮阻断。3)ts突变体作为prp 20和rna 1 -1的抑制基因，在大肠杆菌中表达。为了鉴定RCC 1和RanGAP同源物，我们分离了GTR 1和GTR 2。这些是新的G蛋白，似乎对Ran循环起拮抗作用。少

项目成果

H. Goto: "A single point mutation in HCF causes temperature-sensitive cell-cycle arrest and disrupts VP16 function"Genes & Develop.. 11. 726-737 (1997)

H. Goto：“HCF 中的单点突变会导致温度敏感的细胞周期停滞并破坏 VP16 功能”基因

N. Nakashima: "Saccharomyces cerevisiae G Protein, Gtrlp, which forms complexes with itself and a novel protein designated as Gtr2p, negatively regulates the Ran/Gsp1p G protein cycle through Gtr2p"Genetics. 152. 853-867 (1999)

N. Nakashima：“酿酒酵母 G 蛋白 (Gtrlp) 与其自身形成复合物，并与一种称为 Gtr2p 的新型蛋白形成复合物，通过 Gtr2p 负向调节 Ran/Gsp1p G 蛋白循环”遗传学。

M. Oki: "A novel protein required for nuclear protein import. Moglp, directly interacts with GTP-Gsplp, Saccharomyces cerevisiae Ran-homologue"Proc. Natol. Acad. Science. 95. 15388-15393 (1998)

M. Oki：“核蛋白输入所需的一种新蛋白。Moglp，直接与 GTP-Gsplp、酿酒酵母 Ran 同源物相互作用”Proc。

E.noguchi: "Disruption of YRB2 gene reatands nuclear protein export,causing a profound mitotic delay,and can be rescued by overexpression of XPOI/CRMI." J.Biochem.(in press). (1999)

E.noguchi：“YRB2 基因的破坏会重新导致核蛋白输出，导致有丝分裂严重延迟，并且可以通过 XPOI/CRMI 的过度表达来挽救。”

H.Goto: "A single point mutation in HCF causes temperature-sensitive cell-cycle arrest and disrupts VP16 function." Genes & Develop.11. 726-737 (1997)

H.Goto：“HCF 中的单点突变会导致温度敏感的细胞周期停滞并破坏 VP16 功能。”