Regulatory Factors of Cyclin Functions in the Cell Cycle

细胞周期中细胞周期蛋白功能的调控因素

基本信息

批准号：
08044213
负责人：
KOBAYASHI Hideki
金额：
$ 6.14万
依托单位：
KYUSHU UNIVERSITY
依托单位国家：
日本
项目类别：
Grant-in-Aid for international Scientific Research
财政年份：
1996
资助国家：
日本
起止时间：
1996 至 1998
项目状态：
已结题

项目摘要

According to the proposed research plan, Hideki Kobayashi, Kyushu University in Japan, carried out the collaboration on the cyclin functions in the cell cycle, with Dr.Tim Hunt at ICRF Clare Hall Laboratories, and Dr.Mark Carrington at Cambridge University, England. Hideki Kobayashi isolated Xenopus cyclin A-binding protein by yeast 2-hybrid screen and identified the novel protein THF4 that binds to the N-terminus of cyclin A and inhibits its degradation selectively. Tim Hunt analyzed its biochemical properties on the cell cycle in Xenopus egg extracts. Mark Carrington has isolated murine homolog of THF4 and identified its binding domain to cyclin A.In summary, the N-terminal domain of Xenopus cyclin A1 was used in a 2-hybrid screen to identify proteins that interact with cyclin A.The major interacting species was THF4, a protein that contains a ubiquitin-like domain in its N-terminus shows a significant homology in the C-terminus of S.cerevisiae Dsk2 and S.porn be dph1. THF4 persists … More in Xenopus cells as a nuclear phosphoprotein and its phosphorylation is induced by cyclin A-dependent kinase. THF4 binds to both embryonic and somatic forms of cyclin A (Al and A2) in vivo and in vitro, but not to B-type cyclins. The N-terminal u1biquitin-like domain of THF4, but not the C-temimal conserved domain, is required for interaction with cyclin A.THF4 requires residues 130-160 of cyclin Al for efficient binding, which do not include the destruction box of cyclin A.The addition of bacterially-expressed THF4 protein to_frog egg extract inhibited the Ca2+-induced degradation of cyclin A, but not that of cyclin B.The injection of THF4 protein into the fertilized egg blocked the embryonic cell division.These results suggest that THF4 is a candidate substrate for cyclin A-dependent kinase and that THF4 may act as a negative regulator of cyclin A degradation in mitosis.These results would give a new insight into cyclin proteolysis in the cell cycle This collaborative research project during three years (1996-1998) was quite successful, and we will continue the collaborative research more. Less

根据拟议的研究计划，日本九州大学的Hideki Kobayashi与ICRF Clare Hall Laboratories的Tim Hunt博士和英格兰剑桥大学的Mark Carrington博士一起进行了Cyclin功能的合作。 Hideki Kobayashi通过酵母2杂交筛选分离了Xenopus cyclin a结合蛋白，并鉴定出与细胞周期蛋白A的N末端结合的新型蛋白THF4，并选择性地抑制其降解。蒂姆·亨特（Tim Hunt）在爪蟾卵提取物中分析了其在细胞周期上的生化特性。马克·卡灵顿（Mark Carrington）分离了THF4的鼠同源物，并鉴定了其与细胞周期蛋白A的结合结构域。在总结中，在2杂交筛选中使用了xenopus cyclin a1的N末端结构域，以识别与细胞周期蛋白相互作用的蛋白质。 S.cerevisiae dsk2和s.porn为dph1。 THF4持续……在异氨酸细胞中，更多作为一种核磷酸蛋白，其磷酸化是由细胞周期蛋白A依赖性激酶诱导的。 THF4在体内和体外与细胞周期蛋白A（Al和A2）的胚胎和体细胞形式结合，但不与B型细胞周期蛋白结合。与细胞周期蛋白A.THF4相互作用的THF4的N末端U1Biquitin样结构域，而不是C-阵营的配置结构域，需要保留130-160的细胞周期蛋白Al，以进行有效的结合，这些结合均不包括Cyclin+Cyclin的损坏。细胞周期蛋白A，但不是细胞周期蛋白B。将Thf4蛋白注射到受精卵中，阻塞了胚胎细胞的分裂。这些结果表明THF4是Cyclin a依赖性激酶的候选底物，THF4可能充当有丝分裂中Cyclination A降解的负调控剂。这些结果将为细胞周期中的细胞周期蛋白水解提供新的见解，这一三年（1996-1998）的合作研究项目非常成功，我们将继续进行协作研究。较少的