Control of Mitotic Entry by Regulators of Cdc2

描述（由申请人提供）：在所有真核细胞中，进入有丝分裂是由周期蛋白依赖性激酶Cdc2控制的。Cdc2/Cyclin B复合物由抑制激酶Myt1和Wee1以及活化磷酸酶Cdc25控制。虽然已经确定了许多Cdc25和Myt1/ wee1调控途径，但这些途径如何相互连接以及哪些上游调节剂影响这些途径尚不完全清楚。为了理解生理刺激和检查点控制通路如何调节G2/M转变，我们有必要了解这些不同调节因子之间的因果关系和调节相互作用。本研究利用爪蟾卵提取物提供的强大的体外有丝分裂进入重构系统来研究Cdc25和Wee1的调控。在本提案中，我们解决了G21M调控中的几个突出问题，包括Cdc25通过去除结合14-3-3蛋白在有丝分裂中被激活的机制，Cdc25定向激活磷酸酶是否可能被细胞周期调节，以及Cdc25的不同异构体如何被差异调节。我们还研究了Wee1的几个候选调节因子，包括激酶PIx和甲基转移酶Hsl7。我们的长期目标是获得控制进入有丝分裂的分子事件的机制理解。

DESCRIPTION (provided by applicant): In all eukaryotic cells, entry into mitosis is controlled by the Cyclin-dependent kinase, Cdc2. Cdc2/Cyclin B complexes are controlled by the inhibitory kinases, Myt1 and Wee1 and by the activating phosphatase, Cdc25. Although a number of Cdc25 and Myt1/Wee1-regulatory pathways have been identified, how these pathways are interconnected and which upstream modulators impinge on these pathways is not yet entirely clear. To understand how physiological stimuli and checkpoint control pathways regulate the G2/M transition, it is essential that we understand the causal connections and regulatory interplays between these diverse regulators. This proposal employs the powerful in vitro reconstitution system for mitotic entry provided by Xenopus egg extracts to investigate Cdc25 and Wee1 regulation. In this proposal we address several outstanding issues in G21M regulation, including the mechanism by which Cdc25 is activated at mitosis through removal of bound 14-3-3 proteins, whether Cdc25-directed activating phosphatases may be cell cycle regulated and how different isoforms of Cdc25 may be differentially regulated. We also investigate several candidate regulators of Wee1, including the kinase PIx and the methyltransferase, Hsl7. Our long-term objective is to obtain a mechanistic understanding of the molecular events that control entry into mitosis.