How do cyclins drive the cell cycle?

这个更新申请是为了继续研究细胞周期蛋白如何驱动细胞周期。 G1稳定剂Sic1和Cdh 1的多位点磷酸化的作用。细胞的G1期 由于Sid化学计量比的积累，B型细胞周期蛋白依赖性激酶对细胞周期是难治的。 抑制剂，以及由于Cdh 1引起的高活性B型细胞周期蛋白水解。多位点磷酸化 G1细胞周期蛋白的Sic1和Cdh1都被认为是从G1退出所必需的。不过，我们发现， 来自内源基因座的不可磷酸化的Sic1（所有9个Cdk位点突变）的表达导致 完全可行的菌株，尽管具有延长的G1。我们将类似地测试不可磷酸化的 Cdh 1由内源启动子表达。这些实验将解决动态 结果的多位点磷酸化的G1调节细胞周期蛋白-Cdk复合物，在生理水平。 B型细胞周期蛋白与细胞周期执行机制的相互作用。虽然我们对细胞有很多了解 周期振荡器控制细胞周期蛋白依赖性激酶和后期促进复合物的水平， 这些活动最终如何驱动细胞周期的实际事件，如DMA复制， 或主轴功能。我们最近的研究结果表明，六个酵母B型的高度冗余， 细胞周期蛋白基因是唯一明显的：非必要的“检查点”监视机制和其他调控机制 在缺乏细胞周期蛋白特异性途径的情况下，保护措施变得至关重要。禁用这些监管 保障措施允许集中于由特定细胞周期蛋白控制的细胞生物学途径。 cdc14：靶点与调节因子。Cdc14是退出有丝分裂所需的磷酸酶;它从有丝分裂中释放出来。 在有丝分裂退出之前在核仁中的隔离。Cdc14可能使Cdk靶去磷酸化， 因此有助于逆转有丝分裂状态，但如果Cdc14在体内对一些关键靶点具有特异性， 或者使大部分或全部Cdk底物脱磷酸。我们已经利用突变影响Cdc14 定位以探索Cdc14靶标的光谱。在其他研究中，我们将确定 在内源性表达水平上阻断Pds 1降解的结果，以及功能性 Cdk介导的有丝分裂出口激酶Dbf2磷酸化的意义。这些实验将探索 Clb激酶-Cdc14磷酸酶拮抗作用在细胞周期调控中的动态结果。 总的来说，我们感兴趣的是细胞周期动力学的调节，以及细胞周期蛋白特异性途径， 单个细胞周期事件。

This renewal application is to continue work on how cyclins drive the cell cycle. The role of multi-site phosphorylation of the G1 stabilizers Sic1 and Cdh1. The G1 period of the cell cycle is refractory to B-type cyclin dependent kinase because of accumulation of the Sid stoichiometric inhibitor, and because of highly active B-type cyclin proteolysis due to Cdh1. Multi-site phosphorylation of both Sic1 and Cdh1 by G1 cyclins have been considered essential for exit from G1. We have found, though, that expression of unphosphorylatable Sic1 (all 9 Cdk sites mutated) from the endogenous locus results in a fully viable strain, though with a lengthened G1. We will similarly test the properties of unphosphorylatable Cdh1 expressed from the endogenous promoter. These experiments will address the dynamic consequences of multisite phosphorylation of G1 regulators by cyclin-Cdk complexes, at physiological levels. Interactions of B-type cyclins with cell cycle execution machinery. While a lot is known about the cell cycle oscillator controlling levels of cyclin-dependent kinase and anaphase-promoting complex, much less is known about how these activities eventually drive the actual events of the cell cycle such as DMAreplication or spindle function. Our recent results indicate that the high degree of redundancy of the six yeast B-type cyclin genes is only apparent: non-essential 'checkpoint' surveillance mechanisms and other regulatory safeguards become essential in the absence of cyclin-specific pathways. Disabling these regulatory safeguards allows focus on cell biological pathways controlled by specific cyclins. Cdc14: targets and regulators. Cdc14 is a phosphatase required for exit from mitosis; it is released from sequestration in the nucleolus just before mitotic exit. Cdc14 probably dephosphorylates Cdk targets and thus helps reverse the mitotic state, but it is unresolved if Cdc14 is specific in vivo for a few critical targets, or alternatively dephosphorylates most or all Cdk substrates. We have exploited mutants affecting Cdc14 localization to explore the spectrum of Cdc14 targets. In additional studies we will determine the consequences of blocking Pds1 degradation, at endogenous expression levels, and the functional significance of Cdk-mediated phosphorylation of the mitotic exit kinase Dbf2. These experiments will probe the dynamic consequences of Clb kinase-Cdc14 phosphatase antagonism in cell cycle regulation. Overall, we are interested in regulation of cell cycle dynamics, and in cyclin-specific pathways promoting individual cell cycle events.