DEREGULATING CYCLIN DEPENDENT KINASE

细胞周期蛋白依赖性激酶（Cdk）在所有真核生物的细胞周期调节中是关键的。Cdk受细胞周期蛋白结合的严格调节，因此细胞周期蛋白的可用性是调节细胞周期进程的主要手段之一。这项建议是为了实验，以检查放松管制的后果，这一系统。在一组实验中，提出了一种策略，突变激活的芽殖酵母Cdk Cdc 28独立的细胞周期蛋白的结合要求。这一目标的实现（已取得重大进展）将允许解剖的作用，细胞周期蛋白在简单的酶激活，而不是针对特定的底物的酶。在第二组实验中，我们建议检查的后果，使特定的B型细胞周期蛋白/Cdc 28复合物，Clb 5-Cdc 28，通过细胞周期组成的水平和活动。虽然目前的模型预测，这应该停止在G1期的细胞周期，由于干扰的DNA复制起点的功能，我们的研究结果表明，这可能不是这样的。我们建议通过细胞周期来表征Clb 5降解的控制，以及Clb 5破坏盒在其降解中的作用。我们提出的实验来测试的假设，Clb 5是内在专门的S期进入，而另一个B型细胞周期蛋白，Clb 2，是内在专门进入有丝分裂。这一假设与目前的模型相反，在现有的模型中，所有B型细胞周期蛋白都同样能够对DNA复制进行正向和负向调节。

Cyclin-dependent kinases (Cdk's) are critical in regulation of the cell cycle in all eukaryotes. Cdk's are stringently regulated by cyclin binding, and cyclin availability thus is one of the main means by which cell cycle progression is regulated. This proposal is for experiments to examine the consequences of deregulation of this system. In one set of experiments, a strategy is proposed for mutational activation of the budding yeast Cdk Cdc28 independent of the cyclin binding requirement. Achievement of this goal (towards which significant progress has been made) will allow dissection of the roles of cyclins in simple enzyme activation as opposed to targeting of the enzyme to specific substrates. In a second set of experiments, we propose examining the consequences of making the level and activity of a particular B-type cyclin/Cdc28 complex, Clb5-Cdc28, constitutive through the cell cycle. Although current models predict that this should halt the cell cycle in G1 phase due to interference with function of origins of DNA replication, our results suggest that this may not be the case. We propose to characterize control of Clb5 degradation through the cell cycle, and the role of the Clb5 destruction box in its degradation. We propose experiments to test the hypothesis that Clb5 is intrinsically specialized for S phase entry while another B-type cyclin, Clb2, is intrinsically specialized for entry into mitosis. This hypothesis is in contrast to current models in which all B-type cyclins are equally capable of both positive and negative regulation of DNA replication.