Cell Cycle Regulation of the Yeast HO Gene

描述(申请人提供)：控制细胞周期的机制在所有真核生物中非常保守。哺乳动物和萌芽酵母都承诺在G1期进行另一轮细胞分裂。致癌过程通过干扰G1进展的调节来发挥其最大的影响。我们建议研究控制发芽酵母G1进程的分子机制。我们的实验旨在确定如何调节对萌芽酵母细胞周期的承诺，以应对营养变化、细胞生长和逆境，如DNA损伤。在酵母中，就像在所有高等真核生物中一样，细胞周期转换是由细胞周期蛋白依赖性蛋白激酶(CDK)控制的。已鉴定出九种细胞周期蛋白结合并激活芽殖酵母(CDC28)的CDK。三个周期蛋白Cln1、Cln2和Cln3在调节G1向S的转变过程中起着关键作用。我们的重点是确定是什么控制了这三个周期蛋白在G1期的表达。ECB元件在M/G1边界激活CLN3和其他关键细胞周期调节因子的转录。已经确定了另外两个启动子元件，它们在G1期被Swi4/Swi6和/或MBPL/Swi6激活。这些复合体激活了许多基因的转录，包括cln1和cln2，它们是向S期过渡的限速基因。我们的目标是了解内部和外部信号如何调控这些转录复合体的活性，并控制向S期的过渡。我们已经确定了影响欧洲央行活动的一个激活者和两个抑制者。我们将使用遗传和生化策略来确定这些元素是如何赋予细胞周期转录调控的。我们还将研究Swi6在激活G1转录和抑制转录以应对DNA损伤方面的调节作用。我们已经确定了RAD53激酶磷酸化Swi6以响应DNA损伤的位置。我们将确定这些磷酸化对G1DNA损伤检查点的意义。我们还将确定从我们的研究中获得的Rad53磷酸化的共识位点是否使我们能够预测Rad53修饰其他已知靶点的位点。

DESCRIPTION (provided by applicant): The mechanisms that control the cell cycle are remarkably conserved among all eukaryotes. Both mammals and budding yeast commit to another round of cell division during G1. Oncogenic processes exert their greatest impact by interfering with regulators of G1 progression. We propose to investigate the molecular mechanisms which control G1 progression in budding yeast. Our experiments are designed to determine how commitment to the budding yeast cell cycle is regulated in response to nutritional shifts, cell growth and adversities, such as DNA damage. In yeast, as in all higher eukaryotes, cell cycle transitions are governed by cyclin-dependent kinases (Cdks). Nine cyclins have been identified that bind and activate the Cdk of budding yeast (Cdc28). Three cyclins Cln1, Cln2 and Cln3 play critical roles in modulating the G1 to S transition. Our focus is upon determining what controls the expression of these three cyclins during G1. ECB elements activate transcription of CLN3 and other key cell cycle regulators at the M/G1 boundary. Two other promoter elements have been identified that are activated during G1 by Swi4/Swi6 and/or Mbpl/Swi6. These complexes activate transcription of many genes including CLN1 and CLN2, which are rate limiting for the transition to S phase. Our goal is to understand how internal and external signals modulate the activity of these transcription complexes and control the transition to S phase. We have identified one activator and two repressors that influence ECB activity. We will use genetic and biochemical strategies to determine how cell cycle regulation of transcription is conferred by these elements. We will also investigate the regulatory role of Swi6 in activating transcription in G1 and inhibiting transcription in response to DNA damage. We have identified sites at which Rad53 kinase phosphorylates Swi6 in response to DNA damage. We will determine the significance of these phosphorylations to the G1 DNA damage checkpoint. We will also determine whether the consensus site for Rad53 phosphorylation that we draw from our studies enables us to predict the sites at which Rad53 modifies other known targets.