CONTROL OF ENTRY INTO MEIOSIS IN YEAST

一个基本的生物学问题是细胞如何对 离散的监管信号被识别和集成到精确 操纵基因表达。这个问题直接影响到我们的 对控制发育和分化的认识 高等有机体。酵母菌的产孢子过程 酿酒酵母是防治研究的理想模式系统 真核生物的发育开关。这是一个简单的发展 计划依赖于多个监管影响，这些影响相互作用 促进特定细胞类型的减数分裂诱导和ASCUS形成 (统称为产孢子)。MCK1基因控制着 决定退出有丝分裂细胞周期并进入产孢期 途径，并在介导着丝粒行为中起积极作用 在有丝分裂期间。MCK1p基因产物被认为是 一类新的蛋白激酶具有新的能力 丝氨酸、苏氨酸和酪氨酸上的靶蛋白磷酸化 残留物。事实上，蛋白质磷酸化是一种常见的利用 真核发育中的信号转导机制 受监管的系统。因此，阐明其作用机制 对于理解蛋白激酶是如何 生物体控制着分化、生长和发育。此外， 多种酪氨酸激酶与转化和凋亡的关系 致癌潜力有助于增加诱因。这项建议 描述了旨在阐明MCK1的机制的研究 通过鉴定和控制减数分裂特异基因的表达 其底物、靶标和调节剂的表征。 MCK1p介导进入减数分裂反应的细胞类型和 通过控制减数分裂激活物的去阻遏来传递饥饿信号 基因IME1。因此，MCK1p靶标的磷酸化可以转导 启动ime1基因表达所需的信号(S)。 我们建议：(L)确定和表征可能的底物 采用三种方法相结合的方法：(A)分离突变株和 MCK1缺乏症的剂量依赖性抑制因子；(B)鉴定 与MCK1p物理相互作用的蛋白质；以及(C)鉴定 IME1启动子中MCK1反应位点的变化及其影响因素 与此网站互动。(2)探索监管影响 通过分析激酶活性和底物特异性来调控MCK1p 在已知影响进入减数分裂的条件下；和(3)进行 体外蛋白激酶活性及底物特异性分析 描述了这类独特的蛋白激酶。

One fundamental biological question is how cellular responses to discrete regulatory signals are recognized and integrated to precisely manipulate gene expression. This problem impacts directly on our understanding of the control of development and differentiation in higher organisms. The process of sporulation in the yeast Saccharomyces cerevisiae is an ideal model system for investigation of the control of developmental switches in eukaryotes. This simple developmental program relies on multiple regulatory influences which interact to promote cell-type-specific induction of meiosis and ascus formation (collectively referred to as sporulation). The MCK1 gene governs the decision to exit the mitotic cell cycle and enter the sporulation pathway, and plays a positive role in mediating centromere behavior during mitosis. The MCK1p gene product has been implicated as a member of a new class of protein kinase with the novel ability to phosphorylate target proteins on serine, threonine and tyrosine residues. Indeed, protein phosphorylation is a commonly exploited mechanism of signal transduction in eukaryotic, developmentally regulated systems. Therefore, elucidation of the mechanism of action of protein kinases is of paramount importance to understanding how an organism controls differentiation, growth and development. Furthermore, the association of many tyrosine kinases with transforming and oncogenic potential contributes added incentive. This proposal describes studies intended to elucidate the mechanism by which MCK1 controls meiotic-specific gene expression through identification and characterization of its substrates, targets and regulators. MCK1p mediates entry into meiosis in response to cell-type and starvation signals by controlling derepression of the meiotic activator gene IME1. Thus, phosphorylation of a target of MCK1p transduces the signal(s) required for initiation of IME1 gene expression. We propose to: (l) Identify and characterize putative substrates of MCK1p by a combination of three approaches: (a) isolation of mutant and dosage-dependent suppressors of an mck1 deficiency; (b) identification of proteins that physically interact with MCK1p; and (c) identification of the MCK1-Response Site in the IME1 promoter and factors that interact with this site. (2) Explore the regulatory influences that modulate MCK1p by analyzing kinase activity and substrate specificity under conditions known to affect entry into meiosis; and (3) Conduct in vitro kinase activity and substrate specificity analyses aimed at characterizing this unique class of protein kinase.