YEAST PP2B PHOSPHATASES--ROLE IN PHEROMONE ADAPTATION

描述：钙调磷酸酶（或PP 2B），一种保守的蛋白磷酸酶， 由Ca 2+和钙调蛋白激活，参与信号转导， 影响细胞内磷酸化状态的钙依赖性变化， proteins. 钙调神经磷酸酶是哺乳动物T细胞的关键调节因子， 这种磷酸酶的抑制剂（FK 506，环孢菌素A） 免疫抑制剂 我们的目标是了解 酵母中的Ca 2+依赖性信号传导，特别强调 钙调神经磷酸酶 我们对酵母的研究应该能为我们了解 Ca 2+依赖性信号传导的机制和功能，特别是 钙调神经磷酸酶介导的信号传导，在所有真核细胞中。 交配反应提供了一个机会，以检查的作用， 酵母中的Ca 2+依赖信号。 向单倍体酵母中添加信息素 细胞产生胞质Ca 2+的升高，并激活钙调神经磷酸酶和 钙调素刺激的激酶（CaM激酶）通过这种Ca 2+信号是必需的 为了生存能力 钙调神经磷酸酶的一个基本功能是激活 crz 1 p转录因子，虽然关键的目标， 钙调神经磷酸酶/CRZ 1依赖的转录调控尚未被 鉴定 钙调神经磷酸酶还执行额外的，尚未表征， 信息素处理过的细胞的基本功能。 钙调素激酶的作用 信息素的反应不被理解。 拟议的实验将 检测钙调神经磷酸酶和钙调蛋白激酶在暴露于 费洛蒙 我们将：1）描述信息素诱导的机制 由钙调神经磷酸酶和Crz 1 p介导的转录。 Crz 1 p模式 调节钙调磷酸酶将阐明和其他基因产物 参与钙调神经磷酸酶/CRZ 1反应途径的细胞， 鉴定 2)确定基因产物，修改的要求， 在与信息素孵育期间的Ca 2+依赖信号。 的组件 将研究钙调磷酸酶和CaM激酶依赖性应答。 第三章 确定信息素诱导的钙调神经磷酸酶依赖性转录的靶点 调节并确定这些基因对细胞活力的贡献。 微阵列技术将用于检测所有6000个酵母基因， 钙调神经磷酸酶依赖的基因表达变化。 4)识别底物 钙调磷酸酶通过识别基因产物， 钙调磷酸酶在体内和测试特定的候选蛋白在体外， 钙调神经磷酸酶依赖性去磷酸化。

DESCRIPTION: Calcineurin (or PP2B), a conserved protein phosphatase that is activated by Ca2+ and calmodulin, participates in signal transduction by effecting Ca2+-dependent changes in the phosphorylation state of cellular proteins. Calcineurin is a critical regulator of mammalian T-cells, and inhibitors of this phosphatase (FK506, cyclosporinA) are powerful immunosuppressants. Our goal is to understand the functions of Ca2+-dependent signaling in yeast, with particular emphasis on the role of calcineurin. Our studies of yeast should provide insight into the mechanisms and functions of Ca2+-dependent signaling, especially calcineurin-mediated signaling, in all eukaryotic cells. The mating response provides an opportunity to examine the role of Ca2+-dependent signaling in yeast. Addition of pheromone to haploid yeast cells produces a rise in cytosolic Ca2+, and activation of calcineurin and calmodulin-stimulated kinase (CaM kinase) by this Ca2+ signal is required for viability. One essential function of calcineurin is to activate the Crz1p transcription factor, although the critical targets of calcineurin/CRZ1-dependent transcriptional regulation have not yet been identified. Calcineurin also performs additional, as yet uncharacterized, essential functions in pheromone-treated cells. The role of CaM kinase in the pheromone response is not understood. The proposed experiments will examine the functions of calcineurin and CaM kinase in cells exposed to pheromone. We will: 1) Characterize the mechanism of pheromone-induced transcription mediated by calcineurin and Crz1p. The mode of Crz1p regulation by calcineurin will be elucidated and additional gene products that participate in the calcineurin/CRZ1 response pathway will be identified. 2) Identify gene products that modify the requirement for Ca2+-dependent signaling during incubation with pheromone. Components of both calcineurin and CaM kinase-dependent responses will be studied. 3) Identify targets of pheromone-induced calcineurin-dependent transcriptional regulation and determine the contribution of these genes to cell viability. Microarray technology will be employed to examine all 6000 yeast genes for calcineurin-dependent changes in gene expression. 4) Identify substrates of calcineurin by identifying gene products that physically interact with calcineurin in vivo and by testing specific candidate proteins in vitro for calcineurin-dependent dephosphorylation.