MOLECULAR ASSOCIATIONS OF PHOSPHOLIPASE C

性状（改编自申请人摘要）：磷脂酰肌醇 4，5-二磷酸[PtdIns（4，5）P2]是一种重要的分子， 真核细胞信号转导通路，参与细胞的调控 增殖、分化和致癌转化。 PtdIns（4，5）P2 被磷脂酶C（PLC）水解产生第二信使 1，4，5-三磷酸肌醇[Ins（1，4，5）P3]和甘油二酯（DG）。 Ins（1，4，5）P3触发钙从内部储存中释放， 调节Ca 2+调节途径，而DG激活磷脂-和 钙离子依赖性蛋白激酶C。PtdIns（4，5）P2本身调节其他 重要的功能，如几种肌动蛋白结合蛋白的活性。 显然，PtdIns（4，5）P2的细胞内浓度必须严格控制。 监管，但几乎没有人知道这个规则。 那个女人 长期目标是阐明调节 PtdIns（4，5）P2的稳态，并确定这种调节在 信号转导 该提案的具体目标是利用 双杂交筛选和酵母强大的实验优势 酿酒酵母鉴定与酵母相互作用的蛋白质 磷脂酶C（Plclp），并确定的意义， 酶的体内功能的相互作用。 申请人已经 进行了双杂交筛选，分离出许多质粒， 至少6个不同的组。 申请人将完成这项工作，并将 通过测序鉴定相应的基因（SA 1）。 计划是 证实Plclp和这些基因编码的蛋白质之间的相互作用， 独立生化技术（SA 2）。 目的是确定 负责相互作用的Plclp结构域（SA 3），并确定 这些相互作用对Plclp（SA 4）的体内功能的作用。

DESCRIPTION (Adapted from applicant's abstract): Phosphatidylinositol 4,5-bisphosphate [PtdIns(4,5)P2] is a vitally important molecule in eukaryotic signal transduction pathways, involved in the control of cell proliferation, differentiation, and oncogenic transformation. PtdIns(4,5)P2 is hydrolyzed by phospholipase C (PLC) to produce the second messengers inositol 1,4,5-trisphosphate [Ins(1,4,5)P3] and diacylglycerol (DG). Ins(1,4,5)P3 triggers release of calcium from internal stores and thus modulates Ca2+ -regulated pathways, while DG activates the phospholipid- and Ca2+ -dependent protein kinase C. PtdIns(4,5)P2 itself regulates other important functions, such as the activity of several actin binding proteins. Clearly, PtdIns(4,5)P2's intracellular concentration must be stringently regulated, but almost nothing is known about this regulation. The broad, long term objective is to elucidate the mechanisms regulating the homeostasis of PtdIns(4,5)P2 and to determine the role of this regulation in signal transduction. The specific aims of this proposal employ the two-hybrid screen and the powerful experimental advantages of the yeast Saccharomyces cerevisiae to identify proteins which interact with yeast phospholipase C (Plclp) and to determine the significance of the interaction(s) for the in vivo function of the enzyme. The applicant has performed the two-hybrid screen and isolated many plasmids which fall into at least 6 different groups. The applicant will complete this work and will identify the corresponding genes by sequencing (SA1). The plan is to confirm the interaction between Plclp and proteins encoded by these genes by independent biochemical techniques (SA2). The intent is to identify the domain(s) of Plclp responsible for the interactions (SA3) and to determine the roles of these interactions for the in vivo functions of Plclp (SA4).