SIGNAL TRANSDUCTION AT FERTILIZATION

描述(改编自《调查员摘要》)：此次更新 应用关注的是精子通过何种信号机制 使卵子在受精时开始发育。该项目是 专注于这个过程中发生在鸡蛋上的最早的事件 质膜。这些膜事件导致细胞内钙离子升高， 这反过来又会导致许多后续的发展过程。该项目 包括3个具体目标：(1)研究G-α-Q家族的作用 受精时信号转导中的蛋白质。G-α-Q的量 注射反义基因将减少蛙卵中的家族蛋白 寡核苷酸，以确定这是否抑制钙离子的释放 受精。这些研究还将检查G-α-Q是否 在受精时被激活，使用G-α-Q蛋白的保护 蛋白质分解作为一种分析方法。(2)研究酪氨酸的作用 磷脂酶C-γ在血管内皮生长因子信号转导中的磷酸化 受精。磷脂酶C-γ的激活将被抑制 注射PLC-Gamma SH2结构域，以确定这是否降低了钙离子 在受精时释放。这些研究还将检查PLC-伽马 在受精时被激活，使用该酶的酪氨酸磷酸化 作为一种化验。(3)研究α-v整合素在信号中的作用 受精时的转导。这些研究将调查是否 刺激α-v整合素引起细胞内钙离子升高 两栖虫卵，以及它是否增加了脂质PIP2的量， 磷脂酶C的底物。他们还将检测钙离子是否释放。 通过抑制α-v整合素功能来减少AT受精。

DESCRIPTION (Adapted from the Investigator's Abstract): This renewal application is concerned with the signalling mechanisms by which a sperm causes an egg to begin development at fertilization. The project is focussed on the earliest events of this process, that occur at the egg plasma membrane. These membrane events cause a rise in intracellular Ca2+, which in turn causes many subsequent developmental process. The project includes 3 specific aims: (1) To examine the role of G-alpha-q family proteins in signal transduction at fertilization. The amounts of G-alpha-q family proteins in frog eggs will be reduced by injection of antisense oligonucleotides, to determine if this inhibits Ca2+ release at fertilization. These studies will also examine whether G-alpha-q is activated at fertilization, using protection of the G-alpha-q protein from proteolysis as an assay. (2) To examine the role of tyrosine phosphorylation of phospholipase C-gamma in signal transduction at fertilization. Activation of phospholipase C-gamma will be inhibited by injection of PLC-gamma SH2 domains, to determine if this reduces Ca2+ release at fertilization. These studies will also examine whether PLC-gamma is activated at fertilization, using tyrosine phosphorylation of this enzyme as a assay. (3) To examine the role of alpha-v integrins in signal transduction at fertilization. These studies will investigate whether stimulation of an alpha-v integrin causes a rise in intracellular Ca2+ in amphibian eggs, and whether it increases the amount of the lipid PIP2, the substrate for phospholipase C. They will also examine whether Ca2+ release at fertilization is reduced by inhibition of alpha-v integrin function.