SIGNAL TRANSDUCERS IN SPERM

描述：(从申请者的描述中扫描)受精 哺乳动物在一个已知的过程中依赖于精子的生理启动 作为获能能力。这个项目的长期目标是确定 获能的分子和细胞机制。Trp2是精子的一种钙通道 由精子与卵子透明带接触而激活，并驱动 顶体反应。在精子分化和成熟过程中，Trp2 组装成翻译后修饰的异构体复合体 在获能过程中，从而调节离子通道功能。作为这个频道 在受精过程中扮演确定的角色，并经历确定的 在精子获能过程中的结构和功能调节，它提供了一个 独特的模型系统，用于确定潜在的分子过程。 Trp2在获能过程中的调节机制将在 这个资助期通过结合最先进的电生理方法 和基因组/蛋白质组方法。有四个具体目标。首先， Trp2复合体的蛋白质成分将通过以下方法分离 免疫共沉淀和蛋白质组质谱鉴定。第二, Trp2和Trp2相关蛋白的磷酸化状态为 下定决心。第三，获能对Trp2离子通道活性的影响 将由电生理学研究确定。最后一点是， Trp2相关蛋白及其在离子调控中的蛋白磷酸化 通道活动将通过电生理学方法来确定。 这些研究可能在几个层面上具有重要意义。第一，对 获能是我们受精的基本模式的核心。第二， 无法有效地获得人类精子的能力可能解释了 不孕症以及体外培养过程中的部分失败率 受精。此外，据报道，精子钙通道拮抗剂具有 男性避孕药的行为，从而理解 通道功能是设计通道型抗生育药物所必需的。 最后，trp蛋白在广泛的信号转导过程中发挥作用。 细胞的范围，但我们对这些重要通道的了解有限 由于缺乏特定的敌手。产生一种功能阻断抗体 并用来研究内源性Trp2通道的功能 老鼠的精子。因此，这些研究也可能为 色氨酸通道在其他组织中的功能。

DESCRIPTION: (Scanned from the applicant's description) Fertilization in mammals depends upon the physiological priming of sperm during a process known as capacitation. The long range goal of this project is to determine the molecular and cellular mechanisms of capacitation. TRP2 is a sperm Ca2+ channel that is activated by sperm contact with the egg zona pellucida and drives the acrosome reaction. During differentiation and maturation of sperm, TRP2 assembles into heteromerc complexes that are posttranslationally modified during capacitation, thereby modulating ion channel function. As this channel plays a defined role in the fertilization process, and undergoes defined structural and functional regulation during sperm capacitation, it provides a unique model system in which to determine the underlying molecular processes. The mechanisms of TRP2 regulation during capacitation will be determined in this funding period by combining state-of-the-art electrophysiological methods and genomic/proteomic approaches. There are four Specific Aims. First, the proteins components of TRP2 complexes will be isolated by co-immunoprecipitation and identified by proteomic mass spectroscopy. Second, the phosphorylation state of TRP2 and of TRP2-associated proteins will be determined. Third, the effects of capacitation on TRP2 ion channel activity will be determined by electrophysiological studies. Finally, the role of TRP2-associated proteins and of protein phosphorylation in the control of ion channel activity will be determined by electrophysiological methods. These studies may be significant at several levels. First, an understanding of capacitation is central to our basic models of fertilization. Second, the inability to capacitate human sperm efficiently may account for aspects of infertility, as well as for part of failure rates during in vitro fertilization. Moreover, sperm Ca2+ channel antagonists are reported to have male contraceptive action and hence an understanding of the regulation of channel function is necessary for design of channel-based antifertility agents. Finally, TRP proteins are implicated in signal transduction processes in a wide range of cells, but our knowledge of these important channels has been limited by the lack of specific antagonists. A function-blocking antibody was generated and is utilized here to study the functions of the endogenous TRP2 channel of mouse sperm. Consequently, these studies may also provide a model for the functions of TRP channels in other tissues.