SYSTEM ANALYSIS OF PROTEIN PHOSPHORYLATION INVOLVED IN SPERM MATURATION

这个子项目是利用资源的许多研究子项目之一。 由NIH/NCRR资助的中心拨款提供。对子项目的主要支持 子项目的首席调查员可能是由其他来源提供的， 包括美国国立卫生研究院的其他来源。为子项目列出的总成本可能 表示该子项目使用的中心基础设施的估计数量， 不是由NCRR赠款提供给次级项目或次级项目工作人员的直接资金。 离开哺乳动物睾丸的精子不能活动，也没有使卵母细胞受精的能力。精子成熟始于附睾部，在女性生殖道内继续发育，经过获能过程，最后发生顶体反应。这种成熟过程是独一无二的，因为精子在转录和翻译上都是静止的。精子成熟是在一个“封闭系统”中进行的，其中的变化主要涉及对现有蛋白质的修饰。发生的最大变化之一是许多精子蛋白质的磷酸化状态，这些变化似乎是精子功能和生育能力所必需的。因此，我们建议研究精子磷蛋白质组发生的系统范围的变化。这将通过两种方式来解决：1)使用我们已经从PNNL获得的附睾精的蛋白质组数据和信息学技术来确定哪些蛋白质具有可磷酸化的特定磷酸化基序，以及精子蛋白质组中存在哪些作用于这些基序的激酶和磷酸酶，以及2)分离不同成熟阶段的精子并对其进行磷酸蛋白质组学分析，以确定与成熟相关的重要磷酸化事件。这些数据将被用来组装精子磷酸蛋白质组的系统分析。这些实验将填补我们在理解精子成熟的磷酸化网络方面的一个重大空白。鉴于激酶和磷酸酶作为药物靶点的重要性，这项工作有望快速识别男性潜在的非荷尔蒙避孕靶点。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Spermatozoa that leave the mammalian testis are not motile and do not have the capability to fertilize an oocyte. Sperm maturation begins in the epididymis and continues in the female reproductive tract with the process of capacitation and finally the acrosome reaction. This maturation process is unique in that sperm are transcriptionally and translationally quiescent. Sperm maturation occurs in a "closed system" where changes involve primarily modification of existing proteins. One of the biggest changes that occur is the phosphorylation states of the many sperm proteins, and these changes appear to be required for sperm function and fertility. Consequently, we propose to investigate the system wide changes which occur to the phosphoproteome of the sperm. This will be approached two ways: 1) Using the proteomic data from epididymal sperm that we have already acquired from PNNL and informatics techniques to identify which proteins have specific phosphorylation motifs that can be phosphorylated and which kinases and phosphatases are present in the sperm proteome to act on these motifs, and 2) isolate sperm at various stages of maturation and subject them to phosphoproteomics analysis to identify important maturation-related phosphorylation events. These data will be used to assemble a system analysis of the sperm phospho-proteome. These experiments will fill a significant gap in our understanding of the phosphorylation network of sperm maturation. Given the importance of kinases and phosphatases as drug targets, this work promises rapid identification of potential non-hormonal contraceptive targets in the male.