Characterization of a Testis Kinase Family (Tssk)

描述(申请人提供)：通过磷酸化对蛋白质进行翻译后修饰，在许多细胞过程中发挥作用，如细胞外信号转导、细胞内运输和细胞周期进程。我们小组对涉及精子分化和功能的信号事件的研究感兴趣。虽然减数分裂事件在雌性生殖细胞中有很好的特征，但对雄性生殖细胞减数分裂和分化为成熟精子的调节仍然不是很清楚。最近，在雄性生殖细胞中发现了一个新的丝氨酸/苏氨酸激酶家族(睾丸特异性丝氨酸激酶(Tesk))。我们已经证明在成熟的哺乳动物精子中存在一个或多个TSSK家族成员(Hao等，2004)(和PR)，此外，我们还获得了无脊椎动物的睾丸中表达TSSK同源物的证据。TSSK基因在睾丸中的保守表达模式以及磷酸化在信号传递过程中的重要性，有力地表明TSSK(S)在生殖细胞分化和/或精子功能中发挥着重要作用(S)。这一提议旨在为理解TSSK(S)在生殖细胞和成熟精子中的功能奠定基础。该项目的未来方向包括分析TSSK(S)的零突变体(KO)，以及利用无脊椎动物模型中可用的遗传学方法研究涉及TSSK(S)的信号通路。为了实现这些目标，首先有必要为这些研究创建工具，并确定这些激酶的特征。考虑到这一点，这一应用将集中在：1)针对TSSK家族5个成员的特异性抗体的产生；2)成熟精子中存在的TSSK(S)的鉴定及其亚细胞定位的研究；3)TSSK4激酶活性的生化特征以及潜在相互作用伙伴的研究。尽管不是该应用程序的一部分，但在第n大学的Mitch Eddy博士的合作下，Tssk4的鼠标KO正在生产过程中。重要的是，拟议的研究将有助于开始阐明TSSK基因功能的途径。最近对Tssk6 KO(也称为SSTK)的表型分析证实了Tssks在精子发生和/或精子功能中的预测重要性(Spiridonov等人，2005年)；这些零突变小鼠在雄性中呈现不育表型，在雌性中则不表现。作为这一应用程序的一部分而生成的工具对于分析tssk KO表型和开始研究tssk功能将是非常重要的。Tssks在精子发生和/或精子功能中的预测重要性表明，这项研究最终可以帮助设计新的人类避孕药。

DESCRIPTION (provided by applicant): Post translational modifications of proteins through phosphorylation play a role in many cellular processes such as the transduction of extracellular signals, intracellular transport, and cell cycle progression. Our group is interested in the study of signaling events that are involved in sperm differentiation and function. Although meiotic events are well characterized in the female germ cells, the regulation of male germ cell meiosis and differentiation into mature sperm is still not well understood. Recently, a novel family of serine/threonine kinases (testes specific serine kinases (Tssk)) has been found post meiotically expressed in the male germ cells. We have demonstrated the presence of 1or more members of the Tssk family in mature mammalian sperm (Hao et al., 2004) (and PR) and in addition, we have obtained evidence that Tssk homologues are expressed in the testes of invertebrates. The conserved testicular expression pattern of Tssk genes, as well as the importance of phosphorylation in signaling processes, strongly suggests that Tssk(s) have important role(s) in germ cell differentiation and/or sperm function. This proposal is designed to set the grounds to understand the function(s) of Tssk(s) in germ cells and in mature sperm. Future directions of this project include the analysis of null mutants (KO) of Tssk(s), as well as the investigation of signaling pathways involving Tssk(s) using genetic methods available in invertebrate models. Toward these goals, it will be necessary first to create tools for these studies and to characterize these kinases. Taking this into consideration, this application will focus on: 1) The generation of specific antibodies against the 5 members of the Tssk kinase family; 2) The identification of the Tssk(s) present in mature sperm and the investigation of their subcellular localization; 3) The biochemical characterization of Tssk4 kinase activity and the study of potential interacting partners. Although not part of this application, in collaboration with Dr. Mitch Eddy from NTH, a mouse KO of Tssk4 is in the process of being produced. Importantly, the proposed research will be useful to begin to illuminate the pathways of Tssk gene function. The predicted importance of Tssks in spermiogenesis and/or sperm function has been recently confirmed by phenotypic analysis of Tssk6 KOs (also known as Sstk)(Spiridonov et al., 2005); these null mutant mice present a sterile phenotype in the males but not in the females. The tools generated as part of this application will be very important for the analysis of Tssk KO phenotypes and to start the study of Tssk function. The predicted importance of Tssks in spermiogenesis and/or sperm function suggests that this research can ultimately help to design novel human contraceptives.