The Protein Complex of a Sperm Ion Channel

精子离子通道的蛋白质复合物

• 批准号: 6987909
• 负责人: Dejian Ren
• 金额: $ 7.74万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-12-01 至 2006-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6987909
• 关键词: affinity chromatography; calcium flux; cyclic AMP; electrophysiology; fertility; genetically modified animals; green fluorescent proteins; immunocytochemistry; immunoprecipitation; laboratory mouse; male; membrane channels; molecular cloning; protein protein interaction; protein purification; protein sequence; protein structure function; sperm; sperm motility; transfection; voltage /patch clamp

DESCRIPTION (provided by applicant): The overall goal of this research is to understand the regulation of sperm function by calcium, cyclic nucleotides and ion channels at the molecular level. This proposal focuses on the role of a sperm tail ion channel (CatSper1) that is specifically required for male fertility. The CatSper1 channel that we recently cloned from humans and mice is expressed only in the testes. The protein in sperm is strikingly localized in the principal piece membrane. Mice deficient in CatSper1 show no phenotype except that males are infertile due to reduced sperm motility. cAMP-induced calcium influx into sperm is deficient in CatSper1-null mice. Because of its restricted protein expression in the sperm tail and because of its restricted knockout phenotype in male fertility, CatSper1 represents an excellent target for a safe, non-hormonal, male contraceptive. Our preliminary results indicate that CatSper1 is associated with other proteins to achieve its cAMP-activated signaling. We propose to identify the proteins associated with CatSper1. We will achieve two specific aims: 1) To purify the protein complex containing CatSper1 using affinity purification, and 2) To clone the interacting proteins and test the protein interaction using immunoprecipitation and immunostaining. The interacting proteins discovered from this study will be essential for the functional expression of CatSper1. Such an expression system will allow us to study the regulation of sperm function by ion channels at the molecular level. It may also lead to the development of a safe and efficient male contraceptive.

描述（申请人提供）：本研究的总体目标是在分子水平上了解钙、环核苷酸和离子通道对精子功能的调节。该提案重点关注男性生育能力特别需要的精子尾离子通道 (CatSper1) 的作用。我们最近从人类和小鼠身上克隆的 CatSper1 通道仅在睾丸中表达。精子中的蛋白质明显位于主膜中。 CatSper1 缺陷的小鼠没有表现出任何表型，只是雄性小鼠由于精子活力降低而无法生育。在 CatSper1 缺失小鼠中，cAMP 诱导的钙离子流入精子中存在缺陷。由于其在精子尾部的蛋白质表达受到限制，并且由于其在男性生育力方面的敲除表型受到限制，CatSper1 代表了安全、非激素男性避孕药的绝佳靶标。我们的初步结果表明 CatSper1 与其他蛋白质相关以实现其 cAMP 激活信号传导。 我们建议鉴定与 CatSper1 相关的蛋白质。 我们将实现两个具体目标：1) 使用亲和纯化纯化含有 CatSper1 的蛋白质复合物，2) 克隆相互作用的蛋白质并使用免疫沉淀和免疫染色测试蛋白质相互作用。本研究中发现的相互作用蛋白对于 CatSper1 的功能表达至关重要。这样的表达系统将使我们能够在分子水平上研究离子通道对精子功能的调节。它还可能导致安全有效的男性避孕药的开发。