Calcium signaling nanodomains in sperm motility and fertility

精子活力和生育力中的钙信号纳米结构域

• 批准号: 10392928
• 负责人: Jean-Ju Lucia Chung
• 金额: $ 34.88万
• 依托单位: YALE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-08-01 至 2024-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10392928
• 关键词: Affect; Age; Alkalinization; Animal Model; Binding; Biology; Calcium Signaling; CatSper; Complex; Coupled; Coupling; Cytoskeleton; DNA Sequence Alteration; Data; Defect; Development; Diagnosis; Electrophysiology (science); Feedback; Fertility; Fertilization; Fertilization in Vitro; Goals; Human; Image; In Situ; Individual; Infertility; Knock-out; Knockout Mice; Knowledge; Lead; Light; Male Infertility; Mechanics; Membrane; Methods; Microscopic; Mission; Modeling; Molecular; Motion; Movement; Mus; Outcome; Pathway interactions; Penetration; Persons; Pharmacology; Phenotype; Phosphorylation; Physiology; Prevention; Process; Property; Proteins; Proteomics; Public Health; Quality of life; Race; Regulation; Research; Research Proposals; Resolution; Role; Signal Pathway; Signal Transduction; Signal Transduction Pathway; Signaling Molecule; Site; Sperm Capacitation; Sperm Motility; Sperm Tail; Spermatogenic Cell; System; Testing; Time; United States National Institutes of Health; assisted reproduction; base; cell motility; comparative; contraceptive target; egg; experimental study; imaging modality; improved; innovation; insight; loss of function; male fertility; molecular imaging; novel; patch clamp; prospective; reconstitution; reproductive; sensor; sperm cell; trafficking

PROJECT SUMMARY Since discovery in 2001, flagellar Ca2+ channel CatSper has remained the only Ca2+ channel in which genetic mutations cause male infertility. There are critical knowledge gaps in the current understanding of the functional regulation of this critically important channel and its signaling pathways to trigger hyperactivation, an asymmetric flagellar motion required for male fertility. The long-term goal is to deepen the understanding of sperm physiology and to uncover new pathways that could be exploited to assess and control sperm motility and fertility. CatSper channels form unique multi-protein Ca2+ signaling complexes in four linear “racing stripe” nanodomains along the sperm tail. The presence of this spatially ordered nanodomains serves as a marker of successful sperm hyperactivation. The overall objective of this application is to elucidate how the CatSper channel complex is molecularly defined and structurally organized to integrate signal transduction pathways leading to the mechan- ical transitions required for hyperactivation. The central hypothesis is that the Ca2+ signaling state within CatSper nanodomains determines sperm motility and fertility. The rationale for determining the molecular mechanisms of CatSper channel regulation is that this knowledge will likely offer a strong scientific framework whereby new pharmacological strategies to alter sperm motility, and thus male fertility, can be developed. This application proposes to characterize novel CatSper-interacting molecules implicated in Ca2+ binding and membrane traffick- ing and microscopically visualize the Ca2+ signaling state in sperm ready to fertilize. The central hypothesis will be tested by pursuing three specific aims: 1) Define the function of the novel CatSper components in regulating channel activity and sperm motility; 2) Determine the role of the novel molecules in nanodomain formation; and 3) Elucidate the domain-integrated signaling pathways in sperm that achieve fertilization. Electrophysiological recording and flagellar waveform analysis will be employed to evaluate loss-of-function phenotype effects on channel activity and sperm motility. For the second aim, a different stage of spermatogenic cells from the knock- out models will be used to determine their role in channel complex assembly and nanodomain formation. For the third aim, motility-correlation super-resolution and in situ molecular imaging methods will be used to investigate the signaling state and domain organization of individual sperm cells with proven motility and fertility. The research proposal is innovative, in the applicant’s opinion, because it focuses on new CatSper components using new animal models to test an original concept of coupling signaling domain organization to channel activity in regulating Ca2+ signaling, and incorporates new methods into the sperm biology field. The proposed research is significant because it is expected to provide new mechanistic insights into CatSper channel activity and sperm motility regulation and molecular properties of sperm that are ready to fertilize. Ultimately, such knowledge has the potential to offer new opportunities for improvements in assisted reproduction, diagnosis of male infertility, and development of new targets for contraceptives.

项目总结 自2001年发现以来，鞭毛膜上的钙通道CatSper一直是唯一一条钙离子通道。 突变会导致男性不育。在当前对功能的理解中存在着关键的知识缺口 调节这一至关重要的通道及其信号通路以触发过度激活，不对称 鞭毛运动是男性生育所必需的。长期目标是加深对精子生理学的理解 并发现可以用来评估和控制精子运动和生育能力的新途径。 CatSper通道在四个线性的“赛跑条纹”纳米结构域中形成独特的多蛋白钙信号复合体 沿着精子尾巴。这种空间有序的纳米结构域的存在是成功精子的标志。 过度活跃。本申请的总体目标是阐明CatSper通道复合体是如何 分子的定义和结构的组织，以整合信号转导途径，导致机械- 过度激活所需的体细胞转变。中心假说是CatSper内的钙信号状态 纳米结构域决定了精子的活力和受精率。确定其分子机制的基本原理 CatSper渠道监管是，这一知识可能会提供一个强有力的科学框架，使新的 可以开发药物策略来改变精子的运动能力，从而改变男性的生育能力。此应用程序 建议表征与钙离子结合和膜转运有关的新型CatSper相互作用分子。 ING和显微镜观察准备受精的精子中的钙信号状态。中心假说将 通过追求三个具体目标进行测试：1)确定新型CatSper组件在调节中的功能 通道活性和精子活力；2)确定新分子在纳米结构域形成中的作用；以及 3)阐明精子中实现受精的结构域整合信号通路。电生理学 记录和鞭毛波形分析将被用来评估功能丧失的表型效应 通道活动和精子活动率。对于第二个目的，生精细胞的阶段与敲击不同- OUT模型将用于确定它们在通道复合体组装和纳米结构域形成中的作用。对于 第三个目标，将使用运动相关超分辨和原位分子成像方法进行研究 已证实具有活力和生育能力的单个精子细胞的信号状态和结构域组织。 申请人认为，该研究方案具有创新性，因为它侧重于新的CatSper组件 使用新的动物模型测试信令域组织与通道活动耦合的原始概念 在调节钙信号方面，并将新的方法纳入精子生物学领域。拟议的研究 具有重要意义，因为它有望为CatSper通道活动和精子提供新的机械性见解 准备受精的精子的运动调节和分子特性。归根结底，这些知识已经 在辅助生殖、男性不育诊断、 以及制定新的避孕措施目标。