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Secretory Vesicle Function During Spermatogenesis & Fertilization in C. elegans

精子发生过程中的分泌囊泡功能

基本信息

批准号：
7920958
负责人：
STEVEN W. L'HERNAULT
金额：
$ 28.78万
依托单位：
EMORY UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2007
资助国家：
美国
起止时间：
2007-09-10 至 2012-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7920958
关键词：
Accounting Acrosome Address Affect Animals Binding Biogenesis Biology Bone Diseases Caenorhabditis elegans Cell membrane Cell surface Cells Cellular biology Child Couples Data Defect Development Disease Ensure Event Fanconi Syndrome Fertilization Genes Germ Cells Goals Human Immune System Diseases Infertility Kidney Ligands Malignant Neoplasms Mammals Membrane Protein Traffic Men&apos s Role Molecular Mutation Nature Nerve Degeneration Neurodegenerative Disorders Organelles Osteogenesis Osteoporosis Pharmacology Phenotype Physiological Play Preparation Process Proteins Proton Pump Public Health Regulation Renal function Role Secretory Vesicles Sperm Motility Spermatids Spermatogenesis System Testing Ubiquitin Ubiquitination Work base developmental genetics egg falls human disease infancy innovation loss of function meetings men mutant receptor function sperm cell sperm function ubiquitin-protein ligase vacuolar H+-ATPase zygote

项目摘要

DESCRIPTION (provided by applicant): The long-term goal of this work is to determine the molecular and cellular mechanisms required for sperm-egg fusion during fertilization. This proposal addresses how a sperm secretory vesicle (acrosome) forms properly so that it can fuse with and create a fertilization-competent cell surface on the spermatozoon. Sperm secretory vesicles acquire an acidic internal pH during their maturation due to vacuolar (V-) ATPase activity, and this occurs in many animals, including humans. However, the mechanisms that regulate V-ATPase activity in sperm secretory vesicles are not understood, mostly because a spermatogenesis system suitable for its analysis had not been identified. Our preliminary data show that Caenorhabditis elegans spermatogenesis is ideally suited for analyzing the V-ATPase role and regulation during biogenesis and function of sperm secretory vesicles. In particular, mutations in the spe-5 gene alter a V-ATPase subunit that is spermatogenesis-specific, so acidification of sperm secretory vesicles can be studied in the absence of somatic V-ATPase defects, which are usually lethal. The spe-16 gene encodes a spermatogenesis-specific ubiquitin E3 ligase and how it participates in developmentally controlled acidification of sperm secretory vesicles will be determined. The focused objective of this proposal is to determine how the spe-5 and spe-16 encoded proteins cooperate to regulate internal pH in sperm secretory vesicles and what happens when this regulation is disrupted. We use three specific aims to test the overall hypothesis that the internal pH of sperm secretory vesicles must fall in order for this organelle to fuse with the plasma membrane. Specific aim #1 is to identify the role of the V-ATPase during sperm secretory vesicle maturation. Specific aim #2 is to identify how absence of V-ATPase function affects sperm motility and function during fertilization. Specific aim #3 is to identify why secretory vesicles fail to acidify in spe-16 loss of function mutant sperm. The unusual nature of the spe-16 mutant phenotype ensures it utilizes a previously undescribed way of regulating an animal V-ATPase. Ubiquitin is conjugated to proteins in normal human sperm, including those in the acrosome, and ubiquitination defects occur in infertile men; the role of ubiquitination is not understood in either case. Defects in ubiquitin biology are associated with various cancers, neurodegenerative diseases and many other disease processes. In addition to its role during spermatogenesis, V-ATPase function is defective in diseases that include renal Fanconi syndrome and the bone diseases, infantile ostepetrosis and osteoporosis. This proposal is focused on how sperm develop the ability to fertilize an egg. This is a problem of significant public health impact because ~15% of couples desiring children are infertile and many cases are due to defective sperm. The processes we study, in addition to functioning in sperm, also play a role in proper kidney function and bone formation.

描述（由申请人提供）：这项工作的长期目标是确定受精过程中精卵融合所需的分子和细胞机制。该建议解决了精子分泌囊泡（顶体）如何正确形成，以便它可以与精子融合并在精子上产生受精能力细胞表面。由于液泡（V-）ATP酶活性，精子分泌囊泡在成熟过程中获得酸性内部pH，这发生在许多动物中，包括人类。然而，调节精子分泌囊泡中V-ATP酶活性的机制尚不清楚，主要是因为尚未鉴定出适合其分析的精子发生系统。我们的初步数据表明，秀丽隐杆线虫精子发生是理想的适合于分析V-ATP酶的作用和调节过程中的生物发生和功能的精子分泌囊泡。特别是，突变的spe-5基因改变了一个V-ATP酶亚基，这是精子发生特异性的，所以酸化的精子分泌囊泡可以在体细胞V-ATP酶缺陷，这通常是致命的情况下进行研究。SPE-16基因编码精子发生特异性泛素E3连接酶，它如何参与发育控制的精子分泌囊泡酸化将被确定。该提案的重点目标是确定SPE-5和SPE-16编码的蛋白质如何合作调节精子分泌囊泡的内部pH值，以及当这种调节被破坏时会发生什么。我们使用三个具体的目的来测试的整体假设，即精子分泌囊泡的内部pH值必须下降，以使这个细胞器与质膜融合。具体目标#1是确定V-ATPase在精子分泌囊泡成熟过程中的作用。具体目标#2是确定V-ATP酶功能的缺乏如何影响精子活力和受精过程中的功能。具体目标#3是确定为什么spe-16功能丧失突变精子中分泌囊泡不能酸化。spe-16突变体表型的不寻常性质确保其利用先前未描述的调节动物V-ATP酶的方式。泛素结合在正常人精子中的蛋白质上，包括那些在顶体中的蛋白质，并且在不育男性中发生泛素化缺陷;在这两种情况下，泛素化的作用都不清楚。泛素生物学缺陷与各种癌症、神经退行性疾病和许多其他疾病过程相关。除了在精子发生过程中的作用外，V-ATP酶功能在包括肾范可尼综合征和骨骼疾病、婴儿骨石化症和骨质疏松症在内的疾病中是有缺陷的。这项提案的重点是精子如何发展使卵子受精的能力。这是一个严重影响公共卫生的问题，因为约15%的想要孩子的夫妇是不育的，许多情况是由于精子缺陷。我们研究的过程除了在精子中发挥作用外，还在适当的肾功能和骨形成中发挥作用。