Gamete Interactions in Caenorhabditis elegans

描述（由申请人提供）：受精是具有重要医学、社会和经济影响的生物过程。通过广泛的研究，人们对受精事件有了一些详细的了解。然而，这些事件的分子基础通常仍然难以捉摸。之前大多数有关受精的工作都依赖于生化和免疫学方法。我们的工作在将经典遗传分析应用于这一重要研究领域方面具有开创性。我的实验室一直在帮助开创性地使用线虫来解决精卵相互作用的机制。为线虫开发的许多遗传和分子工具在传统上用于研究受精的其他生物体中不可用或很难利用。线虫最显着的优势之一是我们能够分离和维持影响精子或卵子而不影响其他细胞的突变体。此前，通过对不育突变体的研究，我们已经确定了秀丽隐杆线虫中生产性配子相互作用所需的一些第一个精子分子。这些精子分子包括 SPE-9 和 SPE-38。 SPE-9 是一种精子表面跨膜分子，其胞外结构域包含 10 个表皮生长因子 (EGF) 样重复序列。我们假设 SPE-9 充当卵子表面精子受体的配体。 spe-38 基因编码新型四次跨膜分子。基于蛋白质定位和遗传相互作用研究，我们假设 SPE-38 调节钙通道 (SPE-41/TRP-3) 和其他精子分子的定位和/或活性。我们最近确定了秀丽隐杆线虫受精所需的第一个卵分子。 Egg-1 和 Egg-2 基因编码蛋表面低密度脂蛋白 (LDL) 受体重复序列的蛋白质。我们假设 EGG-1 和 EGG-2 在受精过程中作为精子的卵子表面受体半冗余地发挥作用。本提案的目的是通过进行以下实验目标来进一步了解线虫受精：1）通过开发试剂来测试它们和/或其他分子之间潜在的配体-受体相互作用，进一步表征SPE-9和EGG-1/EGG-2在受精中的作用。 2) 通过蛋白质相互作用研究和诱变研究 SPE-38 和 SPE-41/TRP-3 之间的联系，更好地了解 SPE-38 和 SPE-41/TRP-3 在受精中的功能。 3)克隆精子功能基因spe-36以确定其分子性质和在受精中的作用。 spe-36 基因由与 spe-9、spe-38 和 spe-41/trp-3 突变体相同的不育突变体表型定义。这项工作将补充其他生物体的生育力研究，并提供对细胞间相互作用机制和生殖策略多样性的见解。公共健康相关性：尽管对许多物种的受精事件已有相当详细的描述，但该过程的分子基础尚不清楚。我们正在鉴定秀丽隐杆线虫模型系统中受精的分子和分子机制。这项工作将补充其他物种受精的研究，并最终有助于更好地了解不孕的原因、生殖策略的多样性和新避孕药的设计。

DESCRIPTION (provided by applicant): Fertilization is biological process with important medical, social and economic implications. From extensive study, the events of fertilization are known in some detail. However, the molecular underpinnings of these events generally remain elusive. Most previous work on fertilization has relied on biochemical and immunological approaches. Our work is groundbreaking in the application of classic genetic analysis to this vital area of research. My lab has been helping to pioneer the use of C. elegans for addressing the mechanisms of sperm-egg interactions. Many of the genetic and molecular tools developed for C. elegans are not available or are very difficult to utilize in other organisms traditionally used for studying fertilization. One of the most significant advantages of C. elegans is our ability to isolate and maintain mutants that affect sperm or eggs and no other cells. Previously, through the study of sterile mutants, we have identified some of the first sperm molecules required for productive gamete interactions in C. elegans. These sperm molecules include SPE-9 and SPE-38. SPE-9 is a sperm surface transmembrane molecule with an extracellular domain that contains ten epidermal growth factor (EGF)-like repeats. We hypothesize that SPE-9 functions as a ligand for an egg surface sperm receptor. The spe-38 gene encodes a novel four pass transmembrane molecule. Based on protein localization and genetic interaction studies, we hypothesize that SPE-38 regulates the localization and/or activity of a calcuim channel (SPE-41/TRP-3) and other sperm molecules. We recently identified the first egg molecules required for fertilization in C. elegans. The egg-1 and egg-2 genes encode egg surface Low Density Lipoprotein (LDL) receptor repeat-containing proteins. We hypothesize that EGG-1 and EGG-2 function semi-redundantly as egg surface receptors for sperm during fertilization. The goal of this proposal is to further our understanding of fertilization in C. elegans by conducting the following experimental aims: 1) to further characterize the roles of SPE-9 and EGG-1/EGG-2 in fertilization by developing reagents to test for potential ligand-receptor interactions between them and/or other molecules. 2) to gain a better understanding SPE-38 and SPE-41/TRP-3 function in fertilization by investigating the connection between them through protein interaction studies and mutagenesis. 3) to clone the sperm function gene spe-36 in order to determine its molecular nature and role in fertilization. The spe-36 gene is defined by a sterile mutant phenotype that is identical to spe-9, spe-38 and spe-41/trp-3 mutants. This work will complement fertility studies in other organisms as well as provide insights into the mechanisms of cell-cell interactions and the diversity of reproductive strategies. PUBLIC HEALTH RELEVANCE: Although the events of fertilization are fairly well described for a number of species, the molecular underpinnings of the process are not well understood. We are identifying the molecules and molecular mechanisms of fertilization in the model system Caenorhabditis elegans. This work will complement studies of fertilization in other species and could eventually lead to a better understanding of the causes of infertility, the diversity of reproductive strategies and the design of new contraceptives.