MECHANISMS OF MAMMALIAN FERTILIZATION/POLYSPERMY BLOCK

哺乳动物受精/多精阻断的机制

• 批准号: 6387528
• 负责人: GREGORY S. KOPF
• 金额: $ 31.39万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 1997
• 资助国家: 美国
• 起止时间: 1997-04-01 至 2002-09-18
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6387528
• 关键词: G protein; antisense nucleic acid; calmodulin dependent protein kinase; cell cell interaction; chorionic gonadotropin; confocal scanning microscopy; egg /ovum; fertilization; genetically modified animals; gonadotropins; high performance liquid chromatography; integrins; laboratory mouse; oligonucleotides; sperm

Fertilization of mouse eggs initiates a cascade of signaling events that include a transient rise in intracellular Ca2+, the stimulation of cortical granule exocytosis that results in modifications of the zona pellucida, recruitment of maternal mRNAs, and completion of the meiotic cell cycle and conversion to a mitotic cell cycle. These events constitute a process termed "egg activation". By analogy to differentiated somatic cells, the sperm may initiate these events of egg activation using G protein-coupled signal transduction pathways. the initial events of sperm-egg plasma membrane interaction that ultimately lead to intracellular signaling may involve adhesion and membrane fusion processes mediated by a sperm ligand and an egg receptor. One aim of this proposal will examine the role of the potential downstream effector for Ca2+ action, the calmodulin-dependent protein kinase II, on specific events of egg activation. This will be done by using a constitutively active form of this kinase, as well as a battery of specific inhibitors of this enzyme. Another aim will address directly the role of G proteins in sperm-induced egg activation by using an antisense approach to ablate the function of specific egg G protein subunits and then assessing this effect on events of egg activation. The last aim will use a sperm- specific protein called fertilin that has been implicated as a specific ligand/adhesion molecule involved in sperm-egg plasma membrane interactions to identify by chemical crosslinking experiments the egg binding protein for fertilin. Since fertilin contains a disintegrin domain an egg integrin may serve as the binding protein for fertilin. The role of an egg integrin in mediating this process will be examined using a transgenic antisense approach in which specific egg integrins are targeted and the effects on sperm binding and egg activation are assessed. Results of these studies focus on the molecular basis of sperm-egg interaction and egg activation and will translate directly to the clinical setting of assisted reproductive technologies. Moreover, these studies address fundamental questions in cell biology, namely, cell adhesion-induced signaling between heterologous cells that relate to questions of regulated exocytosis, cell cycle, and translational control.

小鼠卵细胞受精引发一系列信号传导事件 的 包括细胞内Ca 2+的瞬时升高， 皮质颗粒胞吐，导致 zona 透明膜，母体mRNA的募集，以及 减数分裂 细胞周期和转化为有丝分裂细胞周期。 这些事件 这一过程被称为“卵激活”。 通过类似于 在体细胞分化的过程中，精子可能会引发这些事件。 鸡蛋 使用G蛋白偶联的信号转导途径激活。 的 精子-卵子质膜相互作用的初始事件， 最终 导致细胞内信号传导可能涉及粘附和膜 融合 由精子配体和卵子受体介导的过程。 一个目的 的 该提案将审查潜在下游的作用， 执行器 对于Ca 2+作用，钙调蛋白依赖性蛋白激酶II， 具体 卵子激活事件。 这将通过使用 组成型 这种激酶的活性形式，以及一组特定的 抑制剂 这种酶。 另一个目标将直接解决G的作用 蛋白 在精子诱导的卵子激活中， 消融 特定的卵G蛋白亚基的功能，然后评估 这 对卵子激活事件的影响。 最后一个目标是用精子- 一种叫做受精素的特殊蛋白质， 具体 精卵质膜配体/粘附分子 通过化学交联实验确定相互作用， 蛋 受精素结合蛋白。 由于受精素含有 解聚素 卵整合素的结构域可以作为结合蛋白， 受精素 卵整合素在介导这一过程中的作用将是 检查 使用转基因反义方法， 整合素是 有针对性，对精子结合和卵子激活的影响是 评估。 这些研究的结果集中在分子基础上， 精卵相互作用和卵激活，并将翻译 直接向 辅助生殖技术的临床环境。 此外，委员会认为， 这些研究解决了细胞生物学中的基本问题， 即细胞 异源细胞之间的粘附诱导信号传导， 到 调节胞吐、细胞周期和翻译的问题 控制