The origin and compartmentalization of core somatic histones and GSTO2 in sperm and their role in fertilization

精子中核心体细胞组蛋白和 GSTO2 的起源和划分及其在受精中的作用

• 批准号: RGPIN-2017-05305
• 负责人: Oko, Richard
• 金额: $ 2.91万
• 依托单位: Queen's University
• 依托单位国家: 加拿大
• 项目类别: Discovery Grants Program - Individual
• 财政年份: 2019
• 资助国家: 加拿大
• 起止时间: 2019-01-01 至 2020-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=688102
• 关键词: origin; compartmentalization; core; somatic; histones

Fertilization is the cornerstone of development where the two differentiated cells, sperm and egg, set the stage for embryogenesis, for which competency of the gametes is essential. Yet only rudimentary knowledge is available on the molecules and mechanisms involved in sperm-egg interactions leading to zygotic development. Our long term goal is to gain an understanding of these mechanisms. Our approach is to molecularly dissect sperm compartments, such as the acrosome and perinuclear theca, that are implicated in the fertilization process. The identities of these proteins provide clues to their significance and lead to the design of specific probes and experiments to test their predicted fertilization functions. This strategy has been successful as we have discovered novel sperm proteins that are implicated in the penetration of the egg and its activation during fertilization, leading to the development of biomarkers and synthetic molecules for the detection and treatment of infertility. Understanding the events that occur when an egg is first fertilized leads to application of how we can influence those events and improve reproduction in livestock and humans.*******Utilizing this proteomic approach has led my lab to propose that the perinuclear theca (PT), a proteinaceous cap surrounding the sperm nucleus, is divided into two functional regions: the subacrosomal part, containing proteins involved in acrosomal assembly and the postacrosomal part, containing proteins involved in sperm/egg interactions during fertilization. The major goal of this proposal is to determine the significance of several key sperm PT proteins we have identified: namely, a group of core somatic histones (H3, H2B, H2A and H4), whose unprecedented cytosolic location in the postacrosomal region of the sperm head suggests a role in chromatin remodeling of the male pronucleus during fertilization and Glutathione-S-Transferase Omega 2 (GSTO2), whose similar location suggests a role in regulating the oxidative and reductive state of covalently-linked molecules, especially during the disassembly of the sperm nucleus during fertilization. By functional deactivation of these proteins we will test their proposed mechanisms of action during fertilization and determine if they are required for zygotic development.*******The four specific hypotheses to be tested are: 1. The presence of Core histones and GSTO2 in the PT is a common eutherian sperm feature. 2. Core histones and GSTO2 are synthesized de novo during spermiogenesis and assembled as part of the PT by the microtubular manchette. 3. PT core histones dissolve in the ooplasm during fertilization and contribute by stabilizing the DNA of the decondensing sperm nucleus in preparation for pronuclear formation. 4. PT anchored GSTO2 dissolves in the ooplasm during fertilization and utilizes glutathione in the reduction of S-S bonds between protamines to decondense the sperm nucleus.***

受精是发育的基石，其中两个分化的细胞，精子和卵子，为胚胎发生奠定了基础，配子的能力是必不可少的。然而，只有基本的知识是可利用的分子和机制，涉及精卵相互作用，导致合子的发展。我们的长期目标是了解这些机制。我们的方法是从分子水平上解剖精子间室，如顶体和核周膜，它们与受精过程有关。这些蛋白质的身份提供了线索，其意义，并导致特定的探针和实验的设计，以测试其预测的受精功能。这种策略已经取得了成功，因为我们发现了与卵子的穿透及其在受精过程中的激活有关的新型精子蛋白，从而开发了用于检测和治疗不孕症的生物标志物和合成分子。了解卵子第一次受精时发生的事件，可以应用我们如何影响这些事件并改善牲畜和人类的繁殖。利用这种蛋白质组学方法，我的实验室提出，核周膜（PT），围绕精子核的蛋白质帽，分为两个功能区域：顶体下部分，包含参与顶体组装的蛋白质和顶体后部分，包含参与受精过程中精子/卵子相互作用的蛋白质。该提案的主要目标是确定我们已经确定的几种关键精子PT蛋白的重要性：即一组核心体细胞组蛋白（H3，H2 B，H2 A和H4），其在精子头部顶体后区域的前所未有的胞质位置表明在受精和谷胱甘肽-S-转移酶ω 2（GSTO 2）期间雄性原核的染色质重塑中的作用，其相似的位置表明其在调节共价连接分子的氧化和还原状态中的作用，特别是在受精过程中精子核的解体期间。通过这些蛋白质的功能失活，我们将测试它们在受精过程中的作用机制，并确定它们是否是合子发育所必需的。需要检验的四个具体假设是：1。核心组蛋白和谷胱甘肽S-转移酶2在PT中的存在是一种常见的真兽精子特征。2.核心组蛋白和GSTO 2在精子发生过程中重新合成，并通过微管柄组装成PT的一部分。3. PT核心组蛋白在受精过程中溶解在卵质中，并通过稳定解致密精子核的DNA为原核形成做准备而做出贡献。4. PT锚定的GSTO 2在受精过程中溶解在卵质中，并利用谷胱甘肽还原鱼精蛋白之间的S-S键以使精子核去致密化。