An evolutionary and proteomic search for human gamete recognition proteins that mediate zona pellucida dissolution

介导透明带溶解的人类配子识别蛋白的进化和蛋白质组学研究

• 批准号: 10370974
• 负责人: WILLIE J SWANSON
• 金额: $ 22.93万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-09-22 至 2023-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10370974
• 关键词: 3-Dimensional; Address; Affinity; Binding; Biochemical; Bioinformatics; Biological Assay; Biology; Characteristics; Chemotaxis; Classification; Codon Nucleotides; Complementary DNA; Data; Databases; Detection; Distant; Egg Proteins; Event; Evolution; Exploratory/Developmental Grant for Diagnostic Cancer Imaging; Female; Fertilization; Frequencies; Future; Genes; Genetic; Genomics; Germ Cells; Goals; Guanine + Cytosine Composition; Homologous Gene; Homologous Protein; Human; Human Genome; Knock-out; Length; Mass Spectrum Analysis; Measures; Mediating; Molecular; Morphology; Mus; N Domain; Oocytes; Performance; Protein Structure Initiative; Proteins; Proteome; Proteomics; RNA Splicing; Reproduction; Research; Resources; Specificity; Sperm Capacitation; Structure; Surface Plasmon Resonance; Technology; Testing; Testis; Transcript; Variant; Zona Pellucida; abalone; algorithm training; base; egg; env Gene Products; experience; extracellular; genetic approach; genomic data; improved; innovation; lysin; male; mammalian genome; molecular phenotype; nanopore; prediction algorithm; predictive test; reproductive; sperm cell; sperm protein; three dimensional structure; transcriptome; transcriptome sequencing; transcriptomics

Project Summary The identification of interacting male-female reproductive proteins is imperative for a molecular understanding of fertilization. The goal of this R21 proposal is to identify a set of human gamete recognition proteins using genomic and proteomic approaches, and then perform evolutionary guided computational bioinformatic searches for a homolog of abalone sperm lysin. It is known from three-dimensional protein crystallographic studies of the major proteins from the mammalian Zona Pellucida (ZP) and abalone Vitelline Envelope (VE) that they contain homologous ZP-N domains. Given the molecular homology of ZP-N domains and overall biochemical similarity of the egg coats, an obvious question to address is whether sperm use similar mechanisms (or homologous proteins) to generate a path through the egg coat. The dissolution of the abalone egg VE by the sperm protein lysin has been extensively characterized. However, it remains unknown how mammalian sperm penetrate egg coats, and no lysin homolog is currently identifiable in any mammalian genome. Our proposal address this question by (1) using long-read sequencing to generate deep coverage of full-length non-chimeric cDNAs for use in sperm proteomics, (2) using these resources as a database to perform detailed bioinformatic analyses aimed at generating a database of putative human gamete recognition proteins and identification of a human homolog of abalone sperm lysin and (3) functional tests of predicted protein's ability to disrupt the mouse ZP. If we identify a human sperm lysin, it will immediately open up new avenues of research into mammalian gamete recognition and how the human sperm passes the ZP. If we are unsuccessful at identifying a human lysin, we will have generated a valuable genomic and proteomic resource database for future studies of human sperm biology.

项目概要 鉴定相互作用的男性-女性生殖蛋白对于分子理解至关重要 的受精。该 R21 提案的目标是使用以下方法识别一组人类配子识别蛋白： 基因组学和蛋白质组学方法，然后进行进化引导的计算生物信息学 寻找鲍鱼精子溶素的同系物。由三维蛋白质晶体学可知 对哺乳动物透明带 (ZP) 和鲍鱼卵黄膜 (VE) 中主要蛋白质的研究 它们含有同源的 ZP-N 结构域。鉴于 ZP-N 结构域和整体的分子同源性 由于卵子外壳的生化相似性，一个明显的问题是精子是否使用相似的卵子外壳。 产生穿过蛋壳的路径的机制（或同源蛋白质）。鲍鱼的溶解 精子蛋白溶素对卵子 VE 的影响已得到广泛表征。然而，目前尚不清楚如何 哺乳动物精子穿透卵膜，目前在任何哺乳动物中都没有发现溶素同源物 基因组。我们的建议通过（1）使用长读长测序来生成深度覆盖来解决这个问题 用于精子蛋白质组学的全长非嵌合 cDNA，(2) 使用这些资源作为数据库来 进行详细的生物信息分析，旨在生成假定的人类配子识别数据库 鲍鱼精子溶素的蛋白质和人类同源物的鉴定以及（3）预测的功能测试 蛋白质破坏小鼠 ZP 的能力。如果我们识别出人类精子溶素，它将立即开辟新的领域 哺乳动物配子识别以及人类精子如何通过 ZP 的研究途径。如果我们是 如果未能成功识别人类溶素，我们将产生宝贵的基因组和蛋白质组资源 人类精子生物学未来研究的数据库。