Investigating the role of bromodomain-containing proteins in the production of viable spermatozoa and male fertility

研究含溴结构域蛋白在活精子产生和男性生育能力中的作用

基本信息

批准号：
10398876
负责人：
Paula Elaine Cohen
金额：
$ 38.74万
依托单位：
CORNELL UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2021
资助国家：
美国
起止时间：
2021-05-01 至 2026-03-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10398876
关键词：
Alleles Assisted Reproductive Technology Binding Binding Sites Bromodomain Chromatin Chromatin Structure Chromosome Pairing Complex DNA Packaging DNA Polymerase II Data Defect Development Differentiated Gene Ensure Environment Equus caballus Event Failure Fertility Gene Expression Profile Gene Expression Regulation Gene Silencing Genes Genetic Recombination Genetic Transcription Genome Histone Acetylation Histones Human Immunofluorescence Immunologic Infertility Knockout Mice Machine Learning Male Infertility Maps Mediating Meiosis Meiotic Prophase I Modeling Molecular Morphogenesis Morphology Mus Mutant Strains Mice Mutation Nature Nuclear Pathway interactions Pattern Play Process Production Protamines Proteins Repression Role Sampling Seminal fluid Sperm Head Spermatids Spermatocytes Spermatogenesis Spermiogenesis Spo11 protein Stratification Testing Testis Time Transcriptional Regulation base gene repression genetic corepressor genome-wide insight male male fertility men molecular subtypes mutant promoter protein function recruit sperm analysis sperm cell sperm morphology sperm viability tool

项目摘要

Spermatogenesis is a highly unique differentiation process that involves complex mechanisms of gene regulation, particularly at the level of the transcriptional machinery. The complexity of this process is underscored by the fact that 30% of male infertility is attributable to sperm morphology defects and/or poor semen quality. Bromodomain-containing (BD) proteins are critical regulators of transcription, which act by binding acetylated histone residues at their target loci and recruiting the appropriate transcriptional regulators. Mutations in the genes encoding three BD proteins (BRDT, BRD4 and BRWD1), or that of their interacting proteins, result in morphologically abnormal sperm in mice, and have been associated with poor semen quality and infertility in men. We hypothesize that BRDT, BRD4, and BRWD1 play interconnected roles during meiotic prophase I and spermiogenesis, two key stages of spermatogenesis during which stringent transcriptional regulation is exerted, to ensure appropriate transcriptional control and chromatin compaction leading to the production of morphologically normal sperm. We propose the following model: (1) during meiotic prophase I, BRDT ensures appropriate temporo-spatial control of transcriptional repression to allow for the events of recombination and synapsis; (2) then, upon entry into spermiogenesis, BRDT aids in chromatin compaction during histone-to-protamine exchange, by shutting down transcription across the genome; (3) at the same time, BRD4 and BRWD1 are required to overcome the transcriptional silencing imposed by BRDT specifically at genes essential for spermatid development. Studies herein will test this integrated model of BD action in mice and men. In Aim 1, we will elucidate the role of BRDT in mediating progressive transcriptional shut down during meiotic prophase I using a mouse mutant lacking Brdt. We will examine meiotic progression in wildtype and mutant spermatocytes, and will define the genome-wide distribution of BD proteins and components of the transcriptional machinery. We will ask whether BRDT function is dependent on synapsis or recombination. In Aim 2, we will explore the role of BRWD1 and BRD4 in overcoming BRDT-mediated repression and ensuring expression of critical spermatid differentiation genes. We will investigate how BD proteins co-operate to ensure the correct chromatin environment is in place to allow for the progressive nuclear compaction that arises due to the histone-to-protamine exchange. In Aim 3, we will map the genome-wide distribution of BRDT and BRWD1, histone acetylation, and transcription in human testis, with matched analysis of sperm morphology. By combining these data with analysis of sperm chromatin compaction in wildtype and BD knockout mice, we will develop machine learning tools that permit stratification of sperm from infertile men based on BD protein function and chromatin compaction. These studies are the first to elucidate the coordinated roles of BD proteins in ensuring normal sperm morphology and we will apply our understanding of this transcriptional regulation to defining the causes for sperm defects in infertile men.

精子发生是一个高度独特的分化过程，涉及基因的复杂机制调节，特别是在转录机械级别。这个过程的复杂性是强调了30％的男性不育症归因于精子形态缺陷和/或差精液质量。含溴化域的（BD）蛋白是转录的关键调节剂，其作用结合乙酰化组蛋白在其靶基因座的结合，并募集适当的转录调节剂。编码三种BD蛋白（BRDT，BRD4和BRWD1）的基因中的突变，或它们相互作用的突变蛋白质，导致小鼠形态异常的精子，与精液质量差有关和男人不孕。我们假设BRDT，BRD4和BRWD1在减数分裂预言I和精子发生，这是精子发生的两个关键阶段，在此严格发挥转录调节，以确保适当的转录控制和染色质压实导致形态上正常精子的产生。我们提出以下模型：（1）在减数分裂预言I期间，BRDT确保适当的颞空间控制转录抑制允许重组和突触的事件；（2）然后，进入精子发生后，BRDT有助于通过在整个整个过程中关闭转录基因组；（3）同时，需要BRD4和BRWD1来克服转录沉默由BRDT专门针对精子发育必不可少的基因施加。这里的研究将测试小鼠和男性BD动作的综合模型。在AIM 1中，我们将阐明BRDT在中介中的作用在减数分裂预言i期间使用缺乏BRDT的小鼠突变体的进行性转录关闭。我们将检查野生型和突变体精子细胞中的减数分裂进程，并将定义全基因组转录机械的BD蛋白和成分的分布。我们会问是否BRDT 功能取决于突触或重组。在AIM 2中，我们将探索BRWD1和BRD4在克服BRDT介导的抑制并确保表达关键精子分化基因。我们将研究BD蛋白如何合作以确保到位正确的染色质环境以允许由于组蛋白到脑胺的交换而产生的进行性核压实。在AIM 3中，我们将在人睾丸中绘制BRDT和BRWD1，组蛋白乙酰化和转录的全基因组分布，与精子形态的匹配分析。通过将这些数据与精子染色质分析相结合 Wildtype和BD淘汰小鼠的压实，我们将开发机器学习工具以允许分层基于BD蛋白功能和染色质压实的不育男性的精子。这些研究是首先阐明BD蛋白在确保正常精子形态中的协调作用，我们将应用我们对这种转录调节的理解，以定义不育男人的精子缺陷的原因。