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Mechanisms of sperm chromatin remodeling in vivo and in vitro

精子染色质体内外重塑机制

基本信息

批准号：
10753662
负责人：
Saher Sue Hammoud
金额：
$ 53.96万
依托单位：
UNIVERSITY OF MICHIGAN AT ANN ARBOR
依托单位国家：
美国
项目类别：
财政年份：
2023
资助国家：
美国
起止时间：
2023-08-15 至 2027-05-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10753662
关键词：
ATP phosphohydrolase Affect Antibodies Appearance Back Binding Biochemical Biological Assay Cells ChIP-seq Chromatin Chromatin Remodeling Factor Complex Couples DNA DNA Packaging Data Deposition Development Diagnostic Epitopes Etiology Event Excision Fertility Generations Genetic Genome Genomic Segment Genotype Germ Cells Global Change Goals Haploidy Histones ISWI Immunoprecipitation In Vitro Infertility Investigation Kinetics Knock-out Knowledge Label Mass Spectrum Analysis Mediating Meiosis Methods Mitosis Modeling Molecular Molecular Chaperones Mus Nucleosomes Oocytes PRM2 gene Pattern Phenotype Physical condensation Population Positioning Attribute Process Production Protamines Protein Subunits Proteins Publishing Reagent Recombinant Proteins Role SMARCA5 gene Sexual Reproduction Slide Somatic Cell Spermatids Spermiogenesis Testing Testis Woman biochemical tools biophysical properties biophysical tools chromatin remodeling effective therapy egg genomic tools in vivo insight laser tweezer loss of function men mutant novel optic tweezer programs protamine 1 public health relevance reconstitution reproductive fitness single molecule sperm cell

项目摘要

Abstract Unlike oocytes (and all somatic cells), which package their DNA with histones, the DNA of mature sperm is bound by protamines, a highly basic and rapidly evolving protein that is essential for compaction of the paternal genome. This differential packaging of egg and sperm DNA traces back >500 million years, yet our understanding of the principles of genome organization remains poorly understood. To examine this fundamental yet poorly understood process we have developed novel genetic and biophysical tools to identify proteins implicated in the histone-to-protamine exchange and to dissect the molecular mechanisms of chromatin reorganization during spermiogenesis. In preliminary studies we developed epitope-tagged protamine 1 and 2 mice and relevant reagents such as specific antibodies. Immunoprecipitation followed by mass spectrometry has identified multiple potential chromatin remodelers/chaperones, which we will functionally characterize in this proposed study, both in vivo and in vitro. Since protamine packaging results in a super-condensed chromatin state, which does not easily lend itself to mechanistic investigations in vivo, we have developed novel methods to purify protamines from sperm to use in both in vitro DNA curtain assays and in vitro reconstitution assays of the histone-to-protamine exchange process. The overall goal of this proposal is to apply our diverse suite of genetic, molecular, and biophysical tools to gain mechanistic insights into normal sperm chromatin packaging, which is essential for genome fidelity and reproductive fitness of the species.

摘要