Genome-wide chromatin analysis in mature mouse and human spermatozoa.
At the end of mammalian spermatogenesis, chromatin in differentiating germ cells is extensively remodeled, with the majority of nucleosomes being removed and ultimately exchanged by highly basic proteins named protamines. Residual nucleosomes are, to various degrees, retained at regulatory sequences in human and mouse sperm. Moreover, certain histone variants and modifications remain present in regulatory sequences of subsets of genes in spermatozoa, providing opportunities for paternal inheritance of chromatin states and epigenetic control of gene expression in the subsequent generation. Here we describe in detail a method that enables the generation of soluble chromatin samples from mouse and human spermatozoa within 1 d. These samples are amendable to chromatin immunoprecipitation and high-throughput sequencing of nucleosome-associated genomic DNA, which require several additional days. We also provide computational scripts that allow straightforward analysis of large genome-wide data sets by biologists with limited computational experience. This protocol will facilitate studies of mechanisms of chromatin remodeling and epigenetic reprogramming during spermatogenesis and of paternal epigenetic inheritance. Similarly, it will help in the study of the causes of human male infertility.
1] Friedrich Miescher Institute for Biomedical Research (FMI), Basel, Switzerland. [2] [3].
Mizue
M
Hisano
Serap
S
Erkek
Sophie
S
Dessus-Babus
Liliana
L
Ramos
Michael B
MB
Stadler
Antoine H F M
AH
Peters
D000818:Animals; D002843:Chromatin / Q000737:chemistry*; D042002:Chromatin Assembly and Disassembly; D047369:Chromatin Immunoprecipitation / Q000379:methods*; D004247:DNA / Q000737:chemistry; D044127:Epigenesis, Genetic; D059014:High-Throughput Nucleotide Sequencing; D006657:Histones / Q000737:chemistry / Q000378:metabolism; D006801:Humans; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D009707:Nucleosomes / Q000378:metabolism; D015203:Reproducibility of Results; D013094:Spermatozoa / Q000648:ultrastructure*
D002843:Chromatin; D006657:Histones; D009707:Nucleosomes; D004247:DNA