Abstract
BackgroundDuring the process of spermatogenesis, male germ cells undergo dramatic chromatin reorganization, whereby most histones are replaced by protamines, as part of the pathway to compact the genome into the small nuclear volume of the sperm head. Remarkably, approximately 90 % (human) to 95 % (mouse) of histones are evicted during the process. An intriguing hypothesis is that post-translational modifications (PTMs) decorating histones play a critical role in epigenetic regulation of spermatogenesis and embryonic development following fertilization. Although a number of specific histone PTMs have been individually studied during spermatogenesis and in mature mouse and human sperm, to date, there is a paucity of comprehensive identification of histone PTMs and their dynamics during this process.ResultsHere we report systematic investigation of sperm histone PTMs and their dynamics during spermatogenesis. We utilized “bottom-up” nanoliquid chromatography–tandem mass spectrometry (nano-LC–MS/MS) to identify histone PTMs and to determine their relative abundance in distinct stages of mouse spermatogenesis (meiotic, round spermatids, elongating/condensing spermatids, and mature sperm) and in human sperm. We detected peptides and histone PTMs from all four canonical histones (H2A, H2B, H3, and H4), the linker histone H1, and multiple histone isoforms of H1, H2A, H2B, and H3 in cells from all stages of mouse spermatogenesis and in mouse sperm. We found strong conservation of histone PTMs for histone H3 and H4 between mouse and human sperm; however, little conservation was observed between H1, H2A, and H2B. Importantly, across eight individual normozoospermic human semen samples, little variation was observed in the relative abundance of nearly all histone PTMs.ConclusionIn summary, we report the first comprehensive and unbiased analysis of histone PTMs at multiple time points during mouse spermatogenesis and in mature mouse and human sperm. Furthermore, our results suggest a largely uniform histone PTM signature in sperm from individual humans.Electronic supplementary materialThe online version of this article (doi:10.1186/s13072-016-0072-6) contains supplementary material, which is available to authorized users.
Highlights
During the process of spermatogenesis, male germ cells undergo dramatic chromatin reorganization, whereby most histones are replaced by protamines, as part of the pathway to compact the genome into the small nuclear volume of the sperm head
Given the unique progression of spermatogenesis that encompasses a range of processes driven by profound chromatin changes, our aim is to create a comprehensive characterization of histone post-translational modification (PTM) during mouse spermatogenesis (Fig. 1a), including in meiotic spermatocytes, round spermatids, elongating/ condensing spermatids, and mature sperm
While our approach does not provide information regarding the localization of histones in male germ cells, it does lend support to this hypothesis as we found that the relative abundance of specific histone PTMs on H3 and H4 is overall highly consistent between different individual humans
Summary
During the process of spermatogenesis, male germ cells undergo dramatic chromatin reorganization, whereby most histones are replaced by protamines, as part of the pathway to compact the genome into the small nuclear volume of the sperm head. The nuclear organization and chromatin structure of male germ cells undergo a dramatic and highly unique reorganization This initiates with meiotic division of diploid male spermatogonia to produce four haploid round spermatids, which undergo spermiogenesis, comprising compaction and condensation, cytoplasmic shedding, acrosome formation, and nuclear elongation. During these stages, nucleosome acetylation is increased in round and elongating spermatids, which relaxes and increases accessibility of chromatin to facilitate histone eviction [1, 2]. A full understanding of the paternal epigenome is important from a clinical and translational perspective, as increasing numbers of couples utilize assisted reproductive technologies (ART), including in vitro fertilization (IVF) and intracytoplasmic sperm injection (ICSI), to conceive children
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