Characterization of BRD4 during Mammalian Postmeiotic Sperm Development

Abstract

During spermiogenesis, the postmeiotic phase of mammalian spermatogenesis, transcription is progressively repressed as nuclei of haploid spermatids are compacted through a dramatic chromatin reorganization involving hyperacetylation and replacement of most histones with protamines. Although BRDT functions in transcription and histone removal in spermatids, it is unknown whether other BET family proteins play a role. Immunofluorescence of spermatogenic cells revealed BRD4 in a ring around the nuclei of spermatids containing hyperacetylated histones. The ring lies directly adjacent to the acroplaxome, the cytoskeletal base of the acrosome, previously linked to chromatin reorganization. The BRD4 ring does not form in acrosomal mutant mice. Chromatin immunoprecipitation followed by sequencing in spermatids revealed enrichment of BRD4 and acetylated histones at the promoters of active genes. BRD4 and BRDT show distinct and synergistic binding patterns, with a pronounced enrichment of BRD4 at spermatogenesis-specific genes. Direct association of BRD4 with acetylated H4 decreases in late spermatids as acetylated histones are removed from the condensing nucleus in a wave following the progressing acrosome. These data provide evidence of a prominent transcriptional role for BRD4 and suggest a possible removal mechanism for chromatin components from the genome via the progressing acrosome as transcription is repressed and chromatin is compacted during spermiogenesis.

Supplemental material for this article may be found at http://dx.doi.org/10.1128/MCB.01328-14.

ACKNOWLEDGMENTS

We thank the members of the Berger lab, especially Parisha Shah, for all their support and advice. We thank Jérôme Govin for his guidance. We thank Saadi Khochbin for his advice and sharing his BRDT ChIP-seq data. We thank Jan Van Deursen for sharing his Hrb^−/− mice. We thank James Bradner for sharing JQ1-biotin. We thank Andrea Stout of the Cell and Developmental Microscopy Core for her help with IF imaging. We thank Joseph Grubb and Jonathan Schug of the University of Pennsylvania Functional Genomics Core for their help with ChIP-seq.

Support to J.M.B. was from the T32 Genetics Training Grant at the University of Pennsylvania (GM008216). Support to S.L.B. was from NIH grants GM055360 and U54-HD068157. B.A.G. acknowledges funding from NIH grant GM110174 and Innovator grant DP2OD007447 from the Office of the Director. R.G.M. was supported by NIH grants R01HD048837 and U54HD068157.

We have no competing financial interests.