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Differential expression and sex chromosome association of CHD3/4 and CHD5 during spermatogenesis.

ABSTRACT: ATP-dependent nucleosome remodelers of the CHD family play important roles in chromatin regulation during development and differentiation. The ubiquitously expressed CHD3 and CHD4 proteins are essential for stem cell function and serve to orchestrate gene expression in different developmental settings. By contrast, the closely related CHD5 is predominantly expressed in neural tissue and its role is believed to be restricted to neural differentiation. Indeed, loss of CHD5 contributes to neuroblastoma. In this study, we first demonstrate that CHD5 is a nucleosome-stimulated ATPase. We then compare CHD3/4 and CHD5 expression in mouse brain and show that CHD5 expression is restricted to a subset of cortical and hippocampal neurons whereas CHD3/4 expression is more widespread. We also uncover high levels of CHD5 expression in testis. CHD5 is transiently expressed in differentiating germ cells. Expression is first detected in nuclei of post-meiotic round spermatids, reaches a maximum in stage VIII spermatids and then falls to undetectable levels in stage IX spermatids. Surprisingly, CHD3/4 and CHD5 show complementary expression patterns during spermatogenesis with CHD3/4 levels progressively decreasing as CHD5 expression increases. In spermatocytes, CHD3/4 localizes to the pseudoautosomal region, the X centromeric region and then spreads into the XY body chromatin. In postmeiotic cells, CHD5 colocalises with macroH2A1.2 in association with centromeres and part of the Y chromosome. The subnuclear localisations of CHD4 and CHD5 suggest specific roles in regulation of sex chromosome chromatin and pericentromeric chromatin structure prior to the histone-protamine switch.

SUBMITTER: Bergs JW

PROVIDER: S-EPMC4029951 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Deleted in AZoospermia Associated Protein 1 (DAZAP1) is a ubiquitous hnRNP protein that has been implicated in RNA transcription, splicing, and translation. It is highly expressed in testes, predominantly in late stage spermatocytes and post-meiotic spermatids. Dazap1 deficiency in mice results in growth retardation and spermatogenic arrest. The gene produces two major transcripts of 2.4 and 1.8 kb, designated Dazap1-L and Dazap1-S, respectively. Results of our previous RNA in situ hybridization and immunostaining suggested translational regulation of the Dazap1 transcripts during spermatogenesis. The main objectives of the study were to determine the origin of the two Dazap1 transcripts and to investigate whether they were similarly translated. Our Northern and 3' RACE analyses showed that the two transcripts were generated through alternative polyadenylation. In mouse testes, the levels of both transcripts were low at postnatal day 12 (P12), increased significantly at P18, and reached maximum at P27. Sucrose gradient analyses showed that at P12 both transcripts were actively translated. Afterward, an increasing portion of Dazap1-S became associated with the translationally inactive mRNPs, and the translational repression was accompanied by an increase in the length of its poly(A) tail. A much smaller portion of Dazap1-L was also sequestered to mRNPs as testes matured, but there was no changes in its poly(A) tail length. Using RNA pull-down followed by mass spectrometry, we identified DAZL, a germ-cell specific translation regulator, as one of the proteins that bound to the 3'UTR region specific for Dazap1-L. We further showed that DAZL preferentially bound to Dazap1-L in testis lysates and stimulated the translation of a reporter gene carrying Dazap1-L 3'UTR. In summary, our study shows that the translation of the two Dazap1 transcripts is differentially regulated. It also provides a new example of translational repression associated with poly(A) tail elongation during spermatogenesis.

Project description:Hybrid sterility is a complex phenotype that can result from the breakdown of spermatogenesis at multiple developmental stages. Here, we disentangle two proposed hybrid male sterility mechanisms in the house mice, Mus musculus domesticus and M. m. musculus, by comparing patterns of gene expression in sterile F1 hybrids from a reciprocal cross. We found that hybrid males from both cross directions showed disrupted X chromosome expression during prophase of meiosis I consistent with a loss of meiotic sex chromosome inactivation (MSCI) and Prdm9-associated sterility, but that the degree of disruption was greater in mice with an M. m. musculus X chromosome consistent with previous studies. During postmeiotic development, gene expression on the X chromosome was only disrupted in one cross direction, suggesting that misexpression at this later stage was genotype-specific and not a simple downstream consequence of MSCI disruption which was observed in both reciprocal crosses. Instead, disrupted postmeiotic expression may depend on the magnitude of earlier disrupted MSCI, or the disruption of particular X-linked genes or gene networks. Alternatively, only hybrids with a potential deficit of Sly copies, a Y-linked ampliconic gene family, showed overexpression in postmeiotic cells, consistent with a previously proposed model of antagonistic coevolution between the X- and Y-linked ampliconic genes contributing to disrupted expression late in spermatogenesis. The relative contributions of these two regulatory mechanisms and their impact on sterility phenotypes await further study. Our results further support the hypothesis that X-linked hybrid sterility in house mice has a variable genetic basis, and that genotype-specific disruption of gene regulation contributes to overexpression of the X chromosome at different stages of development. Overall, these findings underscore the critical role of epigenetic regulation of the X chromosome during spermatogenesis and suggest that these processes are prone to disruption in hybrids.

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Differential expression and sex chromosome association of CHD3/4 and CHD5 during spermatogenesis.

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