I-40: Male Genome Programming, Infertility and Cancer

Background: During male germ cells differentiation, genomewide re-organizations and highly specific programming of the male genome occur. These changes not only include the large-scale meiotic shuffling of genes, taking place in spermatocytes, but also a complete “re-packaging” of the male genome in post meiotic cells, leading to a highly compacted nucleo-protamine structure in the mature sperm cells. This process is essential to protect the male genome for its journey out of the male body through the hostile environment of the female organism, as well as to prepare its rapid activation after fertilization. Although it is a critical step for successful reproduction in most species, nearly nothing is known on the molecular basis of this process. Our objective is to identify molecular actors driving male genome programming and functionally characterize their roles. Materials and Methods: A combination of strategies, including the generation of several mouse models (RousseauxSBRM11_ pmid21208144), molecular, structural and proteomic approaches, as well as genome-wide analyses of male germ cells at different stages using chromatin immunoprecipitation (ChIP) and new generation sequencing (NGS) are used. The latter also involve the design and usage of dedicated bioinformatics analyses, which we are developing. Results: Early post-meiotic cells, round spermatids, inherit a haploid genome associated with histones, in a somatic-like nucleosome based chromatin, which then undergoes a genome-wide reorganization associated with the replacement of histones by protamines. Our work during the last 12 years has unravelled several key molecular mechanisms involved in this formidable remodelling of the genome. Our initial investigations were based on two observations. First, several specific histone variants are expressed and incorporated into the chromatin of male germ cells. Second, a genome-wide histone hyperacetylation wave occurs in elongating spermatids (HazzouriEJCB2000_ pmid11152286; FaureMHR2003_pmid14614037), which precedes their removal and replacement by transition proteins and protamines. Combining proteomic and bioinformatics screens (RousseauxSBiRM2012_ pmid22788531), we identified several candidate factors. In depth characterization of these factors involved structural and molecular approaches, as well as the use of genetically modified mouse models. One of our major findings regarding the role of histone acetylation is that Brdt, a double bromodomain containing testis specific member of the BET family, guided by acetylated chromatin, is playing essential roles in the stage-specific programming of the male genome (Pivot- PajotMCB2003_pmid12861021; MoriniereNature09_pmid19794495; GaucherEMBOJ12_pmid22922464). Recently, our investigation of the role of the histone acetyl transferases (HAT) p300 and CBP in postmeiotic cells, using a double conditional KO mouse model, highlighted their role in the control of a late post-meiotic gene expression program involved in the metabolic remodelling of male germ cells (Boussouar_ Andrology14_pmid24522976). We also unravelled the role of testisspecific variants in the programming of pericentric regions, as well as in specifically preparing chromatin for post-meiotic packaging, and for post-fertilization events (GovinJCB07_pmid17261847; WuJRD08_ pmid18703863; MontellierGenesDev2013_pmid23884607). Our lab is also involved in collaborative studies with Pr Y.M. Zhao’s lab, leading to the discovery of a large panel of new histone modifications, hugely enlarging the so-called “histone code”, and increasing our understanding of their role in male genome programming (TanCell11_pmid21925322; DaiNCB14_pmid24681537). Conclusion: We are now in a position to propose the first molecular models explaining male genome programming (Goudarzi_ JMB2014_pmid24613302). Medical applications are described below in “Project application and usage”.