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    Spermatogenesis & infertility: Genetic and epigenetic regulation during sperm differentiation

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    Project leader

    +33 144412312
    julie.cocquet@inserm.fr

     

    Research Topics

    Human infertility affects ~15% of couples, each partner being equally likely to be affected. Spermatogenesis defects are commonly found in infertile men and the majority is presumed to have a genetic or epigenetic origin; however, the causal genes and mechanisms remain largely unknown. Our work is mainly focused on the last step of spermatogenesis, called spermiogenesis. When abnormal, it leads to male infertility.

    Spermiogenesis is a fascinating process in term of gene expression dynamics and chromatin remodeling. During this step, postmeiotic germ cells known as spermatids undergo profound morphological and functional changes to become spermatozoa. They acquire their specific shape through the loss of most of their cytoplasm, the biogenesis of a flagellum and an acrosome, and the compaction of their nucleus to ~1/5th of its original size. Many of the morphological and functional changes occurring during sperm differentiation are orchestrated by the spermatid (epi)genetic program, with thousands of genes highly expressed in spermatids, before transcription shuts down due to chromatin compaction. A significant proportion of spermatid-specific genes is located on the X and Y chromosomes and appears to be co-regulated.

    We study the postmeiotic differentiation of male germ cells (i.e. spermiogenesis) at the gene and the chromatin level using omics, in vivo and in vitro approaches, to identify and characterize novel regulators/pathways required for sperm differentiation and male fertility.

    Our project also aims at studying the impact of spermatozoa epigenetic program on reproductive efficiency, embryo development and progeny’s health. Indeed it is now well-known that, upon fertilization, the sperm cell contributes to the embryo with more than its DNA. Epigenetic information (chromatin, RNA molecules and/or DNA modifications) is also transmitted to the embryo and could affect embryo development and offspring health in some cases of abnormal spermiogenesis. It is therefore essential to better understand the (epi)genetic processes occurring during sperm differentiation.

     

     

    Participants

    Julie Cocquet, Principal investigator (CR1 INSERM)

    Côme Ialy-Radio, ITA (AI INSERM)

    Melina Blanco (M2 student)

    Laila El Khattabi (AHU, Cochin Hospital staff)

    Clara Gobé (chercheur non statutaire)




    Scientific questions

    Deletions of the long arm of the mouse Y chromosome (MSYq) lead to a wide range of sperm differentiation defects, such as deformed spermheads, reduced motility, abnormal chromatin compaction and DNA damage, which cause male infertility. Those defects are associated with loss of specific epigenetic marks (such as H3K9me3, i.e. histone H3 lysine 9 trimethylation) and subsequent deregulation of hundreds of sex chromosome-encoded genes, as well as a smaller portion of autosomal genes (such as the genes of the Speer family). We have previously shown that these phenotypes result from deficiency in the MSYq-encoded multicopy gene Sly (Cocquet et al. PLos Biol 2009; Riel et al. J Cell Sci 2013). Interestingly, we have observed that Slx, the X-linked homolog of Sly (Cocquet et al. 2010), has a role opposite to that of Sly on gene expression and associated chromatin marks (Cocquet et al. PLoS Genet 2012). Slx deficiency also leads to sperm differentiation defects characterized by abnormal sperm morphology and increased spermatid apoptosis (Cocquet et al. Mol Biol Cell 2010). Both Slx and Sly are involved in an intragenomic conflict that causes segregation distortion and male infertility. This conflict has influenced the structure of the mouse genome and may have played an important role in mouse speciation (Cocquet et al. PLoS Genet 2012).

    While studying SLY and SLX proteins, we have identified one of their protein partners, SSTY, which is encoded by MSYq region. SSTY is a member of the SPIN/SSTY protein family. It is specifically expressed in postmeiotic male germ cells and could also contribute to gene and chromatin regulation during sperm differentiation (Comptour et al. FEBS J 2014).

    Our aim is to better understand gene and chromatin regulation during postmeiotic sperm differentiation using SLX, SLY and SSTY as entry points. We have recently shown by ChIP Seq that SLY protein is enriched at the promoter of thousands of genes involved in gene expression, chromatin regulation, and the ubiquitin pathway, with the same feature, all those genes are highly expressed in postmeiotic male germ cells. Among those genes is Dot1l, which encodes the only known H3K79 methyltransferase. H3K79 methylation is expected to be important for the chromatin remodeling and compaction process occurring in spermatids which results from the replacement of most histones by protamines. Our data show that Sly deficiency leads to a downregulation of Dot1l and H3K79 methylation in spermatids, associated with abnormal chromatin remodeling. As a result, Sly-deficient spermatozoa are abnormally shaped, less compact and present a higher susceptibility to DNA damage than WT spermatozoa (Moretti et al. Cell Death & Diff 2017).

    In parallel, we have used a global approach (co-immunoprecipitation followed by mass spectrometry) to identify novel SLY protein partners, and found that SLY interacts with SMRT/NcoR, a protein complex involved in transcriptional regulation. (Moretti et al. Cell Death & Diff 2017).Our results provide a model to explain the abnormal gene regulation and chromatin remodeling observed in Sly-deficient spermatids.

     

     

     

    Localization of SLX and SLY proteins in spermatids. SLX and SLY proteins (in green, respectively top and bottom panel) co-localize with the X or Y chromosome (in yellow, X or Y paint) and with Speer gene cluster (in red, DNA FISH). DAPI (blue) was used to stain nuclei. Image Credit: Cocquet et al. PLoS Genetics 2012

     

     

    Figure: From Moretti et al. 2017. Model presenting the mechanism by which SLY controls gene expression and chromatin remodeling during sperm differentiation. In WT round spermatids (left panel), SLY (in blue) interacts with the SMRT/N-CoR complex (which comprises TBL1XR1, TBL1X, NCOR1 and HDAC3) and is located at the start of genes involved in gene regulation, chromatin regulation and the ubiquitin pathway. In particular, SLY directly controls the expression of X-chromosome-encoded genes coding for H2.A variants (such as H2A.B3) and of the H3K79 methyltransferase DOT1L. In elongating spermatids, there is a wave of H3K79 dimethylation (orange circles) and of histone H4 acetylation (green circles); those modifications are expected to be a prerequisite to the efficient removal of nucleosomes (light pink oval) and replacement by protamines (purple oval), a process which is required to achieve optimal compaction of the spermatozoa nucleus. When SLY is knocked down (right panel), X-encoded H2.A variants are upregulated and more incorporated in the spermatid chromatin, while DOT1L is downregulated. DOT1L downregulation leads to a decrease in dimethylated H3K79 and acetylated histone H4 in elongating spermatids. Alterations in the spermatid chromatin structure affect the replacement of nucleosomes by protamines and lead to a higher proportion of nucleosomes and a decreased proportion of protamines. As a result, Sly-deficient spermatozoa are abnormally shaped, less compact and present a higher susceptibility to DNA damage than WT spermatozoa

     

    Selected publications

    Blanco M, Cocquet J. Genetic Factors Affecting Sperm Chromatin Structure. Adv Exp Med Biol. 2019;1166:1-28.

    Riel JM, Yamauchi Y, Ruthig VA, Malinta QU, Blanco M, Moretti C, Cocquet J, Ward MA. Rescue of Sly Expression Is Not Sufficient to Rescue Spermiogenic Phenotype of Mice with Deletions of Y Chromosome Long Arm. Genes (Basel). 2019 Feb 12;10(2).

    Champroux A, Cocquet J, Henry-Berger J, Drevet JR, Kocer A. A Decade of Exploring the Mammalian Sperm Epigenome: Paternal Epigenetic and Transgenerational Inheritance. Front Cell Dev Biol. 2018 May 15;6:50.

    El Kennani S, Adrait A, Permiakova O, Hesse AM, Ialy-Radio C, Ferro M, Brun V, Cocquet J, Govin J, Pflieger D.(2018) Systematic quantitative analysis of H2A and H2B variants by targeted proteomics. Epigenetics Chromatin. 2018 Jan 12;11(1):2.

    Moretti C*, Serrentino ME*, Ialy-Radio C, Delessard M, Soboleva TA, Tores F, Leduc M, Nitschké P, Drevet JR, Tremethick DJ, Vaiman D, Kocer A, Cocquet J. SLY regulates genes involved in chromatin remodeling and interacts with TBL1XR1 during sperm differentiation. Cell Death Differ. 2017 24:1029-1044

    Moretti C, Vaiman D, Tores F, Cocquet J. Expression and epigenomic landscape of the sex chromosomes in mouse post-meiotic male germ cells. Epigenetics Chromatin. 2016 Oct 27;9:47. eCollection 2016

    Comptour A*, Moretti C*, Serrentino ME, Auer J, Ialy-Radio C, Ward MA, Touré A, Vaiman D, Cocquet J. SSTY proteins co-localize with the post-meiotic sex chromatin and interact with regulators of its expression. FEBS J. 2014 281: 1571-84

    Riel JM, Yamauchi Y, Sugawara A, Li HY, Ruthig V, Stoytcheva Z, Ellis PJ, Cocquet J, Ward MA. (2013) Deficiency of the multi-copy mouse Y gene Sly causes sperm DNA damage and abnormal chromatin packaging. J Cell Sci 126: 803-813

    Cocquet J, Ellis PJ, Mahadevaiah SK, Affara NA, Vaiman D, Burgoyne P. (2012) A genetic basis for a postmeiotic X versus Y chromosome intragenomic conflict in the mouse. PLoS Genet 8: e1002900.

    Cocquet J, Ellis PJ, Yamauchi Y, Riel JM, Karacs TP, Rattigan A, Ojarikre OA, Affara NA, Ward MA, Burgoyne PS. (2010) Deficiency in the multicopy Sycp3-like X-linked genes Slx and Slxl1 causes major defects in spermatid differentiation. Mol Biol Cell. 21:3497-505.

    Cocquet J, Ellis PJ, Yamauchi Y, Mahadevaiah SK, Affara NA, Ward MA, Burgoyne P. (2009) The multicopy gene Sly represses the sex chromosomes in the male mouse germline after meiosis. PLoS Biol 7: e1000244.

     

    Funding