The first part of my thesis project focused on the study of genes involved in spermiogenesis. Through the analysis of an interspecific recombinant congenic mouse strain (IRCS), we previously identified a quantitative trait locus (QTL) in mouse, named Mafq1 on chromosome 1, which is associated with male hypofertility due to ultrastructural sperm abnormalities. Within this locus, we identified the Spata3 (Spermatogenesis Associated 3) gene as a relevant candidate. We invalidated it and observed that Spata3-KO male were fertile in vivo but showed significant hypofertility in vitro accompanied by ultrastructural defects in the acrosome and persistence of cytoplasmic residual bodies in the flagellum. Since the invalidation of Spata3 only partially reproduced the phenotype of the hypofertile IRCS strain, we reanalyzed the latest versions of the mouse genome, and identified at the Mafq1 locus a new candidate gene specifically expressed in post-meiotic male germ cells: Tex44 (Testis-expressed protein 44), that we also invalidated. Tex44-KO males were severely hypofertile in vivo and in vitro. Although the phenotype of the hypofertile IRCS strain was again only partially reproduced, we showed that the absence of the Tex44 gene results in the severe malformation and dysfunction of the flagellum (disjunction between the midpiece and the principal piece leading to a 180° folding in this area, loss of some axonemal microtubule doublets and outer dense fibers in the principal piece, reduced mitochondrial activity), severely impacting sperm motility and hence fertility in male mice.

Sperm and oocyte fusion is essential for fertilization in mammals. The interaction between the oocyte JUNO and the sperm IZUMO1 proteins is followed by the dimerization of the latter, which then changes its structure, leading to a loss of affinity between this ligand-receptor pair essential for gamete adhesion. This event appears to be important for the continuation of the gamete interaction process and could be controlled by a protein with disulfide isomerase (PDI) activity: ERp57. The aim of the second part of my thesis project is to elucidate the role of this protein in gamete interaction, in vitro in mouse and in human, then in vivo in a mouse model in which males are invalidated for the ERp57 gene specifically in sperm (conditional knockout: scKO). We performed in vitro fertilization (IVF) experiments in the presence of either inhibitors of the PDI family that are more or less specific for ERp57, in mouse, or antibodies specifically directed against ERp57 protein in mouse and in human. A significant reduction of fertilization was observed in the presence of the latter. In mouse and human, we observed after acrosome reaction a relocation of the ERp57 protein, similar to that of IZUMO1, from the acrosome to the equatorial segment, which is the initial site of gamete fusion. Finally, our experiments on scKO males showed a severe hypofertility in vivo and in vitro. Taken together, the data obtained demonstrate that, in both mouse and human, ERp57 as a sperm protein appears to play an important role during fertilization.