I-13 Infertility with Impaired Zona Pellucida Adhesion of Spermatozoa from Mice LackingTauCstF-64

Background: Fertilization is a multistep process requiring spermatozoa with unique cellular structures and numerous germ cell-specific molecules that function in the various steps. In the highly coordinated process of male germ cell development, RNA splicing and polyadenylation help regulate gene expression to ensure formation of functional spermatozoa. Male germ cells express tauCstF-64 (Cstf2t gene product), a paralog of the Xlinked CstF-64 protein that supports polyadenylation in most somatic cells. We previously showed (PNAS 2007) that loss of tauCstF-64 causes male infertility because of major defects in mouse spermatogenesis. This infertility mouse model was identified as oligoasthenoteratozoospermia, an infertility condition diagnosed in the human population. Materials and Methods: This unique mouse model was produced by targeted disruption of Cstf 2t gene that this encoding protein is related to polyadenylation in germ cells. This present studies used procedures like sperm immunoflurescence, in vitro fertilization and electron microscopy to determine the cause and the severity of this infertility. Results: Surprisingly, although Cstf2t-/- males produce very few recognizable spermatozoa, some of the spermatozoa produced were motile. This led us to ask whether these Cstf2t-/- sperm were fertile. A motile cell-enriched population of spermatozoa from Cstf2tnull males dispersed cumulus cells of cumulus-oocyte complexes normally. However, motile spermatozoa from Cstf2t-null males failed to fertilize cumulus-intact mouse eggs in vitro. In addition, sperm adhesion to the zona pellucida (ZP) of cumulus-free eggs was significantly decreased, indicating tauCstF-64 is required for production of spermatozoa capable of ZP interaction. Acrosomal proteins involved in sperm-ZP recognition, including zonadhesin, proacrosin, SPAM1/PH-20, and ZP3R/sp56, were normally distributed in the apical head of Cstf2t-/- spermatozoa. Conclusion: We conclude that tauCstF-64 is required not only for expression of genes involved in morphological differentiation of spermatids, but also for genes having products that function during interaction of motile spermatozoa with eggs. To our knowledge, this is the first demonstration that a gene involved in polyadenylation has a negative consequence on sperm-ZP adhesion and fertility