ANALYSIS OF SNAP/NSF COMPLEX IN THE MOUSE OOCYTE PHYSIOLOGY

DE PAOLA, M. MATILDE; ARCOS, ALEXIS; BELLO, OSCAR D.; CAPPA, ANDREA I.; RINALDINI, E; MAYORGA, LUIS S.; BÁTIZ, LUIS FEDERICO; MICHAUT, MARCELA ALEJANDRA

Puerto Varas

Congreso; XII Meeting of Pan American Biochemistry & Molecular Biology Societies; XLIX Reunión Anual de la Sociedad Argentina de Investigación en Bioquímica y Biología Molecular; 2013

Following fertilization, oocyte cortical granules (CG) undergo exocytosis (CGE) to release their content into the perivitelline space, avoiding polyspermy and ensuring normal embryonic development. Several studies suggest that CGE is a SNARE-mediated process; however, the molecular mechanism is still incomplete. We hypothesized that SNAP/NSF complex regulates CGE. RT-PCR results showed that alpha- and gamma-SNAP (but not beta), and NSF are expressed in mouse oocyte. Western blot analyses indicate that these proteins are present during oocyte maturation and egg activation. Indirect immunofluorescence reveals that all identified proteins are predominantly observed in the CG-enriched cortical region. Furthermore, we showed that microinjection of specific antibodies against either alpha-SNAP or NSF inhibits SrCl2-stimulated CGE in a dose-dependent manner. To further confirm these results in vivo, we started the characterization of a spontaneous alpha-SNAP mutant mouse known as hyh. So far, we have determined that hyh female have a strongly reduced in vivo fertility and, under hormonal ovary stimulation, hyh female mice yield four times less oocytes than wild type mice. In addition, mature oocytes from hyh mice exhibit a reduced density and an atypical distribution of CG. Altogether, our results strongly suggest that alpha-SNAP/NSF complex regulates CGE, and that a failure of SNAPs and/or NSF might have consequences still not uncovered.