MOLECULAR IDENTIFICATION AND LOCALIZATION ANALYSIS OF SNAPS AND NSF DURING MOUSE OOCYTE MATURATION

DE PAOLA, M. MATILDE; GALLO, GIOVANNA L.; MAYORGA, LUIS S.; MICHAUT, MARCELA A.

Mendoza

Congreso; XLVIII Reunión Anual de la Sociedad Argentina de Investigación en Bioquímica y Biología Molecular; 2012

In mammals, the primary mechanism in preventing polyspermic fertilization involves cortical granules exocytosis (CGE). Several studies have suggested that CGE is a SNARE protein-mediated pathway; however, the molecular characterization of CGE is still incomplete. We hypothesized that the oocyte uses the same conserved membrane fusion machinery as neurons and human sperm, and the regulatory complex SNAP/NSF is present in mouse oocyte. We first investigated the expression of SNAP isoforms: alpha, beta, and gamma, and NSF by RT-PCR. The results show that the 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 showing no variations between the different stages. Indirect immunofluorescence experiments revealed that alpha-SNAP and NSF localized mainly in the cortical region of all stages analyzed. While gamma-SNAP had a similar distribution that alpha-SNAP, this protein also showed a cytoplasmic distribution in immature oocytes. All identified proteins:  alpha-SNAP, gamma-SNAP, and NSF are predominately observed in the cortical region, which is enriched in cortical granules at the mature oocyte, suggesting that these proteins may be involved in CGE. However, more studies are needed to elucidate if both alpha-SNAP and gamma-SNAP are equally important in CGE or have different functions in mouse oocyte.