Molecular identification and localization analysis of SNAPs and NSF during mouse oocyte maturation

DE PAOLA MARIA MATILDE; GALLO GIOVANNA LUCRECIA; MAYORGA LUIS SEGUNDO; MICHAUT MARCELA ALEJANDRA

Mendoza

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

Sociedad Argentina de Investigación en Bioquímica y Biología Molecular

In mammals, the primary mechanism in preventing polyspermicfertilization involves cortical granules exocytosis (CGE). Severalstudies have suggested that CGE is a SNARE protein-mediatedpathway; however, the molecular characterization of CGE is stillincomplete. We hypothesized that the oocyte uses the sameconserved membrane fusion machinery as neurons and humansperm, and the regulatory complex SNAP/NSF is present in mouseoocyte.We first investigated the expression of SNAP isoforms: alfa, beta and gamma and NSF by RT-PCR. The results show that the alfa and gammaSNAP (but not beta), and NSF are expressed in mouse oocyte. Westernblot analyses indicate that these proteins are present during oocytematuration and egg activation showing no variations between thedifferent stages. Indirect immunofluorescence experimentsrevealed that alfa SNAP and NSF localized mainly in the corticalregion of all stages analyzed. While gamma SNAP had a similardistribution that alfa SNAP, this protein also showed a cytoplasmicdistribution in immature oocytes. All identified proteins: alfa SNAP,gamma SNAP, and NSF are predominately observed in the corticalregion, 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 alfa SNAP and gamma SNAPare equally important in CGE or have different functions in mouseoocyte.