Characterization of VAMP isoforms involved in cortical granule exocytosis in mouse oocyte.

CAPPA, ANDREA ISABEL; DE PAOLA, MARIA MATILDE; GARRIDO, FACUNDO; MICHAUT, MARCELA ALEJANDRA

Córdoba

Congreso; LII Reunión Anual de la Sociedad Argentina de Investigación en Bioquímica y Biología Molecular (SAIB).; 2016

Sociedad Argentina de Investigación en Bioquímica y Biología Molecular

Cortical granule exocytosis (CGE) is a secretory process triggered by sperm-oocyte fusion during fertilization. This exocytosis involves the membrane fusion of cortical granule, localized in the oocyte cortex, and oolema. This fusion is regulated by intracellular calcium and is thought to be mediated by the SNARE complex, which is formed by SNAP-25, Syntaxin and VAMP. Syntaxin4 and SNAP-25 have been identified in mouse egg; however it is unknown if VAMP (vesicle-associated membrane protein) participates in CGE. The aim of this work was to identify and characterize the main VAMP isoforms, VAMP1 and VAMP2, involved in exocytosis in other secretory models. RT-PCR assays showed that VAMP1 and VAMP2 are expressed in mouse oocytes. Immunofluorescence analysis showed that VAMP1 had a cortical localization, while VAMP2 had mainly a cytoplasmic distribution. This result suggested that VAMP1 might be involved in CGE. To confirm this hypothesis we performed a functional assay perturbing the function of endogenous VAMP1 and VAMP2 by antibody microinjection. Only VAMP1 antibody was able to inhibit CGE when mouse oocytes were activated parthenogenetically with strontium chloride. These results confirm that VAMP1 is the VAMP isoform involved in CGE. The importance of this work is that we have completed the characterization of SNARE complex during membrane fusion in the CGE of mouse oocytes.