PERMEANTS PROTEINS AS A TOOL TO STUDY CORTICAL GRANULES EXOCYTOSIS IN MOUSE OOCYTES

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

Mendoza

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

One of the most important events that are produced in oocytes as a result of fertilization is cortical granule exocytosis (CGE), a secretory process that produces polispermy blockade and ensures normal embryonic development. Nevertheless, the molecular mechanism of this process remains unknown. Previous results in our laboratory show that MARCKS, a prominent substrate of PKC involved in exocytosis in different cell types, participates in CGE in mouse oocyte. In this cellular model, proteins are introduced by microinjection, which is a very valuable and useful technique, but it causes an injury to the cells which is reflected in high mortality and low yields. We decided to tune up the usefulness of permeant proteins to study exocytotic mechanisms in mouse oocyte. We used the permeant peptide corresponding to MARCKS effector domain (amino acids 154-165) conjugated to thrimethylrhodamine (TMR-ED). Oocytes were incubated with increasing concentrations of this peptide, previous remotion of zona pellucida; then cells were parthenogenetically activated with SrCl2, and cortical granules were quantified with the program Image J. TMR-DE was able to inhibit CGE and this effect was concentration-dependent. These results corroborate previous data obtained in our laboratory by protein microinjection, and show the efficiency of permeant proteins as a tool to study CGE in moouse oocytes.