A Rab-GEF cascade during human sperm exocytosis.

BUSTOS MA; LUCCHESI O; RUETE MC; MAYORGA LS; TOMES CN

Mendoza

Congreso; XLVIII Reunión Anual Sociedad Argentina de Investigaciones en Bioquímica y Biología Molecular (SAIB); 2012

Sociedad Argentina de Investigaciones en Bioquímica y Biología Molecular (SAIB)

Rab3 and Rab27 are the two major ?secretory Rabs? implicated in calcium-regulated exocytosis in a variety of cells. Understanding how their actions are coordinated is a challenge in the field. Sperm contain a single, large dense-core granule that is released by regulated exocytosis (termed the acrosome reaction, AR) during fertilization. The acrosomal contents are released using the same conserved fusion machinery utilized by neurons and neuroendocrine cells.  Two Rab proteins, Rab3 and Rab27, are required for sperm exocytosis. The levels of GTP-bound Rab27 and Rab3 increase upon initiation of exocytosis. With a fluorescence microscopy-based method developed in the laboratory, we found that challenging with an AR inducer increases the population of cells exhibiting GTP-bound Rabs in the acrosomal region of human sperm. Interestingly, introducing recombinant Rab27A loaded with GTP-γ-S into streptolysin-O permeabilized sperm elicits a remarkable increase in the number of cells evincing GTP-bound Rab3A. In the converse condition, recombinant Rab3A does not modify the percentage of Rab27-GTP-containing cells. Furthermore, Rab27A-GTP recruits a Rab3 GDP/GTP exchange factor (GEF) activity from human sperm extracts. Our findings suggest that Rab27/Rab3A constitute a Rab-GEF cascade, the first ever described in dense-core granule exocytosis.