IHEM   20887
INSTITUTO DE HISTOLOGIA Y EMBRIOLOGIA DE MENDOZA DR. MARIO H. BURGOS
Unidad Ejecutora - UE
congresos y reuniones científicas
Título:
THE SIGNALING MODULE RAB27-RABPHILIN-3AGRAB- RAB3 IN SPERM EXOCYTOSIS
Autor/es:
BUSTOS MA; LUCCHESI O; QUEVEDO MF; TOMES CN
Lugar:
Buenos Aires
Reunión:
Congreso; REUNIÓN CONJUNTA DE SOCIEDADES DE BIOCIENCIAS; 2017
Institución organizadora:
10 Sociedades
Resumen:
Exocytosis is a fundamental cellular process used by eukaryotic cells to release biological compounds such as peptide hormones and neurotransmitters and to insert specific lipids and proteins in the plasma membrane. Mammalian sperm exocytose their acrosome (the acrosome reaction or AR) at fertilization. The AR depends on members of the standard fusion machinery, including small GTPases and SNAREs; two such GTPases, Rabs27 and 3, are organized in a Rab-GEF cascade. Western blot and indirect immunofluorescence were used to detect previously undescribed proteins and their localization in sperm. Antibodies and a variety of recombinant proteins were applied in functional assays in intact and streptolysin O-permeabilized sperm to investigate the role of members of the fusion cascade in the AR. The gene encoding Rab27A was subcloned downstream the TAT sequence of the HIV virus to express permeable Rab27A in Escherichia coli. Immobilized Rabs loaded with GDP or GTP and Rabphilin-3a were used to pull down interacting proteins from human sperm extracts. Pull down was also the method of choice to determine Rab3-GEF activity. Here we show that a permeable, recombinant version of human Rab27A elicits the AR when introduced into capacitated human sperm. Rab27A promotes the exchange of GDP for GTP on Rab3 in the acrosomal region of intact sperm, so does the catalytic domain of GRAB, a GEF for Rab3, when introduced into permeabilized sperm. Rab27 accomplishes these functions by recruiting its effector Rabphilin-3a, which, in turn, interacts with GRAB.