Sequence of Events Leading to Membrane Fusion during Acrosomal Exocytosis in Human Spermatozoa.

DE BLAS, GERARDO A.; ROGGERO, CARLOS M.; BELMONTE, S.A.; TOMES, CLAUDIA N.; MAYORGA, LUIS S.

Merida, Mexico

Otro; Meeting of Internacional Research Scholars. (HHMI 2005); 2005

The acrosome is a membrane-limited granule that overlies che nucleus of the mature spermatozoon. In response to stimuli, it undergoes a special type of calcium-dependent exocytosis termed the acrosorne reacrion, which is an absolute prerequisite for fertilization. SNARE (SNAP receptor) complexes play a central role in mosc membrane fusion events. Acrosnmal exocytosis has several unique features that allow for a detailed characterization of the dynamics (and its regulation) of assembly disassembly of SNARE complexes during membrane Fusion. The release of the acrosomal content is an all-or-nothing event that occurs as wave, requires neurotoxin-sensitive SNAREs. furlctional assay and immunofluorescence techniques in combination with neurotoxins and a photosensitive Ca2+ chelator, we have shown that SNAREs are associatd in c ic~o mplexes in resting sperm. Upon Ca2+ entry into the cytoplasm, Rab3A is activated and anchored to the membranes in a process that is rnodulaced by cholesterol. Interestingly, this sterol does not affect late steps of the membrane fusion process. Active Rab3A triggers the disassembly of cis SNARE complexes by means of N-ethlymaleimide-sensitive factor (NSF) and alphaSNAP (soluble NSF attachment protein). SNAREs are then free to reassemble in loose m s complexes that are resistant to NSF/alphaSNAP action. Ac this point Ca2+ must be released from inside the acrosome to initiate the find steps of membrane fusion, which require fully assembled trans SNARE complexes and synaptotagmin VI. l´he function of this Ca2+-sensing protein is regulated by PKC-mediated pho~phor~latioonf a conserved polybasic domain rlecmsary for interaction with multiple effectors. Synaptotagmin desphosphnrylation is probably an early event during exo- ~ytosisa, llowing the interaction of this protein with other components of the fusion machinery. Our results indicate that rhe unidirectional and sequcntial disas~ernhyl and assembly of SNARE complexes drives membrane fusion during the acrosomal exocytosis of human spermatozoa.