Sequence of events leading to membrane fusion during acrosomal exocytosis

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

Villa Carlos Paz, Cordoba., Argentina

Workshop; Internetional Workshop on Membrane Trafficking.; 2005

The acrosome is a membrane-limited granule which overlies the nucleus of the maturespermatozoon. In response to stimuli, it undergoes a special type of calcium-dependentexocytosis termed the acrosome reaction, which is an absolute prerequisite forfertilization. SNARE complexes play a central role in most membrane fusion events.Acrosomal exocytosis has several unique features that allow for a detailedcharacterization of the dynamics of assembly and disassembly of SNARE complexesand its regulation during membrane fusion. It is an all-or-nothing event that occurs as asynchronized wave, with no recycling of co~nponents and requires neurotoxin-sensitiveSNAREs. By using a functional assay and immunofluorescence techniques incombination with neurotoxins and a photosensitive ca2' chelator, we have shown that inresting sperm, SNAREs are associated in cis complexes. Upon ca2' entrance into thecytoplasm, Rab3A is activated and triggers the disassembly of cis SNARE co~nplcxesby means of NSF and DSNAP. SNAREs are now free to re-assemble in loose trans complexes that are resistant to NSF/OSNAP action. At this point, ca2' must be released from inside the acrosome to initiate the final steps of membrane fusion thatrequire full T+ assembled trans SNARE complexes and synaptotagmin VI. The function of this Ca -sensoring protein is regulated by PKC-mediated phospI~orylation of a conserved polybasic domain necessary for interaction with multiple erectors.Synaptotagmin dephosphorylation is probably an early event during exocytosis thatallows the interaction of this protein with other components of the fusion machinery.Our results indicate that the unidirectional and sequential disassembIy and assembly ofSNARE complexes drives membrane fusion during the acrosornal exocytosis of humanspermatozoa.