The intraacrosomal calcium pool plays a direct role in acrosomal exocytosis.
The acrosome reaction is a unique type of regulated exocytosis. The single secretory granule of the sperm fuses at multiple points with the overlying plasma membrane. In the past few years we have characterized several aspects of this process using streptolysin O-permeabilized human spermatozoa. Here we show that Rab3A triggers acrosomal exocytosis in the virtual absence of calcium in the cytosolic compartment. Interestingly, exocytosis is blocked when calcium is depleted from intracellular stores. By using a membrane-permeant fluorescent calcium probe, we observed that the acrosome actually behaves as a calcium store. Depleting calcium from this compartment by using a light-sensitive chelator prevents secretion promoted by Rab3A. UV inactivation of the chelator restores exocytosis. Rab3A-triggered exocytosis is blocked by calcium pump and inositol 1,4,5-trisphosphate (IP(3))-sensitive calcium channel inhibitors. Calcium measurements inside and outside the acrosome showed that Rab3A promotes a calcium efflux from the granule. Interestingly, release of calcium through IP(3)-sensitive calcium channels was necessary even when exocytosis was initiated by increasing free calcium in the extraacrosomal compartment in both permeabilized and intact spermatozoa. Our results show that a calcium efflux from the acrosome through IP(3)-sensitive channels is necessary downstream Rab3A activation during the membrane fusion process leading to acrosomal exocytosis.
De Blas
Gerardo
G
Laboratorio de Biologia Celular y Molecular, Instituto de Histologia y Embriologia (IHEM-CONICET), Facultad de Ciencias Médicas, CC 56, Universidad Nacional de Cuyo, Casilla de Correo 56, 5500 Mendoza, Argentina.
Michaut
Marcela
M
Treviño
Claudia L
CL
Tomes
Claudia N
CN
Yunes
Roberto
R
Darszon
Alberto
A
Mayorga
Luis S
LS
D000177:Acrosome / Q000378:metabolism*; D020101:Acrosome Reaction; D002118:Calcium / Q000378:metabolism*; D003600:Cytosol / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004533:Egtazic Acid / Q000031:analogs & derivatives* / Q000494:pharmacology; D005089:Exocytosis; D006801:Humans; D007211:Indoles / Q000494:pharmacology; D015544:Inositol 1,4,5-Trisphosphate / Q000378:metabolism; D008024:Ligands; D008027:Light; D008297:Male; D011994:Recombinant Proteins / Q000378:metabolism; D013050:Spectrometry, Fluorescence; D013094:Spermatozoa / Q000378:metabolism; D019284:Thapsigargin / Q000494:pharmacology; D013997:Time Factors; D014466:Ultraviolet Rays; D020826:rab3A GTP-Binding Protein / Q000378:metabolism
D007211:Indoles; D008024:Ligands; D011994:Recombinant Proteins; C070379:1,2-bis(2-aminophenoxy)ethane N,N,N',N'-tetraacetic acid acetoxymethyl ester; D004533:Egtazic Acid; D019284:Thapsigargin; D015544:Inositol 1,4,5-Trisphosphate; D020826:rab3A GTP-Binding Protein; D002118:Calcium; C000543:cyclopiazonic acid