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Biology of Reproduction 62, 1084-1089 (2000)
© 2000 Society for the Study of Reproduction, Inc.


Articles

Rab3A Triggers the Acrosome Reaction in Permeabilized Human Spermatozoa1

R. Yunes2,a, M. Michauta, C. Tomesa, and L.S. Mayorgaa

a Laboratorio de Biología Celular y Molecular, Instituto de Histología y Embriología (IHEM-CONICET), Facultad de Ciencias Médicas, Universidad Nacional de Cuyo, 5500 Mendoza, Argentina

The acrosome reaction is a regulated exocytotic process leading to a massive fusion between the outer acrosomal membrane and the cell membrane. In spite of the great amount of information available related to the acrosome reaction in several species, there is a remarkable paucity about the role of monomeric guanosine triphosphatases (GTPases) of the Rab family—well-established participants in exocytosis in other cell types—in the acrosome reaction. Western blot and immunofluorescence analysis indicate that Rab3A is present in human spermatozoa and localizes to the acrosomal region in the sperm head. One difficulty in studying the role of proteins in intact cells is the fact that they are unable to cross the cell membrane. Therefore, we established a working model of streptolysin O-permeabilized human spermatozoa. Permeabilized spermatozoa were able to respond in a regulated way to different stimuli, such as G protein activators and calcium. An acrosomal reaction was also triggered by a Rab3A peptide corresponding to the effector region. More important, recombinant Rab3A protein in the GTP-bound form caused acrosome exocytosis. The same protein loaded with GDP or Rab11 in the GTP-bound form was inactive. Also, recombinant GDI (GDP dissociation inhibitor)—a protein that releases Rab proteins from membrane—inhibited a GTP{gamma}S-stimulated acrosome reaction. Our results indicate that 1) permeabilized spermatozoa can be used to study the role of macromolecules in the acrosome reaction, 2) Rab3A is present in human spermatozoa, and 3) Rab3A or another Rab3 isoform is involved in the exocytosis of the acrosomal granule in human spermatozoa.

First decision: 6 October 1999.

1 This work was partially supported by an International Research Scholar Award from the Howard Hughes Medical Institute, CONICET PIP (4276/96) and PEI (0423/97), and CIUNC.

2 Correspondence: Roberto Yunes, Laboratorio de Biología Celular y Molecular, Instituto de Histología y Embriología (IHEM-CONICET), CC 56, Facultad de Ciencias Médicas, Universidad Nacional de Cuyo, 5500 Mendoza, Argentina. FAX: 54 261 4494117; ryunes{at}fmed2.uncu.edu.ar




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