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The Journal of Cell Biology, Vol 135, 1741-1747, Copyright © 1996 by The Rockefeller University Press
ARTICLES |
N Masumoto, T Sasaki, M Tahara, A Mammoto, Y Ikebuchi, K Tasaka, M Tokunaga, Y Takai and A Miyake
Department of Obstetrics and Gynecology, Osaka University Medical School, Japan. masumoto@gyne.med.osaka-u.ac.jp
Rabphilin-3A is a putative target protein for Rab3A, a member of the small GTP-binding protein superfamily that has been suggested to play a role in regulated exocytosis in presynapses. In this study we determined the expression and the function of Rabphilin-3A in mouse eggs at fertilization. Rabphilin-3A mRNA and protein were detected by reverse transcriptase-PCR and immunoblot analysis, respectively, in metaphase II mouse eggs. Immunofluorescence analysis showed that Rabphilin-3A protein was distributed in the cortical region in eggs. Sperm induces cortical granule (CG) exocytosis via an increase in cytosolic Ca2+ at fertilization. We microinjected the NH2- or COOH- terminal fragment of recombinant Rabphilin-3A into metaphase II eggs. Neither treatments altered the sperm-induced cytosolic Ca2+ increase, but both inhibited CG exocytosis in a dose-dependent manner. The NH2- terminal fragment was more effective than the COOH-terminal fragment. Full-length Rabphilin-3A did not affect CG exocytosis, but it attenuated the inhibition of CG exocytosis by the NH2-terminal fragment. These results show that Rabphilin-3A is involved in Ca(2+)- dependent CG exocytosis at fertilization in mouse eggs.
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