IHEM   20887
INSTITUTO DE HISTOLOGIA Y EMBRIOLOGIA DE MENDOZA DR. MARIO H. BURGOS
Unidad Ejecutora - UE
congresos y reuniones científicas
Título:
Phosphorylation /dephosphorylation of synaptotagmin 6 during acrosomal exocytosis.
Autor/es:
CASTILLO BENNETT J, ROGGERO CM, MANCIFESTA F, MAYORGA LS.
Lugar:
Carlos Paz Argentina
Reunión:
Congreso; XLIV Reunión Anual de la Sociedad Argentina de Investigación en Bioquímica y Biología Molecular.; 2008
Institución organizadora:
Sociedad Argentina de Investigación en Bioquímica y Biología Molecular
Resumen:
The acrosomal exocytosis (AE) is a calcium regulated exocytosis essential for fertilization. We have demonstrated that synaptotagmin 6 (Syt6) is phosphorylated in resting sperm, and that its C2B domain presents a polybasic region which is a target for PKC. As we predected that threonine 419 (T419) has more probability to be phosphorylated by this kinase, we created a phosphomimetic mutant (T419E). Using a FRET assay we demostrated that T419E lost the ability to bind liposomes in a Ca2+- dependent way and its inhibitory effect in an AE assay. These results suggest that the PKC mediated regulation of C2Bwt activity depends on T419 phosphorylation. However, T419 is not the only target for PKC; the T419E mutant was still able to incorporate 32P in an in vitro phosphorylation assay. We previously demonstrated that Syt 6 is dephosphorylatedafter sperm stimulation. Calcineurin (CaN), a calcium/calmodulin-activated protein phosphatase that is present in sperm and is required for AE, could be responsible of Syt 6 dephosphorylation. In an in vitro phosphorylation/dephosphorylation assay with 32P, using a constitutive active form of caN, we show rhat the Syt 6 C2B domain was dephosphorylated by CaN. We conclude that Syt 6 activity is regulated by PKC phosphorylation at T419 and needs to be dephosphorytlated by CaN to participate in AE.