Signal transduction during shrimp oocyte activation by extracellular Mg2+: roles of inositol 1,4,5-trisphosphate, tyrosine kinases and G-proteins

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ABSTRACT Oocytes of the shrimp Sicyoniaingentis are naturally activated upon contact with seawater Mg2+. We investigated the mechanism of Mg2+-induced intracellular Ca2+ release and cortical contraction through treatment of oocytes with various activators and inhibitors of signal transduction pathways. Injection of oocytes with the second messenger inositol 1,4,5-trisphosphate resulted in an immediate rise in intracellular Ca2+ and normal cortical contraction. By contrast, injection of the GTP analog guanosine 5′-O-(3′-thiotriphosphate) to activate G-proteins did not affect intracellular Ca2+ levels but did induce cortical contraction. Tyrosine kinase inhibitors (tyrphostin and staurosporine) suppressed the Mg2+-induced Ca2+ rise and contraction, and the inhibition could be overcome by injection with inositol 1,4,5-trisphosphate. Immunoprecipitation of phosphotyrosine-containing proteins from oocyte lysates showed that a 110×103Mr protein was phosphorylated within seconds of oocyte exposure to Mg2+, and tyrphostin inhibited the phosphorylation of this protein. Pre-treatment of oocytes with the protease trypsin abolished their ability to release Ca2+ in response to extracellular Mg2+, indicating a role for a cell surface protein during normal oocyte activation; treated oocytes could be rescued by inositol 1,4,5-trisphosphate injection. These results suggest that S. ingentis oocytes are activated through a Mg2+ ‘receptor’ which activates a tyrosine kinase and results in the production of inositol 1,4,5-trisphosphate to release intracellular Ca2+ stores and induce cortical con-traction. A G-protein/GTPase may also be involved in the pathway leading to cortical contraction.