Polyspermy block in jellyfish eggs: collaborative controls by Ca(2+) and MAPK

Jellyfish eggs neither undergo apparent cortical reaction nor show any significant change in the membrane potential at fertilization, but nevertheless show monospermy. Utilizing the perfectly transparent eggs of the hydrozoan jellyfish Cytaeis uchidae, here we show that the polyspermy block is accomplished via a novel mechanism: a collaboration between Ca(2+) and mitogen-activated protein kinase (MAPK). In Cytaeis, adhesion of a sperm to the animal pole surface of an egg was immediately followed by sperm-egg fusion and initiation of an intracellular Ca(2+) rise from this site. The elevated Ca(2+) levels lasted for several minutes following the sperm-egg fusion. The Ca(2+) rise proved to be necessary and sufficient for a polyspermy block, as inhibiting a Ca(2+) rise with EGTA promoted polyspermy, and conversely, triggering a Ca(2+) rise by inositol 1,4,5-trisphosphate (IP3) or excess K(+) immediately abolished the egg's capacity for sperm-egg fusion. A Ca(2+) rise at fertilization or by artificial stimulations evoked dephosphorylation of MAPK in eggs. The eggs in which phosphorylated MAPK was maintained by injection of mRNA for MAPK kinase kinase (Mos), like intact eggs, exhibited a Ca(2+) rise at fertilization or by IP3 injection, and shut down the subsequent sperm-egg fusion. However, the Mos-expressing eggs became capable of accepting sperm following the arrest of Ca(2+) rise. In contrast, addition of inhibitors of MAPK kinase (MEK) to unfertilized eggs caused MAPK dephosphorylation without elevating Ca(2+) levels, and prevented sperm-egg fusion. Rephosphorylation of MAPK by injecting Mos mRNA after fertilization recovered sperm attraction, which is known to be another MAPK-dependent event, but did not permit subsequent sperm-egg fusion. Thus, it is possible that MAPK dephosphorylation irreversibly blocks sperm-egg fusion and reversibly suppresses sperm attraction. Collectively, our data suggest that both the fast and late mechanisms dependent on Ca(2+) and MAPK, respectively, ensure a polyspermy block in jellyfish eggs.