Human glucosamine-6-phosphate isomerase, a homologue of hamster oscillin, does not appear to be involved in Ca2+ release in mammalian oocytes

Injections of cytosolic preparations from mammalian sperm into oocytes have been shown to trigger calcium [Ca2+](i) oscillations and initiate activati on of development. Recently, a protein isolated from hamster sperm has been suggested to be involved in the generation of these oscillations and it wa s named "oscillin." The human homologue of hamster oscillin is glucosamine 6-phosphate isomerase (GPI, EC no. 5.3.1.10), an enzyme so far described to be involved in hexose phosphate metabolism, To assess the role of GPI on C a2+ signaling, a human recombinant protein was generated in a prokaryotic s ystem and injected into fura-2-dextran-loaded metaphase II (MII) mouse oocy tes. Injection of recombinant GPI failed to induce Ca2+ responses in 12/12 injected MII oocytes despite the fact that the recombinant GPI was active a s assessed by an enzymatic assay. Injection of buffer (0/6 oocytes) or fruc tose-6-phosphate, a product of GPI enzymatic reaction (0/5 oocytes), also f ailed to initiate Ca2+ responses. Conversely, injections of sperm cytosolic factor induced [Ca2+]i oscillations in all 17/17 oocytes. In addition, inj ection of recombinant GPI or GPI mRNA failed to induce parthenogenetic acti vation (0/30 oocytes). Immunofluorescence studies using an anti-GPI polyclo nal antibody (GK) resulted in localization of GPI to the sperm's equatorial region. Incubation of the GK antibody with sperm extracts failed to block the [Ca2+](i) responses induced by these extracts. Moreover, near complete depletion of GPI from sperm fractions by immunoprecipitation did not impair the ability of these fractions to induce [Ca2+](i) oscillations. In summar y, our results support the role of a sperm cytosolic component(s) in the ge neration of [Ca2+](i) oscillations during mammalian fertilization, although a protein other than GPl/oscillin is likely to be the active calcium relea sing factor, Mol. Reprod. Dev. 52:277-287, 1999. (C) 1999 Wiley-iiss, Inc.