Roles of protein kinase C isotypes during seawater‐versus cAMP‐induced oocyte maturation in a marine worm

Abstract

Based on immunoblotting analyses using phospho-specific antibodies, follicle-free oocytes of the marine nemertean worm Cerebratulus sp. activate protein kinase C (PKC) when induced to mature by either seawater (SW) or cAMP-elevating drugs. In SW-stimulated oocytes, the onset of maturation (=germinal vesicle breakdown, "GVBD") can be inhibited by broadly acting PKC antagonists such as bisindoylmaleimide (BIM)-I or BIM-IX. Conversely, co-treatment with SW solutions of BIM-I or BIM-IX plus a cAMP elevator (forskolin, serotonin, or a phosphodiesterase inhibitor) restores GVBD, indicating that the blockage of SW-induced GVBD by PKC antagonists is not simply due to oocyte morbidity and that such inhibition is somehow reversible by cAMP signaling. In tests to determine which specific PKC may be involved in regulating GVBD, immunoblots fail to provide strong evidence for the presence of conventional or novel PKCs, which are characteristically activated by 12-O-tetradecanoylphorbol-13-acetate (TPA). Moreover, inhibitors of TPA-sensitive PKCs do not prevent SW-induced GVBD, and TPA itself serves to downregulate, rather than stimulate, GVBD. Alternatively, maturing oocytes apparently possess phosphorylated forms of TPA-insensitive isotypes, including an approximately 67-kDa atypical PKC and an approximately 130-kDa PKC-related kinase (PRK). Accordingly, inhibitors of atypical PKC signaling block SW-but not cAMP-induced GVBD, collectively suggesting that instead of depending on a conventional or novel isotype, SW-induced GVBD may require atypical PKC and/or PRK. In addition, such findings provide further support for the view that GVBD in nemertean oocytes can be achieved via multiple mechanisms, with SW triggering different signaling pathways than are stimulated in the presence of cAMP-elevating drugs.