Co‐operation of G_sα and G_βγ in maintaining G₂ arrest in <i>xenopus</i> oocytes

<jats:title>Abstract</jats:title><jats:p>Progesterone‐induced oocyte maturation is thought to involve the inhibition of an oocyte adenylyl cyclase and reduction of intracellular cAMP. Our previous studies demonstrated that injection of inhibitors of G protein βγ complex induces hormone‐independent oocyte maturation. In contrast, over‐expression of <jats:italic>Xenopus</jats:italic> G<jats:sub>β1</jats:sub> (xG<jats:sub>β1</jats:sub>), alone or together with bovine G<jats:sub>γ2</jats:sub>, elevates oocyte cAMP and inhibits progesterone‐induced oocyte maturation. To further investigate the mechanism of G<jats:sub>βγ</jats:sub>‐induced oocyte maturation, we generated a mutant xG<jats:sub>β1</jats:sub>, substituting Asp‐228 for Gly (D228G). An equivalent mutation in the mammalian G<jats:sub>β1</jats:sub> results in the loss of its ability to activate adenylyl cyclases. Indeed, co‐injection of xG<jats:sub>β1D228G</jats:sub> with G<jats:sub>γ2</jats:sub> failed to increase oocyte cAMP or inhibit progesterone‐induced oocyte maturation. To directly demonstrate that oocytes contained a G<jats:sub>βγ</jats:sub>‐regulated adenylyl cyclase, we analyzed cAMP formation in vitro by using oocyte membrane preparations. Purified brain G<jats:sub>βγ</jats:sub> complexes significantly activated membrane‐bound adenylyl cyclase activities. Multiple adenylyl cyclase isoforms were identified in frog oocytes by PCR using degenerate primers corresponding to highly conserved catalytic amino acid sequences. Among these we identified a partial <jats:italic>Xenopus</jats:italic> adenylyl cyclase 7 (xAC7) that was 65% identical in amino acid sequence to human AC7. A dominant‐negative mutant of xAC7 induced hormone‐independent oocyte maturation and accelerated progesterone‐induced oocyte maturation. Theses findings suggest that xAC7 is a major component of the G<jats:sub>2</jats:sub> arrest mechanism in <jats:italic>Xenopus</jats:italic> oocytes. © 2005 Wiley‐Liss, Inc.</jats:p>