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A G protein-coupled receptor kinase induces Xenopus oocyte maturation
1Ottawa Health Research Institute, Ottawa Hospital Civic Campus, 725 Parkdale Avenue, Ottawa, Ontario K1Y 4E9, Canada.
Abstract:
Several recent studies have suggested that resumption of oocyte meiosis, indicated by germinal vesicle breakdown or GVBD, involves inhibition of endogenous heterotrimeric G proteins in both frogs and mice. These studies imply that a heterotrimeric G protein(s), and hence its upstream activator (a G protein-coupled receptor or GpCR), is activated in prophase oocytes and is responsible for maintaining meiosis arrest. To test the existence and function of this putative GpCR, we utilized a mammalian G-protein-coupled receptor kinase (GRK3) and beta-arrestin-2, which together are known to cause GpCR desensitization. Injection of mRNA for rat GRK3 caused hormone-independent GVBD. The kinase activity of GRK3 was essential for GVBD induction as its kinase-dead mutant (GRK3-K220R) was completely ineffective. Another GRK3 mutant (GRK3-DeltaC), which lacked the C-terminal G(betagamma)-binding domain and which was not associated with oocyte membranes, also failed to induce GVBD. Furthermore, injection of rat beta-arrestin-2 mRNA also induced hormone-independent GVBD. Several inhibitors of clathrin-mediated receptor endocytosis (the clathrin-binding domain of beta-arrestin-2, concanavalin A, and monodansyl cadaverine) significantly reduced the abilities of GRK3/beta-arrestin-2 to induce GVBD. These results support the central role of a yet-unidentified GpCR in maintaining prophase arrest in frog oocytes and provide a potential means for its molecular identification.
Insights
Researches show that G protein-coupled receptors (GpCRs) maintain oocyte meiosis arrest. Inhibiting these receptors with GRK3 or beta-arrestin-2 triggers germinal vesicle breakdown (GVBD), suggesting a key role for GpCRs in this process.
Area of Science:
- Cell Biology
- Developmental Biology
- Reproductive Biology
Background:
- Recent studies suggest heterotrimeric G proteins regulate oocyte meiosis.
- These G proteins, when inhibited, lead to germinal vesicle breakdown (GVBD), indicating a role in maintaining meiotic arrest.
- This implies an activated G protein-coupled receptor (GpCR) in prophase oocytes maintains arrest.
Purpose of the Study:
- To investigate the existence and function of a putative GpCR involved in maintaining meiotic arrest.
- To explore the role of G-protein-coupled receptor kinase 3 (GRK3) and beta-arrestin-2 in oocyte maturation.
Main Methods:
- Injection of messenger RNA (mRNA) for rat GRK3 and beta-arrestin-2 into frog oocytes.
- Utilized kinase-dead (GRK3-K220R) and truncated (GRK3-DeltaC) GRK3 mutants.
- Assessed the effect of inhibitors of clathrin-mediated endocytosis on GRK3/beta-arrestin-2-induced GVBD.
Main Results:
- Injection of GRK3 mRNA induced hormone-independent GVBD, dependent on GRK3 kinase activity and its G(betagamma)-binding domain.
- Injection of beta-arrestin-2 mRNA also induced hormone-independent GVBD.
- Inhibitors of clathrin-mediated endocytosis significantly reduced GVBD induction by GRK3/beta-arrestin-2.
Conclusions:
- These findings strongly support the existence of an unidentified GpCR maintaining meiotic arrest in prophase frog oocytes.
- GRK3 and beta-arrestin-2 play a crucial role in desensitizing this putative GpCR, leading to meiosis resumption.
- This study provides a foundation for the molecular identification of the GpCR involved in oocyte meiotic arrest.