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Association of the proto-oncogene product dbl with G protein betagamma subunits
1Faculty of Bioscience and Biotechnology, Tokyo Institute of Technology, 4259 Nagatsuta-cho, Midori-ku, Yokohama, Japan.
Abstract:
The Rho family of GTP-binding proteins has been implicated in the regulation of various cellular functions including actin cytoskeleton-dependent morphological change. Its activity is directed by intracellular signals mediated by various types of receptors such as G protein-coupled receptors. However, the mechanisms underlying receptor-dependent regulation of Rho family members remain incompletely understood. The guanine nucleotide exchange factor (GEF) Dbl targets Rho family proteins thereby stimulating their GDP/GTP exchange, and thus is believed to be involved in receptor-mediated regulation of the proteins. Here, we show the association of Dbl with G protein betagamma subunits (Gbetagamma) in transient co-expression and cell-free systems. An amino-terminal portion conserved among a subset of Dbl family proteins is sufficient for the binding of Gbetagamma. In fact, Ost and Kalirin, which contain this Gbetagamma-binding motif, also associate with Gbetagamma. c-Jun N-terminal kinase was synergistically activated upon co-expression of Dbl and Gbeta in a dominant-negative Rho-sensitive manner. However, GEF activity of Dbl toward Rho as measured by in vitro GDP binding assays remained unaffected following Gbetagamma binding, suggesting that additional signals may be required for the regulation of Dbl.
Insights
Guanine nucleotide exchange factor Dbl binds G protein betagamma subunits, impacting cellular signaling. This interaction, however, did not directly alter Dbl
Area of Science:
- Molecular Biology
- Cell Signaling
- Biochemistry
Background:
- Rho family GTP-binding proteins regulate actin cytoskeleton and cell morphology.
- Receptor-mediated signals control Rho family protein activity, but mechanisms are unclear.
- Guanine nucleotide exchange factor (GEF) Dbl is implicated in receptor-mediated Rho regulation.
Purpose of the Study:
- To investigate the interaction between Dbl and G protein betagamma (Gbetagamma) subunits.
- To determine if Gbetagamma binding affects Dbl's guanine nucleotide exchange factor (GEF) activity.
- To explore the role of Dbl-Gbetagamma interaction in cellular signaling pathways.
Main Methods:
- Transient co-expression systems in cells.
- Cell-free binding assays.
- In vitro GDP binding assays to measure GEF activity.
- Analysis of c-Jun N-terminal kinase activation.
Main Results:
- Dbl directly associates with Gbetagamma subunits via its amino-terminal domain.
- Other Dbl family proteins (Ost, Kalirin) also bind Gbetagamma.
- Co-expression of Dbl and Gbeta synergistically activated c-Jun N-terminal kinase.
- Gbetagamma binding did not alter Dbl's intrinsic GEF activity towards Rho.
Conclusions:
- Dbl interacts with Gbetagamma subunits, suggesting a role in G protein-coupled receptor signaling.
- The direct GEF activity of Dbl is not modulated by Gbetagamma binding alone.
- Additional regulatory signals are likely required for Dbl activation in response to Gbetagamma.