Visualization of G Protein βγ Dimers Using Bimolecular Fluorescence Complementation Demonstrates Roles for Both β and γ in Subcellular Targeting*
By T. Hynes, L. Tang, Stacy M. Mervine, Jonathan L. Sabo, E. Yost, P. Devreotes, C. Berlot. Journal of Biological Chemistry, 2004
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To investigate the role of subcellular localization in regulating the specificity of G protein βγ signaling, we have applied the strategy of bimolecular fluorescence complementation (BiFC) to visualize βγ dimers in vivo. We fused an amino-terminal yellow fluorescent protein fragment to β and a carboxyl-terminal yellow fluorescent protein fragment to γ. When expressed together, these two proteins produced a fluorescent signal in human embryonic kidney 293 cells that was not obtained with either subunit alone. Fluorescence was dependent on βγ assembly in that it was not obtained using β2 and γ1, which do not form a functional dimer. In addition to assembly, BiFC βγ complexes were functional as demonstrated by more specific plasma membrane labeling than was obtained with individually tagged fluorescent β and γ subunits and by their abilities to potentiate activation of adenylyl cyclase by αs in COS-7 cells. To investigate isoform-dependent targeting specificity, the localization patterns of dimers formed by pair-wise combinations of three different β subunits with three different γ subunits were compared. BiFC βγ complexes containing either β1 or β2 localized to the plasma membrane, whereas those containing β5 accumulated in the cytosol or on intracellular membranes. These results indicate that the β subunit can direct trafficking of the γ subunit. Taken together with previous observations, these results show that the G protein α, β, and γ subunits all play roles in targeting each other. This method of specifically visualizing βγ dimers will have many applications in sorting out roles for particular βγ complexes in a wide variety of cell types.
Subfields: Biophysics, Molecular Biology
View Paper: https://www.semanticscholar.org/paper/836784fbe40dec49495702a8989f6a79ec6a352c
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Citations in the corpus
- Visualization of protein interactions in living plant cells using bimolecular fluorescence complementation. (2004, background): “As demonstrated by our study and recent reports (Grinberg et al., 2004; Hynes et al., 2004) BiFC-generated fluorescence signals can also be quantified.”