Gialpha and Gbeta subunits both define selectivity of G protein activation by alpha2-adrenergic receptors.

Previous studies of the specificity of receptor interactions with G protein subunits in living cells have relied on measurements of second messengers or other downstream responses. We have examined the selectivity of interactions between alpha2-adrenergic receptors (alpha2R) and various combinations of Gialpha and Gbeta subunit isoforms by measuring changes in ...
FRET between Gialpha-yellow fluorescent protein and cyan fluorescent protein-Gbeta chimeras in HeLa cells. All combinations of Gialpha1, -2, or -3 with Gbeta1, -2, or -4 were activated to some degree by endogenous alpha2Rs as judged by agonist-dependent decreases in FRET. The degree of G protein activation is determined by the combination of Gialpha and Gbeta subunits rather than by the identity of an individual subunit. RT-PCR analysis and small interfering RNA knockdown of alpha2R subtypes, followed by quantification of radiolabeled antagonist binding, demonstrated that HeLa cells express alpha2a- and alpha2b-adrenergic receptor isoforms in a 2:1 ratio. Increasing receptor number by overexpression of the alpha2aR subtype minimized the differences among coupling preferences for Gialpha and Gbeta isoforms. The molecular properties of each Gialpha, Gbeta, and alpha2-adrenergic receptor subtype influence signaling efficiency for the alpha2-adrenergic receptor-mediated signaling pathway.
Mesh Terms:
Enzyme Activation, Fluorescence Resonance Energy Transfer, Fluorescent Dyes, GTP-Binding Protein alpha Subunits, Gi-Go, GTP-Binding Protein beta Subunits, HeLa Cells, Humans, Microscopy, Fluorescence, RNA, Small Interfering, Receptors, Adrenergic, alpha-2, Reverse Transcriptase Polymerase Chain Reaction, Signal Transduction, Substrate Specificity
Proc. Natl. Acad. Sci. U.S.A.
Date: Jan. 03, 2006
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