Ga12 Structural Determinants of Hsp90 Interaction Are Necessary for Serum Response Element–Mediated Transcriptional Activation

The G12/13 class of heterotrimeric G proteins, comprising the a-subunits Ga12 and Ga13, regulates multiple aspects of cellular behavior, including proliferation and cytoskeletal rearrangements. Although guanine nucleotide exchange factors for the monomeric G protein Rho (RhoGEFs) are well characterized as effectors of this G protein class, a variety of other downstream targets has been reported. To identify Ga12 determinants that mediate specific protein interactions, we used a structural and evolutionary comparison between the G12/13, Gs, Gi, and Gq classes to identify “class-distinctive” residues in Ga12 and Ga13. Mutation of these residues in Ga12 to their deduced ancestral forms revealed a subset necessary for activation of serum response element (SRE)–mediated transcription, a G12/13-stimulated pathway implicated in cell proliferative signaling. Unexpectedly, this subset of Ga12 mutants showed impaired binding to heat-shock protein 90 (Hsp90) while retaining binding to RhoGEFs. Corresponding mutants of Ga13 exhibited robust SRE activation, suggesting a Ga12-specific mechanism, and inhibition of Hsp90 by geldanamycin or small interfering RNA–mediated lowering of Hsp90 levels resulted in greater downregulation of Ga12 than Ga13 signaling in SRE activation experiments. Furthermore, the Drosophila G12/13 homolog Concertina was unable to signal to SRE in mammalian cells, and Ga12:Concertina chimeras revealed Ga12-specific determinants of SRE activation within the switch regions and a C-terminal region. These findings identify Ga12 determinants of SRE activation, implicate Ga12:Hsp90 interaction in this signaling mechanism, and illuminate structural features that arose during evolution of Ga12 and Ga13 to allow bifurcated mechanisms of signaling to a common cell proliferative