Heterotrimeric guanine-nucleotide binding proteins (G proteins) of the G12/13 subfamily play key signaling roles in cell growth, oncogenic transformation, migration, and invasion. In this subfamily, mammals encode two distinct G protein alpha subunits (Gα12 and Gα13) which share 67% amino acid identity. Gα12/13 also share several downstream signaling pathways, including transcriptional activation via Serum Response Factor (SRF). However, both proteins have multiple, unique binding partners, suggesting divergent signaling mechanisms within this subfamily. Amino acid substitutions from non-SRF signaling invertebrate homologs were utilized to identify different structural features required for growth signaling in Gα12 and Gα13. Previous work showed that replacing the N-terminal region of these mammalian α subunits with invertebrate amino acid sequence caused no disruption of SRF signaling. Conversely, invertebrate substitution of a highly conserved sequence downstream of the Switch III region abolished SRF signaling by Gα12 and Gα13. In current studies of this “Post-Switch region” of Gα12, a single amino acid substitution disrupting signaling to SRF, while the same mutation in Gα13 had no effect. Replacement of additional amino acids in this region were required to uncouple Gα13 from SRF activation, suggesting divergent growth signaling mechanisms in the G12/13 subfamily. Additionally, invertebrate substitution in the Post-Switch region disrupted Gα13 interaction with cancer-implicated proteins, termed RhoGEFs, whereas the corresponding SRF-uncoupled Gα12 mutant retained binding to these effector proteins. These findings reveal novel C-terminal motifs in Gα12 and Gα13 that are necessary for their non-overlapping mechanisms of RhoGEF binding and SRF-mediated cell growth. Since certain cancers selectively overexpress Gα12 or Gα13, further characterization of these proteins could be used to guide the development of Gα12 or Gα13-specific inhibitors.
