Role of a polybasic motif in plasma membrane targeting and signaling of G protein α subunits

Heterotrimeric G proteins are involved in a wide variety of physiological processes and diseases. They exist as a heterotrimer of α, β, and γ subunits, with the Gα bound to GDP or GTP, depending on its activation state. Heterotrimeric G proteins transmit signals from extracellular stimuli to intracellular effectors. They must be localized at the cytoplasmic face of the plasma membrane to do so. Membrane targeting signals for Gα include myristoylation and/or palmitoylation, as well as interaction with Gβγ. It has also been postulated that other membrane targeting signals exist for the α subunits that aren\u27t myristoylated. One possibility is a polybasic motif present in the N-termini of these a subunits. Mutations in the basic residues in the N-termini of αs and αq and inhibit palmitoylation and plasma membrane localization. Myristoylation can recover these defects, supporting the proposal that myristoylation and this polybasic motif have complementary roles as a membrane targeting signal for G protein aα subunits. Co-expression of Gβγ was able to recover plasma membrane localization, though not to the same extent as wild type Gα, indicating that the defects in plasma membrane localization caused by mutations in the polybasic motif are observed even in the context of the heterotrimer. Polybasic mutants of αs and αq defective in plasma membrane localization were also defective in signaling through receptors. Myristoylation and co-expression of Gβγ did not recover signaling of these mutants, even though they recovered plasma membrane localization. Also, a polybasic mutant of αq was defective in signaling even though it was properly localized at the plasma membrane. This mutant was able to interact with Gβγ, RGS4, and was found to properly couple to receptors and activate effectors. This mutant displayed a diminished localization in plasma membrane microdomains compared to wild type αq, which may account for the defect in signaling of this mutant. Further mutagenesis analysis confirmed that it was the positive charges of these basic residues that were important for plasma membrane localization and signaling. Overall, basic residues in the N-termini of αs and αq are required for proper plasma membrane localization, palmitoylation, signaling, and plasma membrane microdomain localization of α q