Understanding Gαq/11 Localization and Trafficking in Uveal Melanoma

Abstract

Constitutively activating mutations in Gαq and Gα11 (Gαq/11) have been reported in up to 93% of uveal melanomas. Although constitutively active Gαq/11 promote uveal melanoma tumorigenesis through activation of multiple downstream pathways, no therapies inhibit constitutively active Gαq/11. Studies suggest that increased palmitate turnover and increased cytoplasmic localization occur upon activation of some Gα subunits. The purpose of our studies is to understand the role of palmitoylation in trafficking and signaling of constitutively active Gαq/11 in uveal melanoma. Using live cell imaging and cellular fractionation of HEK 293 cells and uveal melanoma cells, constitutively active GαqQ209L and GαqQ209P show decreased localization at membranes compared to wild type Gαq, suggesting increased turnover of attached palmitate in constitutively active mutants. Moreover, immunofluorescence microscopy revealed that a palmitoylation‐deficient constitutively active mutant displays a complete loss of plasma membrane localization and an inability to signal as measured by Rho‐ and Rac‐dependent YAP translocation into the nucleus, TEAD‐dependent luciferase activity, and ERK phosphorylation. These studies demonstrate that palmitoylation of mutationally activated Gαq/11 is required for its signaling functions. Although live cell imaging and cellular fractionation experiments reveal decreased membrane localization of constitutively active Gαq compared to wild‐type Gαq, our studies suggest that complete loss of membrane localization abolishes constitutively active Gαq‐dependent signaling. Disruption of constitutively active Gαq/11 palmitoylation and localization may be an effective strategy for inhibiting constitutively active Gαq/11 oncogenic signaling in uveal melanoma.