Understanding the Role of Gβγ in the Cellular Regulation of Mutationally Activated Gαq/11.

Uveal melanoma (UM) is the most common intraocular malignancy in adults. UM metastasizes to the liver in 50% of patients, and there are currently no effective therapies for metastatic UM. Approximately 90% of UM cases harbor mutually exclusive mutations in the GNAQ and GNA11 genes, which code for the heterotrimeric G protein subunits Gαq and Gα11 , respectively. The most common mutations within these genes are within residues Q209 and R183, resulting in Q209L, Q209P, or R183C mutations in UM patients. These mutations prevent GTP hydrolysis and render the proteins constitutively active (CA) to promote oncogenic signaling. The Ras/mitogen-activated protein kinase (MAPK) and Hippo/YAP pathways are commonly stimulated via CA Gαq/11 . Although it is generally thought that active, GTP-bound Gα subunits signal independently of Gβγ, we have found that these CA mutants of Gαq/11 retain some level of binding to Gβγ and that this interaction is necessary for oncogenic signaling. The main objective of this project is to understand the role of Gβγ in the regulation of CA Gαq/11 . We hypothesized that disrupting the interaction between CA Gαq/11 and Gβγ can ultimately inhibit oncogenic signaling in UM. To test this hypothesis, we introduced a single point mutation within the N-terminus of the CA Gαq mutants, which has been shown previously to disrupt the interaction between wild-type Gαq and Gβγ and prevent agonist-dependent signaling of Gαq. We also overexpressed another Gα subunit, Gαo , to bind endogenous Gβγ and thereby prevent Gβγ binding by CA Gαq mutants. We found that these methods disrupted binding of CA Gαq to Gβγ and further disrupted oncogenic signaling via the Hippo/YAP and MAPK pathways. Interestingly, we found that the Gαq Q209L mutant binds more strongly to Gβγ, and signaling by the Gαq Q209P and Gαq R183C mutants are more sensitive to disruption of Gβγ binding. This was further confirmed within UM cell lines where siRNA knockdown of Gβ1 and Gβ2 had a more profound effect on inhibition of signaling in cells with the Gαq Q209P mutation compared to cell lines with the Gαq Q209L mutation. In conclusion, our study challenges the idea that CA Gαq/11 signals independently of Gβγ. Our work also suggests a novel difference in sensitivity between the Gαq Q209L, Gαq Q209P, and Gαq R183C mutants. We propose that disrupting the interaction between CA Gαq/11 and Gβγ can be a novel therapeutic target in UM. © FASEB.

Citation

Jenna Aumiller, Asuka Inoue, Philip Wedegaertner. Understanding the Role of Gβγ in the Cellular Regulation of Mutationally Activated Gαq/11. FASEB journal : official publication of the Federation of American Societies for Experimental Biology. 2022 May;36 Suppl 1

PMID: 35552720

search → result