Identification of G protein subunit alpha i3 as a promising oncotarget of LUAD

Exploring new oncotargets essential for lung adenocarcinoma (LUAD) cell growth is important. Here the bioinformatical studies revealed that Gαi3 expression is elevated in LUAD tissues and its overexpression correlates with poor survival of the patients. Moreover, overexpression of Gαi3 mRNA and protein was detected in LUAD tissues of patients as well as in primary/immortalized LUAD cells. In both primary and immortalized LUAD cells, genetic silencing (by viral shRNA) or knockout ("KO", through CRISPR/Cas9 method) of Gαi3 potently inhibited LUAD cell proliferation and mobility. The results of caspase-3 activity assay, caspase-9 activity assay, histone DNA ELISA, TUNEL nuclear staining and Annexin V staining showed that inhibition of Gαi3 expression promoted apoptosis. In addition, a significant decrease in mitochondrial membrane potential was found in Gαi3-deficient LUAD cells by JC-1 staining. Overexpression of Gαi3 strengthened the proliferation and migration of LUAD cell. Gene set enrichment analysis revealed that Gαi3 was closely related to PI3k/Akt/mTOR, which we validated experimentally. Akt-S6K phosphorylation was downregulated following Gαi3 silencing or KO, but augmented after Gαi3 overexpression in primary LUAD cells. Restoring Akt-S6K phosphorylation by a S473D constitutively-active mutant Akt1 ameliorated Gαi3 KO-induced LUAD cell proliferation inhibition, migration suppression and apoptosis. In vivo, the growth of subcutaneous LUAD xenografts was largely inhibited after intratumoral injection of Gαi3 shRNA-expressing adeno-associated virus (AAV). Gαi3 downregulation, Akt-mTOR inhibition, proliferation inactivation and apoptosis were detected in the Gαi3 shRNA-treated LUAD xenografts. Together, targeting Gαi3 potently inhibited LUAD cell growth in vitro and in vivo.