Gα_12/13 inhibition enhances the anticancer effect of bortezomib through PSMB5 downregulation

Bortezomib is a proteasome inhibitor approved for anticancer therapy. However, variable sensitivity of tumor cells exists in this therapy probably due to differences in the expression of proteasome subunits. Gα_12/13 serves modulators or signal transducers in diverse pathways. This study investigated whether cancer cells display differential sensitivity to bortezomib with reference to Gα_12/13 expression, and if so, whether Gα_12/13 affects the expression of proteasome subunits and their activities. Bortezomib treatment exhibited greater sensitivities in Huh7 and SNU886 cells (epithelial type) than SK-Hep1 and SNU449 cells (mesenchymal type) that exhibited higher levels of Gα_12/13. Overexpression of an active mutant of Gα₁₂ (Gα₁₂QL) or Gα₁₃ (Gα₁₃QL) diminished the ability of bortezomib to induce cytotoxicity in Huh7 cells. Moreover, transfection with the minigene that disturbs G protein-coupled receptor-G protein coupling (CT12 or CT13) increased it in SK-Hep1 cells. Consistently, MiaPaCa2 cells transfected with CT12 or CT13 exhibited a greater sensitivity to bortezomib. Evidence of Gα_12/13's antagonism on the anticancer effect of bortezomib was verified in the reversal by Gα₁₂QL or Gα₁₃QL of the minigenes' enhancement of cytotoxity. Real-time polymerase chain reaction assay enabled us to identify PSMB5, multicatalytic endopeptidase complex-like-1, and proteasome activator subunit-1 repression by CT12 or CT13. Furthermore, Gα_12/13 inhibition enhanced the ability of bortezomib to repress PSMB5, as shown by immunoblotting and proteasome activity assay. Moreover, this inhibitory effect on PSMB5 was attenuated by Gα₁₂QL or Gα₁₃QL. In conclusion, the inhibition of Gα_12/13 activities may enhance the anticancer effect of bortezomib through PSMB5 repression, providing insight into the Gα_12/13 pathway for the regulation of proteasomal activity.