Galpha12 stimulates apoptosis in epithelial cells through JNK1-mediated Bcl-2 degradation and up-regulation of IkappaBalpha.

Apoptosis is an essential mechanism for the maintenance of somatic tissues, and when dysregulated can lead to numerous pathological conditions. G proteins regulate apoptosis in addition to other cellular functions, but the roles of specific G proteins in apoptosis signaling are not well characterized. Galpha12 stimulates protein phosphatase 2A (PP2A), a serine/threonine phosphatase that modulates essential signaling pathways, including apoptosis. Herein, we examined whether Galpha12 regulates apoptosis in epithelial cells. Inducible expression of Galpha12 or constitutively active (QL)alpha12 in Madin-Darby canine kidney cells led to increased apoptosis with expression of QLalpha12, but not Galpha12. Inducing QLalpha12 led to degradation of the anti-apoptotic protein Bcl-2 (via the proteasome pathway), increased JNK activity, and up-regulated IkappaBalpha protein levels, a potent stimulator of apoptosis. Furthermore, the QLalpha12-stimulated activation of JNK was blocked by inhibiting PP2A. To characterize endogenous Galpha12 signaling pathways, non-transfected MDCK-II and HEK293 cells were stimulated with thrombin. Thrombin activated endogenous Galpha12 (confirmed by GST-tetratricopeptide repeat (TPR) pull-downs) and stimulated apoptosis in both cell types. The mechanisms of thrombin-stimulated apoptosis through endogenous Galpha12 were nearly identical to the mechanisms identified in QLalpha12-MDCK cells and included loss of Bcl-2, JNK activation, and up-regulation of IkappaBalpha. Knockdown of the PP2A catalytic subunit in HEK293 cells inhibited thrombin-stimulated apoptosis, prevented JNK activation, and blocked Bcl-2 degradation. In summary, Galpha12 has a major role in regulating epithelial cell apoptosis through PP2A and JNK activation leading to loss of Bcl-2 protein expression. Targeting these pathways in vivo may lead to new therapeutic strategies for a variety of disease processes.