Activation of STAT3 by G alpha(s) distinctively requires protein kinase A, JNK, and phosphatidylinositol 3-kinase.

Abstract Signal transducer and activator of transcription 3 (STAT3) can be stimulated by several Gs-coupled receptors, but the precise mechanism of action has not yet been elucidated. We therefore examined the ability of GαsQ226L (GαsQL), a constitutively active mutant of Gαs, to stimulate STAT3 Tyr705 and Ser727 phosphorylations in human embryonic kidney 293 cells. Apart from GαsQL, the stimulation of Gαs by cholera toxin or β2-adrenergic receptor and the activation of adenylyl cyclase by forskolin, (Sp)-cAMP, or dibutyryl-cAMP all promoted both STAT3 Tyr705 and Ser727 phosphorylations. Moreover, the removal of Gαs by RNA interference significantly reduced the β2-adrenergic receptor-mediated STAT3 phosphorylations, denoting its capacity to regulate STAT3 activation by a G protein-coupled receptor. The possible downstream signaling molecules involved were assessed by using specific inhibitors and dominant negative mutants. Induction of STAT3 Tyr705 and Ser727 phosphorylations by GαsQL was suppressed by inhibition of protein kinase A, Janus kinase 2/3, Rac1, c-Jun N-terminal kinase (JNK), or phosphatidylinositol 3-kinase, and a similar profile was observed in response to β2-adrenergic receptor stimulation. In contrast to the Gα16-mediated regulation of STAT3 in HEK 293 cells (Lo, R. K., Cheung, H., and Wong, Y. H. (2003) J. Biol. Chem. 278, 52154–52165), the Gαs-mediated responses, including STAT3-driven luciferase activation, were resistant to inhibition of phospholipase Cβ. Surprisingly, Gαs-mediated phosphorylation at Tyr705, but not at Ser727, was resistant to inhibition of c-Src, Raf-1, and MEK1/2 as well as to the expression of dominant negative Ras. Therefore, as with other Gα-mediated activations of STAT3, the stimulatory signal arising from Gαs is transduced via multiple signaling pathways. However, unlike the mechanisms employed by Gαi and Gα14/16, Gαs distinctively requires protein kinase A, JNK, and phosphatidylinositol 3-kinase for STAT3 activation.