Abstract

Activation of mitogen-activated protein kinase (MAPK) is induced by adding thyrotropin-releasing hormone (TRH) to COS-7 cells cotransfected with TRH receptors and an epitope-tagged MAPK. Long term treatment of the cells with pertussis toxin has no effect on TRH-induced MAPK activation. Incubation of the cells with the protein kinase C (PKC) inhibitor GF109203X causes an almost complete inhibition of MAPK activation by the PKC activator phorbol-12-myristate-13-acetate. In contrast, only ∼50% of the TRH-induced MAPK activity is inhibited by GF109203X, indicating that activation of MAPK by TRH is only partially dependent on PKC. The inhibitory effect of GF109203X is additive with that of p21^N17ras, a dominant negative mutant of p21^ras that exerts little effect on PKC-dependent MAPK activation by phorbol-12-myristate-13-acetate. The TRH-induced activation of MAPK also is inhibited partially by overexpression of transducin α subunits (αt), an agent known to sequester free G protein βγ dimers. However, the inhibitory potency of αt on TRH-induced activation is about half of that obtained in cells transfected with m2 muscarinic receptors, which activate MAPK exclusively through βγ dimers. The effect of αt is also additive with that of GF109203X but not with that of p21^N17ras. MAPK activation is not induced by the constitutively active form of G_αq due to an inhibitory effect of its expression at a step downstream of that at which PKC-dependent and -independent routes to MAPK converge. Our results demonstrate that TRH receptors activate MAPK by a pathway only partially dependent on PKC activity. Furthermore, they indicate that βγ dimers of a pertussis and cholera toxin-insensitive G protein are involved in the PKC-independent fraction of the dual signaling route to MAPK initiated in the TRH receptor.