Glucagon receptor activates extracellular signal-regulated protein kinase 1/2 via cAMP-dependent protein kinase

We prepared a stable cell line expressing the glucagon receptor to characte rize the effect of G(s)-coupled receptor stimulation on extracellular signa l-regulated protein kinase 1/2 (ERK1/2) activity. Glucagon treatment of the cell line caused a dose-dependent increase in cAMP concentration, activati on of cAMP-dependent protein kinase (PKA), and transient release of intrace llular calcium. Glucagon treatment also caused rapid dose-dependent phospho rylation and activation of mitogen-activated protein kinase kinase/ERK kina se (MEK1/2) and ERK1/2. Inhibition of either PKA or MEK1/2 blocked ERK1/2 a ctivation by glucagon. However, no significant activation of several upstre am activators of MEK, including Ras, Rap1, and Raf, was observed in respons e to glucagon treatment. In addition, chelation of intracellular calcium re duced glucagon-mediated ERK1/2 activation. In transient transfection experi ments, glucagon receptor mutants that bound glucagon but failed to increase intracellular cAMP and calcium concentrations showed no glucagon-stimulate d ERK1/2 phosphorylation. We conclude that glucagon-induced MEK1/2 and ERK1 /2 activation is mediated by PKA and that an increase in intracellular calc ium concentration is required for maximal ERK activation.