Protein Kinase C Deficiency Blocks Recovery from Agonist-induced Desensitization

Abstract

Protein phosphorylation is central to agonist-induced attenuation of the function of G-protein-linked receptors. Stable expression of RNA antisense to specific protein kinase mRNAs permitted analysis of loss-of-function mutants of A431 human epidermoid carcinoma cells, lacking protein kinase A, protein kinase C, or beta-adrenergic receptor kinase. Deficiency of protein kinase C, but not the others, amplified rather than attenuated agonist-induced desensitization. In wild-type cells, the t1/2 for recovery from desensitization was approximately 25 min following removal of agonist. In the protein kinase C-deficient cells, no resensitization was observed even 60 min after agonist removal. Like protein kinase C-deficiency, inhibition of protein kinase C with bisindolylmaleimide or calphostin C blocked resensitization. Resensitization was suppressed by FK506, an inhibitor of protein phosphatase 2B, mimicking protein kinase C-deficiency, but in a non-additive manner. The data reveal protein kinase C and protein phosphatase 2B to be critical elements of resensitization.