Role of Tyrosine Phosphorylation in the Muscarinic Activation of the Cystic Fibrosis Transmembrane Conductance Regulator (CFTR)
Résumé
Cystic fibrosis transmembrane conductance regulator (CFTR) is a chloride (Cl) channel, which plays an important
role in physiological anion and fluid secretion, and is defective in several diseases. Although its activation by PKA and PKC has
been studied extensively, its regulation by receptors is less well understood. To study signaling involved in CFTR activation, we
measured whole-cell Cl currents in BHK cells cotransfected with GPCRs and CFTR. In cells expressing the M3 muscarinic
acetylcholine receptor, the agonist carbachol (Cch) caused strong activation of CFTR through two pathways; the canonical
PKA-dependent mechanism and a second mechanism that involves tyrosine phosphorylation. The role of PKA was suggested by partial inhibition of cholinergic stimulation by the specific PKA inhibitor Rp-cAMPS. The role of tyrosine kinases was suggested by Cch stimulation of 15SA-CFTR and 9CACFTR, mutants that lack 15 PKA or 9 PKC consensus sequences and are unresponsive to PKA or PKC stimulation, respectively.
Moreover the residual Cch response was sensitive to inhibitors of the Pyk2 and Src tyrosine kinase family. Our results suggest
that tyrosine phosphorylation acts on CFTR directly and through inhibition of the phosphatase PP2A. Results suggest
that PKA and tyrosine kinases contribute to CFTR regulation by GPCRs that are expressed at the apical membrane of intestinal
and airway epithelia.