PROTEIN KINASE-A (PKA) STILL ACTIVATES CFTR CHLORIDE CHANNEL AFTER MUTAGENESIS OF ALL 10 PKA CONSENSUS PHOSPHORYLATION SITES

The cystic fibrosis transmembrane conductance regulator (CFTR) plays a central role in transepithelial ion transport by acting as a tightly regulated apical chloride channel. Regulation is achieved by the conce rted action of ATP at conserved nucleotide binding folds and serine ph osphorylation at multiple sites by protein kinases A (PKA) and C (PKC) . A previous investigation concluded that activation by PKA is critica lly dependent on phosphorylation at four of the nine predicted PKA sit es in the R domain (S660A, S737A, S795A, S813A), because a ''Quad'' mu tant lacking these sites could not be activated. We show in the presen t work that not only can this mutant be phosphorylated and activated, but a mutant in which all 10 predicted PKA sites have been altered sti ll retains significant PKA-activated function. Potentiation of the PKA response by PKC is also preserved in this mutant. Thus CFTR may be re gulated by cryptic PKA sites which also mediate interactions between d ifferent kinases. Such hierarchical phosphorylation of CFTR by obvious and cryptic PKA sites could provide a metered response to secretagogu es.