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 concerted action of ATP at conserved nucleotide binding folds and serine phosphorylation at multiple sites by protein kinases A (PKA) and C (PKC). A previous investigation concluded that activation by PKA is critically dependent on phosphorylation at four of the nine predicted PKA sites in the R domain (S660A, S737A, S795A, S813A), because a "Quad" mutant lacking these sites could not be activated. We show in the present work that not only can this mutant be phosphorylated and activated, but a mutant in which all 10 predicted PKA sites have been altered still retains significant PKA-activated function. Potentiation of the PKA response by PKC is also preserved in this mutant. Thus CFTR may be regulated by cryptic PKA sites which also mediate interactions between different kinases. Such hierarchical phosphorylation of CFTR by obvious and cryptic PKA sites could provide a metered response to secretagogues.
Chang
X B
XB
Research Institute, Hospital for Sick Children, Toronto, Ontario, Canada.
Tabcharani
J A
JA
Hou
Y X
YX
Jensen
T J
TJ
Kartner
N
N
Alon
N
N
Hanrahan
J W
JW
Riordan
J R
JR
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D016466:CHO Cells; D018118:Chloride Channels; D003062:Codon / Q000235:genetics; D006224:Cricetinae; D003550:Cystic Fibrosis / Q000235:genetics / Q000378:metabolism; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D006801:Humans; D007454:Iodides / Q000378:metabolism; D007473:Ion Channels / Q000502:physiology*; D007700:Kinetics; D008564:Membrane Potentials; D008565:Membrane Proteins / Q000235:genetics* / Q000378:metabolism*; D016297:Mutagenesis, Site-Directed; D010710:Phosphates / Q000378:metabolism; D010766:Phosphorylation; D016133:Polymerase Chain Reaction / Q000379:methods; D011494:Protein Kinases / Q000378:metabolism*; D017433:Protein Structure, Secondary; D011994:Recombinant Proteins / Q000378:metabolism; D015183:Restriction Mapping; D014162:Transfection
C505032:CFTR protein, human; D018118:Chloride Channels; D003062:Codon; D007454:Iodides; D007473:Ion Channels; D008565:Membrane Proteins; D010710:Phosphates; D011994:Recombinant Proteins; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D011494:Protein Kinases