Stimulation of CFTR activity by its phosphorylated R domain.
Phosphorylation controls the activity of ion channels in many tissues. In epithelia, the cystic fibrosis transmembrane conductance regulator (CFTR) chloride channel is activated by phosphorylation of serine residues in its regulatory (R) domain and then gated by binding and hydrolysis of ATP by the nucleotide-binding domains. Current models propose that the unphosphorylated R domain serves as an inhibitory particle that occludes the pore, much like the inhibitory 'ball' in Shaker K+ channels; presumably, phosphorylation relieves this inhibition. Here we test this by adding an R-domain peptide to a CFTR variant in which much of the R domain had been deleted (CFTR-deltaR/S660A): in contrast to predictions, we found that adding an unphosphorylated R domain to CFTR-deltaR/S660A did not inhibit activity, whereas a phosphorylated R-domain peptide stimulated activity. To investigate how phosphorylation controls activity, we studied channel gating and found that phosphorylation of the R domain increases the rate of channel opening by enhancing the sensitivity to ATP. Our results indicate that CFTR is regulated by a new mechanism in which phosphorylation of one domain stimulates the interaction of ATP with another domain, thereby increasing activity.
Winter
M C
MC
Howard Hughes Medical Institute, Department of Internal Medicine, University of Iowa College of Medicine, Iowa City 52242, USA.
Welsh
M J
MJ
D000255:Adenosine Triphosphate / Q000378:metabolism; D001665:Binding Sites; D003001:Cloning, Molecular; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli; D006367:HeLa Cells; D006801:Humans; D015640:Ion Channel Gating; D016297:Mutagenesis, Site-Directed; D010766:Phosphorylation; D012694:Serine / Q000378:metabolism
C505032:CFTR protein, human; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D012694:Serine; D000255:Adenosine Triphosphate; D017868:Cyclic AMP-Dependent Protein Kinases