title: Mechanism of dysfunction of two nucleotide binding domain mutations in cystic fibrosis transmembrane conductance regulator that are associated with pancreatic sufficiency.
abstract: Variability in the severity of cystic fibrosis (CF) is in part due to specific mutations in the CF transmembrane conductance regulator (CFTR) gene. To understand better how mutations in CFTR disrupt Cl- channel function and to learn about the relationship between genotype and phenotype, we studied two CF mutants, A455E and P574H, that are associated with pancreatic sufficiency. A455E and P574H are located close to conserved ATP binding motifs in CFTR. Both mutants generated cAMP-stimulated apical membrane Cl- currents in heterologous epithelial cells, but current magnitudes were reduced compared with wild-type. Patch-clamp analysis revealed that both mutants had normal conductive properties and regulation by phosphorylation and nucleotides. These mutants had normal or increased Cl- channel activity: A455E had an open-state probability (Po) similar to wild-type, and P574H had an increased Po because bursts of activity were prolonged. However, both mutants produced less mature glycosylated protein, although levels were greater than observed with the delta F508 mutant. These changes in channel activity and processing provide a quantitative explanation for the reduced apical Cl- current. These data also dissociate structural requirements for channel function from features that determine processing. Finally, the results suggest that the residual function associated with these two mutants is sufficient to confer a milder clinical phenotype and infer approaches to developing treatments.
authors.affiliation: Howard Hughes Medical Institute, Department of Physiology and Biophysics, University of Iowa College of Medicine, Iowa City 52242.
authors.forename: D N
authors.initials: DN
authors.lastname: Sheppard
authors.forename: L S
authors.initials: LS
authors.lastname: Ostedgaard
authors.forename: M C
authors.initials: MC
authors.lastname: Winter
authors.forename: M J
authors.initials: MJ
authors.lastname: Welsh
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism*; D000818:Animals; D018118:Chloride Channels / Q000378:metabolism*; D000242:Cyclic AMP / Q000819:agonists; D003550:Cystic Fibrosis / Q000235:genetics; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D004847:Epithelial Cells; D004848:Epithelium / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D015640:Ion Channel Gating; D008565:Membrane Proteins / Q000235:genetics* / Q000378:metabolism; D009154:Mutation / Q000502:physiology*; D010179:Pancreas / Q000378:metabolism; D018408:Patch-Clamp Techniques; D010766:Phosphorylation; D051381:Rats; D011916:Rats, Inbred F344; D013961:Thyroid Gland / Q000502:physiology
chemical_list: C505032:CFTR protein, human; D018118:Chloride Channels; D008565:Membrane Proteins; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D000255:Adenosine Triphosphate; D000242:Cyclic AMP