The major cystic fibrosis causing mutation exhibits defective propensity for phosphorylation

Proteomics. 2015 Jan;15(2-3):447-61. doi: 10.1002/pmic.201400218. Epub 2014 Dec 17.


The major cystic fibrosis causing mutation, F508del-CFTR (where CFTR is cystic fibrosis transmembrane conductance regulator), impairs biosynthetic maturation of the CFTR protein, limiting its expression as a phosphorylation-dependent channel on the cell surface. The maturation defect can be partially rescued by low-temperature (27°C) cell culture conditions or small-molecule corrector compounds. Following its partial rescue, the open probability of F508del-CFTR is enhanced by the potentiator compound, VX-770. However, the channel activity of rescued F508del-CFTR remains less than that of the Wt-CFTR protein in the presence of VX-770. In this study, we asked if there are allosteric effects of F508del on the phosphorylation-regulated R domain. To identify defects in the R domain, we compared the phosphorylation status at protein kinase A sites in the R domain of Wt and F508del-CFTR. Here we show that phosphorylation of Ser-660, quantified by SRM-MS, is reduced in F508del-CFTR. Although the generation of a phosphomimic at this site (substituting aspartic acid for serine) did not modify the maturation defect, it did enhance F508del-CFTR channel function after pharmacological rescue with corrector VX-809, and treatment with the potentiator, VX-770. These findings support the concept that defective phosphorylation of F508del-CFTR partially accounts for its altered channel activity at the cell surface.

Keywords: Cell biology; Cystic fibrosis; Mass spectrometry; Phosphorylation; Protein kinase A (PKA); Protein misfolding.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Cell Line
  • Cricetinae
  • Cyclic AMP-Dependent Protein Kinases / metabolism
  • Cystic Fibrosis / genetics*
  • Cystic Fibrosis / metabolism*
  • Cystic Fibrosis Transmembrane Conductance Regulator / chemistry
  • Cystic Fibrosis Transmembrane Conductance Regulator / genetics*
  • Cystic Fibrosis Transmembrane Conductance Regulator / metabolism*
  • HEK293 Cells
  • Humans
  • Molecular Sequence Data
  • Phosphorylation
  • Protein Structure, Tertiary
  • Sequence Deletion


  • Cystic Fibrosis Transmembrane Conductance Regulator
  • Cyclic AMP-Dependent Protein Kinases