Rescue of CFTR NBD2 mutants N1303K and S1235R is influenced by the functioning of the autophagosome

J Cyst Fibros. 2018 Sep;17(5):582-594. doi: 10.1016/j.jcf.2018.05.016. Epub 2018 Jun 20.


The missing phenylalanine at position 508, located in nucleotide-binding domain (NBD1) of the cystic fibrosis transmembrane regulator (CFTR), is the most common cystic fibrosis mutation. Severe disease-causing mutations also occur in NBD2. To provide information on potential therapeutic strategies for mutations in NBD2, we used a combination of biochemical, cell biological and electrophysiological approaches and newly created cell lines to study two disease-causing NBD2 mutants, N1303K and S1235R. We observed that neither was sensitive to E64, a cysteine protease inhibitor. However, further investigation showed that when treated with a combination of correctors, C4 + C18, both mutants also responded to E64. Further exploration to assess aggresome throughput using the autophagy regulator LC3 as a marker showed that, in the absence of correctors, N1303K showed a stalled throughput of LC3-II to the aggresome. The throughput became active again after treatment with the corrector combination C4 + C18. Confocal microscopic studies showed that the N1303K and S1235R mutant proteins both co-localized with LC3, but this co-localization was abolished by the corrector combination and, to a lesser extent, by VX-809. Both the corrector combination and VX-809 increased the CFTR chloride channel function of both mutants. We conclude that correctors have a dual effect, particularly on N1303K: they improve trafficking and function at the plasma membrane and reduce the association with autophagosomes. After treatment with correctors persistent degradation by the autophagosome may limit restoration of function. Thus, mutations in NBD2 of CFTR, in contrast to ΔF508-CFTR, may require additional personalized strategies to rescue them.

Keywords: Autophagy; CFTR; Correctors; Degradation; Mutations; Short circuit current; Western blot.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Aminopyridines / pharmacology*
  • Animals
  • Autophagosomes / physiology*
  • Autophagy
  • Benzodioxoles / pharmacology*
  • Biological Transport
  • Blotting, Western
  • Cell Line
  • Cystic Fibrosis / drug therapy*
  • Cystic Fibrosis / genetics*
  • Cystic Fibrosis Transmembrane Conductance Regulator / drug effects*
  • Cystic Fibrosis Transmembrane Conductance Regulator / genetics*
  • Electric Conductivity
  • Leucine / analogs & derivatives*
  • Leucine / pharmacology
  • Mutant Proteins / drug effects
  • Mutant Proteins / genetics
  • Mutation
  • Small Molecule Libraries / pharmacology


  • Aminopyridines
  • Benzodioxoles
  • Mutant Proteins
  • Small Molecule Libraries
  • Cystic Fibrosis Transmembrane Conductance Regulator
  • lumacaftor
  • Leucine
  • E 64