CFTR potentiators partially restore channel function to A561E, a cystic fibrosis mutant with a similar mechanism of dysfunction as F508del-CFTR

Yiting Wang, J Liu, Avgi Loizidou, Luc A Bugeja, Ross Warner, Bethan R Hawley, Zhiwei Cai, Ashley M Toye, David N Sheppard, Hongyu Li

Research output: Contribution to journalArticle (Academic Journal)peer-review

23 Citations (Scopus)


BACKGROUND AND PURPOSE: Dysfunction of the cystic fibrosis transmembrane conductance regulator (CFTR) Cl(-) channel causes the genetic disease cystic fibrosis (CF). Towards the development of transformational drug therapies for CF, we investigated the channel function and action of CFTR potentiators on A561E, a CF mutation found frequently in Portugal. Like the most common CF mutation F508del, A561E causes a temperature-sensitive folding defect that prevents CFTR delivery to the cell membrane and is associated with severe disease.

EXPERIMENTAL APPROACH: Using baby hamster kidney (BHK) cells expressing recombinant CFTR, we investigate CFTR expression by cell surface biotinylation, and function and pharmacology with the iodide efflux and patch-clamp techniques.

KEY RESULTS: Low temperature incubation delivered a small proportion of A561E-CFTR protein to the cell surface. Like F508del-CFTR, low temperature-rescued A561E-CFTR exhibited a severe gating defect characterized by brief channel openings separated by prolonged channel closures. A561E-CFTR also demonstrated thermoinstability, losing function more quickly than F508del-CFTR in cell-free membrane patches and intact cells. Using the iodide efflux assay, CFTR potentiators, including genistein and the clinically-approved small-molecule ivacaftor, restored partial function to A561E-CFTR. Interestingly, ivacaftor restored wild-type levels of channel activity (as measured by open probability) to single A561E- and F508del-CFTR Cl(-) channels. However, it accentuated the thermoinstability of both mutants in cell-free membrane patches.

CONCLUSIONS AND IMPLICATIONS: Like F508del-CFTR, A561E-CFTR perturbs protein processing, thermostability and channel gating. CFTR potentiators partially restore channel function to low temperature-rescued A561E-CFTR. Transformational drug therapy for A561E-CFTR will likely require CFTR correctors, CFTR potentiators and special attention to thermostability.

Original languageEnglish
JournalBritish Journal of Pharmacology
Publication statusPublished - 5 Jun 2014

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