Transcomplementation by a truncation mutant of cystic fibrosis transmembrane conductance regulator (CFTR) EnhancesδF508 processing through a biomolecular interaction

Abstract

We previously showed that a truncation mutant of CFTR missing the first four transmembrane segments of TMD1, 264 CFTR, binds to key elements in the ER quality control mechanism to increase the amounts of the mature C band of both wt and F508 CFTR through transcomplementation. Here, we created a new construct, 27-264 CFTR. Even though 27-264 CFTR is rapidly degraded in the proteasome, steady state protein can be detected by Western blot. 27-264 CFTR can also increase the amounts of the mature C band of both wt and F508 CFTR through transcomplementation. Electrophysiology experiments show that 27-264 CFTR can restore chloride channel currents. Further experiments with the conduction mutant S341A show conclusively that currents are indeed generated by rescued channel function of F508 CFTR. Immunoprecipitation studies show that 27-264 binds to F508-CFTR, suggesting a bimolecular interaction. Thus the adeno-associated viral vector, rAAV- 27-264 CFTR, is a highly promising CF gene therapy vector, because it increases the amount of mature band C protein both from wt and F508 CFTR, and rescues channel activity of F508 CFTR. © 2013 by The American Society for Biochemistry and Molecular Biology, Inc.