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Chaperone displacement from mutant cystic fibrosis transmembrane conductance regulator restores its function in human airway epithelia

Fei Sun^*, Zhibao Mi, Steven B. Condliffe^*,1, Carol A. Bertrand^*, Xiaoyan Gong^*,1, Xiaoli Lu, Ruilin Zhang^*, Joseph D. Latoche, Joseph M. Pilewski^*,, Paul D. Robbins and Raymond A. Frizzell^*,2

^* Department of Cell Biology and Physiology,

Department of Microbiology and Molecular Genetics, and

Department of Medicine, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania, USA

² Correspondence: Department of Cell Biology and Physiology, University of Pittsburgh School of Medicine, Pittsburgh, PA 15261, USA. E-mail: frizzell{at}pitt.edu

Mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene cause cystic fibrosis (CF). The most common mutation, F508, omits the phenylalanine residue at position 508 in the first nucleotide binding domain (NBD1) of CFTR. The mutant protein is retained in the endoplasmic reticulum and degraded by the ubiquitin-proteasome system. We demonstrate that expression of NBD1 plus the regulatory domain (RD) of F508 CFTR (FRD) restores the biogenesis of mature F508 CFTR protein. In addition, FRD elicited a cAMP-stimulated anion conductance response in primary human bronchial epithelial (HBE) cells isolated from homozygous F508 CF patients. A protein transduction domain (PTD) could efficiently transduce (90%) airway epithelial cells. When fused to a PTD, direct addition of the FRD peptide conferred a dose-dependent, cAMP-stimulated anion efflux to F508 HBE cells. Hsp70 and Hsp90 associated equally with WT and F508 CFTR, whereas nearly twice as much of the Hsp90 cochaperone, Aha1, associated with F508 CFTR. Expression of FRD produced a dose-dependent removal of Aha1 from F508 CFTR that correlated with its functional rescue. These findings indicate that disruption of the excessive association of the cochaperone, Aha1, with F508 CFTR is associated with the correction of its maturation, trafficking and regulated anion channel activity in human airway epithelial cells. Thus, PTD-mediated FRD fragment delivery may provide a therapy for CF.—Sun, F., Mi, Z., Condliffe, S. B., Bertrand, C. A., Gong, X., Lu, X., Zhang, R., Latoche, J. D., Pilewski, J. M., Robbins, P. D., Frizzell, R. A. Chaperone displacement from mutant cystic fibrosis transmembrane conductance regulator restores its function in human airway epithelia.

Key Words: CFTR • Aha1 • rescue • PTD