The ΔF508 CFTR defect: molecular mechanism of suppressor mutation V510D and the contribution of transmembrane helix unraveling

• Published in: bioRxiv
• Main Authors: Simon, Kailene S, Nagarajan, Karthi, Mechin, Ingrid, Duffy, Caroline, Manavalan, Partha, Altmann, Steve, Majewski, Aliza, Foley, Joseph, Kaczmarek, J Stefan, Bercury, Scott, Maderia, Matthew, Hilbert, Brendan J, Batchelor, Joseph D, Ziegler, Robin, Bajko, Jeffrey, Kothe, Michael, Scheule, Ronald K, Nair, Anil, Hurlbut, Gregory D
• Format: Paper
• Language: English
• Published: Cold Spring Harbor Cold Spring Harbor Laboratory Press 20.04.2020
• Subjects: Cell surface; Channel gating; Cystic fibrosis; Epithelial cells; Gene deletion; Genetic suppression; Homology; Mutation; Phenylalanine; Protein structure; Tertiary structure; Transmembrane domains
• DOI: 10.1101/2020.04.19.049338

Cystic fibrosis (CF) results from mutations within the gene encoding the Cystic Fibrosis Transmembrane Conductance Regulator (CFTR), a transmembrane chloride channel found on the apical surface of epithelial cells. The most common CF-causing mutation results in a deletion of phenylalanine 508 (ΔF508-CFTR), a residue normally found within the NBD1 domain. Loss of F508 causes NBD1 to be less thermodynamically stable and prevents proper tertiary folding of CFTR. As a result, CFTR is not properly trafficked to the cell surface. Recently, progress has been made towards the development of small molecule correctors that can restore CFTR tertiary structure and stabilize the channel to overcome the instability inherent in ΔF508-CFTR. However, the resultant improvement in channel activity has been modest, and the need for potent correctors remains. To fully inform such efforts, a better understanding of the molecular pathology associated with ΔF508-CFTR is required. Here we present a comprehensive study of the impact of F508 deletion on both purified NBD1 and full-length CFTR. Through the use of homology modeling, molecular dynamics simulations, mutational analysis, biochemical, biophysical and functional characterization studies, we obtained insight into how the ΔF508 mutation may lead to helical unraveling of transmembrane domains 10 and 11 (TM10, TM11), and how the known suppressor mutations V510D and R1070W, as well as novel second site suppressor mutations (SSSMs) identified in this work, may act to rescue ΔF508-CFTR maturation and trafficking.