Cystic fibrosis mutations lead to carboxyl-terminal fragments that highlight an early biogenesis step of the cystic fibrosis transmembrane conductance regulator

Inefficient delivery of the cystic fibrosis transmembrane conductance regul ator (CFTR) to the surface of cells contributes to disease in the majority of cystic fibrosis patients. Analysis of cystic fibrosis-associated missens e mutations in the first nucleotide binding domain (NBD1), including A455E, S549R, Y563N, and P574H, revealed reduced levels of mature CFTR with eleva ted levels of carboxyl-terminal polypeptide fragments of 105 and 90 kDa, Th ese fragments appear early in biogenesis and degrade rapidly in four distin ct cell types tested including the bronchial epithelial IB3-1 cell Line. Th ey were detected at highest levels with CFTRA455E where the 105-kDa fragmen t accounted for 40% of newly synthesized polypeptide but for only 20 and 7% of nascent wild type and mutant Delta F508 proteins, respectively. The ban ds represent core- and unglycosylated forms of the same CFTR fragment suppo rting that precursor forms are correctly inserted into the membrane of the endoplasmic reticulum, Proteolytic cleavage would be predicted to occur on the cytosolic face of the endoplasmic reticulum within the NBD1-R domain se gment, but pharmacological testing did not support involvement of the 26 S proteasome. The examined missense mutations in NBD1 manifest differently th an the major mutant, Delta F508, and highlight a critical conformational as pect of biogenesis of CFTR.