Cl transport in complemented CF bronchial epithelial cells correlates with CFTR mRNA expression levels

Abstract

Little is known about the relationship between CF transmembrane conductance regulator (CFTR) gene expression and the corresponding transport of Cl. The phenotypic characteristics of polarized DeltaF508 homozygote CF bronchial epithelial (CFBE41o-) cells were evaluated following transfection with episomal expression vector containing either full-length (6.2kb) wild type (wt) and (4.7kb) DeltaF508CFTR cDNA. Forskolin-stimulated Cl secretion in two clones expressing the full-length wild type CFTR was assessed; clone c7-6.2wt gave 13.4+/-2.5 microA/cm(2) and clone c10-6.2wt showed 41.3+/-25.3 microA/cm(2). Another clone (c4-4.7DeltaF) complemented with the DeltaF508 CFTR cDNA showed high and stable expression of vector-derived DeltaF508 CFTR mRNA and a small cAMP-stimulated Cl current (4.7+/-0.7 microA/cm(2)) indicating DeltaF508CFTR trafficking to the plasma membrane at physiological temperatures. Vector-driven CFTR mRNA levels were 5-fold (c7-6.2wt), 14-fold (c10-6.2wt), and 27-fold (c7-4.7DeltaF) higher than observed in normal bronchial epithelial cells (16HBE14o-) endogenously expressing wtCFTR. Assessment of CFTR mRNA levels and CFTR function showed that cAMP-stimulated CFTR Cl currents were 33%, 167% and 24%, respectively, of those in 16HBE14o- cells. The data suggest that transgene expression needs to be significantly higher than endogenously expressed CFTR to restore functional wtCFTR Cl transport to levels sufficient to reverse CF pathology.