Cystic fibrosis gene and protein expression during fetal lung development

The cystic fibrosis transmembrane conductance regulator (CFTR) transports Cl- in the apical membrane of secretory epithelial cells. Normal lung development is critically dependent on active Cl- secretion by developing airway epithelia. In this study, the polymerase chain reaction (PCR) was used to detect CFTR mRNA expression in rabbit lung at different stages of development. Using nested amplification in exons 7 and 11, a 494-bp fragment was isolated from cDNA prepared from lungs obtained at gestational days 22 (pseudoglandular) and 29 (terminal sac). Nucleotide sequencing verified the identification of rabbit CFTR sequences. At day 22 of gestation, CFTR was detected in fetal liver, kidney, heart, intestine, and brain. CFTR could not be amplified from adult rabbit brain. CFTR protein was detected using a polyclonal antibody raised in chickens against a synthetic peptide from the R domain conjugated to thyroglobulin. Rabbit CFTR migrated as a 165-kD protein on 5% sodium dodecyl sulfate polyacrylamide gels when detected either by chicken anti-R IgG or a previously characterized rabbit anti-R antibody. In frozen sections of fetal rabbit lung at day 22 of gestation, CFTR was localized to the membrane regions of primitive tubular epithelial cells and absent from surrounding connective tissue. At day 29, CFTR was concentrated in the apical region of bronchiolar epithelial cells and absent from alveoli and vessels. These results suggest that CFTR gene expression begins very early in lung development and, with differentiation of the airways, becomes confined to differentiated bronchiolar epithelium, the presumed site for cAMP-mediated Cl- secretion.