Journal article
Abnormal localization of cystic fibrosis transmembrane conductance regulator in primary cultures of cystic fibrosis airway epithelia
The Journal of cell biology, Vol.118(3), pp.551-559
08/01/1992
DOI: 10.1083/jcb.118.3.551
PMCID: PMC2289545
PMID: 1379244
Abstract
Cystic fibrosis (CF) is caused by mutations in the cystic fibrosis transmembrane conductance regulator (CFTR), a membrane glycoprotein that forms Cl- channels. Previous work has shown that when some CF-associated mutants of CFTR are expressed in heterologous cells, their glycosylation is incomplete. That observation led to the hypothesis that such mutants are not delivered to the plasma membrane where they can mediate Cl- transport. Testing this hypothesis requires localization of CFTR in nonrecombinant cells and a specific determination of whether CFTR is in the apical membrane of normal and CF epithelia. To test the hypothesis, we used primary cultures of airway epithelia grown on permeable supports because they polarize and express the CF defect in apical Cl- permeability. Moreover, their dysfunction contributes to disease. We developed a semiquantitative assay, using nonpermeabilized epithelia, an antibody directed against an extracellular epitope of CFTR, and large (1 microns) fluorescent beads which bound to secondary antibodies. We observed specific binding to airway epithelia from non-CF subjects, indicating that CFTR is located in the apical membrane. In contrast, there was no specific binding to the apical membrane of CF airway epithelia. These data were supported by qualitative studies using confocal microscopy: the most prominent immunostaining was in the apical region of non-CF cells and in cytoplasmic regions of CF cells. The results indicate that CFTR is either missing from the apical membrane of these CF cells or it is present at a much reduced level. The data support the proposed defective delivery of some CF-associated mutants to the plasma membrane and explain the lack of apical Cl- permeability in most CF airway epithelia.
Details
- Title: Subtitle
- Abnormal localization of cystic fibrosis transmembrane conductance regulator in primary cultures of cystic fibrosis airway epithelia
- Creators
- G M Denning - Howard Hughes Medical Institute, Department of Internal Medicine, University of Iowa College of Medicine, Iowa City 52242L S Ostedgaard - Howard Hughes Medical Institute, Department of Internal Medicine, University of Iowa College of Medicine, Iowa City 52242M J Welsh - Howard Hughes Medical Institute, Department of Internal Medicine, University of Iowa College of Medicine, Iowa City 52242
- Resource Type
- Journal article
- Publication Details
- The Journal of cell biology, Vol.118(3), pp.551-559
- DOI
- 10.1083/jcb.118.3.551
- PMID
- 1379244
- PMCID
- PMC2289545
- ISSN
- 0021-9525
- eISSN
- 1540-8140
- Language
- English
- Date published
- 08/01/1992
- Academic Unit
- Neurology; Molecular Physiology and Biophysics; Pulmonary, Critical Care, and Occupational Medicine; Emergency Medicine; Fraternal Order of Eagles Diabetes Research Center; Neurosurgery; Internal Medicine
- Record Identifier
- 9984021000302771
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