Published ahead of print on January 27, 2005, doi:10.1165/rcmb.2004-0306OC Am. J. Respir. Cell Mol. Biol., Volume 32, Number 5, May 2005, 453-461 A more recent version of this article appeared on May 1, 2005
Submitted on September 30, 2004 Mucin glycosylation and sulphation in airway epithelial cells is not influenced by CFTR expressionShih-Hsing Leir1,1 Department of Paediatric Molecular Genetics, University of Oxford, Weatherall Institute of Molecular Medicine, Oxford, United Kingdom, 2 Department of Biological Sciences, Imperial College, London, United Kingdom * To whom correspondence should be addressed. E-mail: ann.harris{at}paediatrics.ox.ac.uk.
Abnormalities in mucus properties and clearance make a major contribution to the pathology of Cystic Fibrosis (CF). Our aim was to test the hypothesis that the defects in CF mucus are a direct result of mutations in the CFTR protein. We evaluated a single mucin molecule MUC1F/5ACTR that carries tandem repeat sequence from MUC5AC, a major secreted airway mucin, in a MUC1 mucin vector. To establish whether the presence of mutant or normal CFTR directly influences the O-glycosylation and sulphation of mucins in airway epithelial cells, we used the CFT1-LC3 (
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F508 CFTR mutant) and CFT1-LCFSN (wild-type CFTR corrected) human airway epithelial cell lines. MUC1F/5ACTR mucin was immunoprecipitated, centricon purified, and O-glycosylation was evaluated by Matrix-assisted laser desorption ionization (MALDI) and electrospray (ES) tandem mass spectrometry to determine the composition of different carbohydrate structures. Mass spectrometry data showed the same O-glycans in both CFTR mutant and wild-type CFTR corrected cells. Metabolic labelling assays were performed to evaluate gross glycosylation and sulphation of the mucins and showed no significant difference in mucin synthesized in six independent clones of these cell lines. Our results show that the absence of functional CFTR protein causes neither an abnormality in mucin O-glycosylation nor an increase in mucin sulphation.
