QUANTITATION OF MUCIN MESSENGER-RNA IN RESPIRATORY AND INTESTINAL EPITHELIAL-CELLS

Mucin glycoproteins (mucins) are the major macromolecular constituents of mucus gels in mammalian respiratory, gastrointestinal, and reprodu ctive tracts. Disorders of mucin glycosylation, which may result from either abnormal post-translational processing or differences in mucin protein gene expression, have been indicated in several diseases. Quan titation of mucin gene expression has been hindered by two features of human mucin genes: variable numbers of tandemly repeating nucleotides per mRNA molecule and polydisperse mRNA transcripts. We report here a method to quantitate mucin mRNA levels in epithelial cells and have e valuated three mucin genes, MUC1, MUC2, and MUC5, which are expressed in respiratory epithelium. The method uses the 3' non-tandem repeat mu cin cDNA sequences, as they were shown to have a single-size transcrip t when amplified by the polymerase chain reaction, consistent with a o ne-to-one relationship with the mRNA molecule. The 3' non-tandem repea t cDNA sequences were cloned and transcribed in vitro to prepare compl ementary RNA (cRNA) standards. By comparison to a cRNA standard curve, mucin gene expression was evaluated in colon adenocarcinoma, pancreat ic adenocarcinoma, and transformed respiratory epithelial cells and in nasal polyp tissue by slot blot analysis. CFPAC-1, a pancreatic adeno carcinoma cell line, expressed the highest MUC1 transcript levels. Col on adenocarcinoma cell lines varied in MUC2 expression levels, and one colon adenocarcinoma cell line, HT-29, had higher levels of MUC5 than MUC2. Nasal polyp tissue expressed more MUC5 mRNA than MUC1 or MUC2 m RNA. This mucin mRNA slot blot method provides a quantitative method f or investigating the regulation of mucin gene expression in health and disease.