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The human MUC3 gene is highly expressed in small intestine and gallbladder. Thus far only 646 basepairs of its cDNA encoding 17 amino acid repeats have been cloned. In order to further clone the human MUC3 cDNA, a human small intestinal cDNA library was constructed and screened with a cDNA probe encompassing the 17 amino acid tandem repeat region of human MUC3. In two subsequent screenings of the library resulting positive clones were used as probes. In total, 27 partial MUC3 cDNA clones were isolated and sequenced that define a semi-unique region and a novel 177 nucleotide tandem repeat region, located upstream of the region encoding the 17 amino acid tandem repeats. The 177 nucleotide repeat region is at least 5 kb in length and encodes 59 amino acid repetitive peptides with a consensus sequence of VSTTPVASSEASTLSTTPVDTSTPVTTSTQASSSPTTAEGTSMPTSTPSEGSTPLTSMP, that is notably different from the 17 amino acid repeat of MUC3 or any other known mucin repeat.

Mucin expression was studied during proliferation and differentiation of the enterocyte-like Caco-2 and goblet cell-like LS174T cell lines. Caco-2 cells express mRNAs of MUC1, MUC3, MUC4 and MUC5A/C whereas MUC2 and MUC6 mRNAs are virtually absent. Furthermore, MUC3 mRNA is expressed in a differentiation dependent manner, as is the case for enterocytes. Concomitantly MUC3 protein precursor (approximately 550 kDa) was detected in Caco-2 cells. In LS174T cells mucin mRNAs of MUC1, MUC2 and MUC6 are constitutively expressed at high levels, whereas MUC3, MUC4 and MUC5A/C mRNAs are present at low levels. At the protein level LS174T cells express the goblet cell specific mucin protein precursors MUC2, MUC5A/C and MUC6 with apparent molecular masses of about 600 kDa, 470/500 kDa and 400 kDa respectively. MUC3 protein is not detectable. Furthermore, human gallbladder mucin protein (approximately 470 kDa precursor), of which the gene has not yet been identified, is expressed in LS174T cells. In addition, synthesis and secretion of the goblet cell specific mature MUC2, MUC5A/C and human gallbladder mucin was demonstrated in LS174T cells. It is concluded that Caco-2 and LS174T cell lines provide excellent in vitro models to elucidate the cell-type specific mechanisms responsible for mucin expression.