Biosynthesis of two distinct types of mucin in HM3 human colon cancer cells.
Mucins, high-M(r) glycoproteins with a large amount of O-glycosidically linked carbohydrate, protect the colonic epithelial surface and are altered in ulcerative colitis and colon cancer. At least two mucin genes, MUC2 and MUC3, are expressed at high levels in the human intestine. As an experimental model for studying the biosynthesis of human intestinal mucins, we used HM3 colon cancer cells. When mature mucins labelled with [3H]glucosamine or [3H]threonine were analysed by gel filtration, it was found that secreted mucins (M(r) > 10(8) were larger than soluble cellular mucins (M(r) approx. 5 x 10(6)). Only secreted mucin was sensitive to reduction. Both MUC2 and MUC3 proteins, identified by labelling with [3H]threonine or [35S]cysteine and immunoprecipitation with antibodies to synthetic mucin peptides, were already of large size (M(r) > 180,000) by the earliest labelling time (5 min). The MUC3 precursor was completely degraded by trypsin, but the MUC2 precursor had a trypsin-resistant fragment of M(r) approx. 240,000 containing threonine and cysteine. The trypsin-resistant MUC2 fragment contained N-linked carbohydrate, as indicated by a decrease in size as a result of peptidyl N-glycosidase digestion or tunicamycin treatment of HM3 cells. These results show that HM3 colon cancer cells produce at least two distinct human intestinal mucins. They also indicate that the mechanisms of biosynthesis of intestinal mucins differ from those of other mucin-like glycoproteins that have been studied.
Ohara
S
S
Gastrointestinal Research Laboratory, VA Medical Center, San Francisco, CA.
Byrd
J C
JC
Gum
J R
JR
Kim
Y S
YS
D000595:Amino Acid Sequence; D003110:Colonic Neoplasms / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D008969:Molecular Sequence Data; D009077:Mucins / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics; D009844:Oligosaccharides / Q000032:analysis; D014407:Tumor Cells, Cultured
D009077:Mucins; D009844:Oligosaccharides