Early steps in the biosynthesis of MUC2 epithelial mucin in colon cancer cells.
Expression of the MUC2 mucin has been demonstrated in normal gastrointestinal and respiratory epithelium and in carcinomas of the gastrointestinal and respiratory tracts, breast, ovary, and bladder using RNA probes and (or) monoclonal antibodies reactive with peptide epitopes on the 23 amino acid tandem repeat. Mouse monoclonal antibodies 4F1 and 3A2 were previously obtained by immunization with mucin derived from the LS174T colon cancer cell line and a KLH conjugate of a synthetic MUC2 VNTR peptide. These antibodies react with distinct epitopes on synthetic VNTR peptides and with normal and malignant epithelial tissues. In the present study, we examined the biosynthesis of MUC2 in LS174T colon cancer cells, using these antibodies to immunoprecipitate labelled mucin. A very high molecular mass protein was immunoprecipitated following 1 min pulse labelling with [3H]threonine and [3H]proline. A slight increase in molecular mass was observed over the next 16 min; however, unlike the MUC1 mucin, there was no large difference in apparent molecular mass between the MUC2 protein precursor and fully processed mucin using separation by SDS-PAGE. O-Glycosylation began within 1 h of synthesis of the protein core. Mucin secretion into the culture medium was detected in the 2nd hour following synthesis and was largely completed within 4 h of synthesis. Secreted mucin was far less reactive with these monoclonal antibodies than the precursor protein.
McGuckin
M A
MA
Department of Obstetrics and Gynaecology, University of Queensland, Royal Brisbane Hospital, Herston, Australia.
Devine
P L
PL
Ward
B G
BG
D000230:Adenocarcinoma / Q000378:metabolism*; D000911:Antibodies, Monoclonal; D015153:Blotting, Western; D003110:Colonic Neoplasms / Q000378:metabolism*; D017077:Culture Media, Conditioned; D006031:Glycosylation; D006801:Humans; D055262:Mucin-2; D009077:Mucins / Q000096:biosynthesis*; D011233:Precipitin Tests; D014407:Tumor Cells, Cultured
D000911:Antibodies, Monoclonal; D017077:Culture Media, Conditioned; C524625:MUC2 protein, human; C524626:Muc2 protein, mouse; D055262:Mucin-2; D009077:Mucins