Biosynthesis of two subunits of type IV procollagen and of other basement membrane proteins by a human tumor cell line.
The major collagenous component secreted into the medium of cultured HT-1080 tumor cells was identified as type IV procollagen by specific antibodies and characteristic ratios of incorporated labeled 3-hydroxyproline and 4-hydroxyproline. The disulfide-bonded molecules consisted of two subunits, pro-alpha 1(IV) and pro-alpha 2(IV) chains with apparent molecular weights of 180 000 and 165 000. No conversion of the procollagen to collagen or to procollagen intermediates was detected in the cell cultures. The two subunits apparently represent different gene products, since enzymatic digestion of the separated chains produced quite different peptide maps. Pepsin degraded native type IV procollagen successively into several fragments, some still disulfide-linked, giving rise to a complex set of polypeptide chains (Mr = 30 000-140 000). This agrees with similar diverse patterns produced by pepsin from authentic type IV collagens. The ratio between the pro-alpha 1(IV) and pro-alpha 2(IV) chains varied in several experiments between 1.3 and 1.8, suggesting that the two chains belong to different triple-helical molecules. The cells also produced distinct amounts of fibronectin (subunit Mr = 230 000) and of the basement membrane glycoprotein laminin. The latter showed three subunits with Mr = 220 000, 210 000, and 400 000. A further disulfide-bonded, non-collagenous polypeptide (Mr = 160 000) was detected but not yet identified. Immunofluorescence demonstrated these proteins within the cells but not in a pericellular matrix. The production of basement membrane components by HT-1080 cells and lack of interstitial collagens disagree with the original classification of the cell line as a fibrosarcoma.
D000596:Amino Acids / Q000032:analysis; D000954:Antigens, Surface / Q000032:analysis; D001485:Basement Membrane / Q000378:metabolism*; D002460:Cell Line; D005455:Fluorescent Antibody Technique; D006801:Humans; D008565:Membrane Proteins / Q000096:biosynthesis*; D003012:Microbial Collagenase; D010434:Pepsin A; D010446:Peptide Fragments / Q000032:analysis; D011347:Procollagen / Q000096:biosynthesis*; D012509:Sarcoma
D000596:Amino Acids; D000954:Antigens, Surface; D008565:Membrane Proteins; D010446:Peptide Fragments; D011347:Procollagen; D010434:Pepsin A; D003012:Microbial Collagenase