Biosynthesis and regulation of type V collagen in diploid human fibroblasts.
The biosynthesis of type V collagen and its regulation were studied using diploid human gingival fibroblasts. Cells were metabolically labeled with radioactive amino acids and labeled proteins were subjected to limited pepsin digestion, DEAE-cellulose chromatography at 15 degrees C, and polyacrylamide gel electrophoresis. Proteins eluted from DEAE-cellulose columns by 0.25 M NaCl contained a collagen species which was resistant to mammalian collagenase and had alpha chains with hydroxylysine/lysine ratios and CNBr peptide patterns similar to alpha 1(V) and alpha 2(V). Procollagen(V) fractions obtained by DEAE-cellulose chromatography and immunoprecipitates of type V collagen antibody contained polypeptides with Mr = 239,000, 219,000, 198,000, 174,000, 157,000, and 132,000. By comparing the CNBr peptide maps of these proteins with those of standard alpha 1(V) and alpha 2(V) chains, the first three polypeptides were shown to be related to alpha 1(V) and the others to alpha 2(V). It was concluded that the gingival fibroblasts synthesize type V collagen, that the pro alpha 1(V) and the pro alpha 2(V) chains have Mr = 239,000 and 174,000, respectively, and that the alpha 1(V) and alpha 2(V) chains laid in the form of fibrils have Mr = 198,000 and 132,000, respectively. A detectable amount of type V collagen was synthesized only at high cell density, and it was associated with the cell layer. The amount and proportion of type V synthesized were increased when the cells were labeled in the presence of serum, and the increase was accompanied by a decrease in type III. This effect was dependent on serum concentration. Serum obtained from platelet-poor plasma failed to elicit this effect, and it was restored by the addition of platelet-derived growth factor. Platelet-derived growth factor was effective in medium with and without platelet-poor serum. Thus, it appears that platelet-derived growth factor may be an important regulatory factor in the synthesis of types V and III collagens.
D000596:Amino Acids / Q000032:analysis; D002478:Cells, Cultured; D003094:Collagen / Q000235:genetics* / Q000302:isolation & purification; D004171:Diploidy; D005347:Fibroblasts / Q000378:metabolism; D005881:Gingiva / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D003012:Microbial Collagenase; D010446:Peptide Fragments / Q000032:analysis; D011347:Procollagen / Q000235:genetics / Q000302:isolation & purification
D000596:Amino Acids; D010446:Peptide Fragments; D011347:Procollagen; D003094:Collagen; D003012:Microbial Collagenase