Colchicine-induced modulation of collagenase in human skin fibroblast cultures. I. Stimulation of enzyme synthesis in normal cells.
Microtubule-active agents affect the secretion of a variety of proteins, including collagenase. To gain insight into the mechanisms involved in this process, we examined the effects of colchicine on the synthesis, secretion, and activity of human skin collagenase. When added to monolayer cultures of human skin fibroblasts, 10(-6) M colchicine produced a mean 3-fold increase in trypsin-activatable collagenase in the culture medium. Stimulation was not observed with lumicolchicine. The enhanced accumulation of collagenase was dose-dependent with 10(-9), 10(-8), 10(-7), and 10(-6) M colchicine giving collagenase activities/mg protein that were 100 +/- 6%, 165 +/- 20%, 186 +/- 34%, and 297 +/- 62% of control, respectively. Although the effect on collagenase was seen under conditions independent of cellular growth (i.e., in serum-free medium), maximum stimulation occurred in subconfluent cultures. The colchicine-induced increase in activity was paralleled by an increase in immunoreactive enzyme protein, suggesting stimulation of enzyme synthesis. The catalytic efficiency of the enzyme (activity per unit immunoreactive protein) was unchanged, however, indicating that a structurally normal enzyme was being synthesized. To examine the process in more detail, the biosynthesis of 3H-labeled collagenase was quantitated in these cultures by specific immunoprecipitation. Although 10(-6) M colchicine produced no increase in total protein synthesis, an increased rate of collagenase synthesis was seen after only 1.5 hr. These data suggest that colchicine has a specific effect on the synthesis of collagenase and may be a useful probe for studying its regulation.
D002478:Cells, Cultured; D003078:Colchicine / Q000494:pharmacology*; D004591:Electrophoresis, Polyacrylamide Gel; D004790:Enzyme Induction; D005347:Fibroblasts / Q000201:enzymology; D006801:Humans; D003012:Microbial Collagenase / Q000096:biosynthesis*; D008970:Molecular Weight; D012867:Skin / Q000187:drug effects / Q000201:enzymology*
D003012:Microbial Collagenase; D003078:Colchicine