Tumorigenic Cell Lines Type IV Collagenase and Interstitial Collagenase in Human

The regulation of M, 72,000 type IV collagenase and interstitial collagenase expression was studied in vitro. Three tumorigenic human cell lines were used, together with human fetal lung fibroblasts as a nontumorigenic control. M, 72,000 type IV collagenase was expressed constitutively by all four cell lines, whereas only A20S8 melanoma cells exhibited constitutive expression of interstitial collagenase. Treatment of cells with transforming growth factor 01 (TGF-/31) and 12-O-tetradecanoylphorbol-13-acetate (TPA) revealed an opposite pattern of regulation of these two metalloproteinases. Specifically, TPA increased interstitial collagenase mRNA levels in each cell line and decreased type IV colla genase mRNA levels in control fibroblasts and the tumorigenic cell lines, HT-1080 and A2058. TGF-/J1 treatment increased type IV collagenase mRNA levels in each cell line and decreased interstitial collagenase mRNA levels in A2058 melanoma cells. Interstitial collagenase mRNA induction was accompanied in all cell lines by elevated interstitial procollagenase in the conditioned medium, as detected by zymography. Changes in \1, 72,000 type IV collagenase expression revealed a more complex pattern of regulation. TPA and TGF-jSl treatment of HT-1080 cells resulted in the appearance of two bands of gelatinolytic activity with a molecular weight of approximately 62,000 and 59,000. The M, 62,000 species was also induced by TGF-01 treatment of A2058 cells. Addition of affinity-purified radiolabeled M, 72,000 type IV procollagenase to TPA-treated HT-1080 cells demon strated that both species were products of the \1, 72,000 proenzyme and that exogenous proenzyme could be processed by these cells. Western blot analysis with specific antipeptide antibodies revealed that both the M, 62,000 and 59,000 species were derived from the M, 72,000 proenzyme by ammo-terminal cleavage. There was no evidence for cellular processing of either interstitial procollagenase or the M, 92,000 type IV procollagenase. These results demonstrate that the M, 72,000 type IV collagenase is under the control of different regulatory elements from interstitial collagenase, at the level of both mRNA expression and cellular processing, and that this processing appears to be the result of a phorbol ester and TGF-01-inducible cellular activation mechanism. The ratio of active enzyme species to latent V/, 72,000 proenzyme may provide a better correlation with invasive potential than overall levels of this widely expressed metalloproteinase.