Heparin is a potent inhibitor of arterial smooth muscle cell (SMC) migration and proliferation in vivo and in vitro. We propose that heparin affects these SMC functions by interfering with either the expression or the activity of secreted proteases required for cell movement. We have reported that heparin selectively inhibits the expression of tissue-type plasminogen activator in SMCs during mitogenesis. In this study we show that the gene expression of another kind of protease, interstitial collagenase, is induced by fetal bovine serum and is also suppressed by heparin. The inhibitory effect on the induced collagenase mRNA is specific to heparin-like molecules and does not depend on the anticoagulant activity of heparin. The induction of the collagenase gene depends on the protein kinase C pathway, since it can be induced by phorbol esters such as phorbol 12-myristate 13-acetate and blocked by inhibitors such as H-7 and staurosporine. In transient transfection assays with chloramphenicol acetyltransferase constructs containing the phorbol ester-responsive element introduced into baboon SMCs, heparin inhibits transcription induced by serum or phorbol 12-myristate 13-acetate. These results support the conclusion that, in primate SMCs, interstitial collagenase gene transcription mediated by the phorbol ester-responsive element is blocked by heparin.