L-TRYPTOPHAN INDUCES EXPRESSION OF COLLAGENASE GENE IN HUMAN FIBROBLASTS - DEMONSTRATION OF ENHANCED AP-1 BINDING AND AP-1 BINDING SITE-DRIVEN PROMOTER ACTIVITY

Collagenase, a prototypic matrix metalloproteinase, plays a major role in the degradation of the extracellular matrix. The essential amino a cid L-tryptophan was recently shown to stimulate the expression of col lagenase gene in human dermal fibroblast cultures. In this study, we f ocused our attention on the mechanisms responsible for activation of c ollagenase transcription by L-tryptophan. Incubation of fibroblasts wi th L-tryptophan resulted in a dose- and time-dependent elevation of co llagenase and tissue inhibitor of metalloproteinase mRNA levels. The m aximum enhancement in collagenae mRNA was approximately 50-fold. This effect was not abolished by cycloheximide, suggesting independence fro m ongoing protein synthesis. Transient cell transfections with a promo ter/reporter gene construct containing 3.8 kb of 5' flanking DNA of th e human collagenase gene linked to the chloramphenicol acetyl transfer ase (CAT) gene or a construct containing three phorbol ester-responsiv e AP-1 binding sequences (12-O-tetradecanoyl-phorbol-13-acetate-respon sive element) in front of the thymidine kinase promoter linked to the CAT gene indicated enhancement of promoter activity by L-tryptophan. F urthermore, electrophoretic DNA mobility shift assays demonstrated enh anced DNA-protein complex formation specific for an AP-1 binding site probe with nuclear extracts prepared from cells incubated with L-trypt ophan. These results collectively suggest that activation of collagena se gene expression in dermal fibroblasts by L-tryptophan is mediated t hrough AP-1 binding elements in the collagenase gene promoter that are sufficient for gene response.