Keloids are benign collagenous tumors that develop during an abnormal wound healing process. The genetic predisposition to form keloids is found predominantly in black and oriental populations. Our studies suggest that keloids result when an alternate regulatory program is invoked in dermal fibroblasts during wound healing. Fibroblasts from keloid tissue are refractory to down-regulation of matrix synthesis and growth in response to several effectors. Hydrocortisone coordinately down-regulates mRNA levels of alpha1(I), alpha2(I), alpha1(III) and alpha2(V) collagen genes in fibroblasts from normal human dermis and scars, but not in fibroblasts from keloids. The abnormal mechanism affects transcription rate. A simple hypothesis consistent with these findings is that a common trans-acting factor involved in coordinate regulation of the several collagen genes is altered in keloid cells, an that the cis-sequences (response elements) are similar in different collagen genes and in the two cell types. We have found differences in the binding of nuclear proteins from normal and keloid cells to synthetic oligonucleotides containing the phorbol ester, c-AMP, and glucocorticoid response elements which suggest that an alteration in the AP-1 family of transcription factors may account for abnormal regulation of growth and matrix synthesis in keloids. Using gel mobility shift assays, Western analysis and immunoprecipitation of protein-DNA complexes we will characterize trans- and cis-acting elements involved in abnormal regulation of gene expression in keloid cells. We will examine regulation of these transcription factors by hydrocortisone, phorbol ester and c-AMP. We will attempt to show a direct connection between this alteration in transcription factors and abnormal glucocorticoid down-regulation of collagen genes using mobility shift assays to show altered binding of transcription factors to cis- elements in the context of collagen genes and altered glucocorticoid regulation of appropriate reporter plasmids in transient transfections. We will attempt to confirm results of preliminary studies suggesting that abnormal glucocorticoid regulation in keloids also involves stability of collagen mRNA. If so, RNase protection assays of transcripts from the 3' and 5' end of collagen cDNA's will be used to examine binding of cytoplasmic factors from normal and keloid cells and to identify responsible sequences. A proliferin promoter-reporter construct will be used to help elucidate the mechanism whereby the same cis-acting sequence can be up- or down- regulated by glucocorticoids depending on AP-1 activity.