Regulation of transcription factor AP‐1 in human fibroblasts in response to cytokines: role in regulation of Matrix Metalloproteinase‐3

Abstract

Previously we showed that IL‐4 inhibits the IL‐1 induction of MMP‐3, and that there was decreased binding in gelshift assays to a probe containing the MMP‐3 AP‐1 site in the presence of IL‐4. In the present study, the AP‐1 Family TransAm Transcription Factor Assay Kit from Active Motif, as well as chromatin immunoprecipitation studies, are utilized to determine the composition of AP‐1 dimers a)available in fibroblasts and b)actually binding to the endogenous MMP‐3 promoter in the presence of IL‐1 and/or IL‐4. Results show that FosB, Fra‐1, c‐Jun and JunB are all present and capable of binding in fibroblasts stimulated with IL‐1, and that their binding is inhibited by IL‐4. JunD binding is present as well, but is relatively unaffected by these cytokines. Interestingly, there was no increase in cFos binding in response to IL‐1 at the 1 or 3 hour timepoints. Preliminary ChIP experiments using an antibody specific for the active form of c‐Jun, show binding to the endogenous MMP‐3 promoter induced by IL‐1 and inhibited by IL‐4, and this binding correlates with levels of acetylated histone H3 binding the endogenous promoter. Further ChIP experiments will be done with other members of the AP‐1 family. In addition, the signal transduction pathways leading to activation of AP‐1 binding will also be examined. This work is supported by grant R15DE16277 from the NIDCR to R.C. Borghaei.