DIFFERENTIAL-EFFECTS OF PROTEIN-KINASE-C INHIBITORS ON FIBRONECTIN-INDUCED INTERLEUKIN-BETA GENE-TRANSCRIPTION, PROTEIN-SYNTHESIS AND SECRETION IN HUMAN MONOCYTIC CELLS

Human monocytic cells express interleukin-1 beta (IL-1 beta) when stim ulated with the extracellular matrix glycoprotein, fibronectin (FN). P rotein kinase C (PKC) activation is considered important for this proc ess; however, the metabolic steps at which PKC acts upon to mediate th e FN-induced IL-1 beta response remain unclear. We performed an analys is of the mechanisms by which two PKC inhibitors, Calphostin C and Sta urosporine, prevent the FN-induced IL-I beta response. Both inhibitors blocked the secretion of IL-I beta protein into the media of peripher al blood mononuclear cells exposed to FN. Immunoprecipitation analysis revealed that under these circumstances, Calphostin C inhibited the p roduction of IL-I beta protein, whereas Staurosporine allowed protein production, but inhibited its secretion. To determine the mechanisms r esponsible for these differences, we turned to human U937 promonocytic cells. U937 cells transfected with the human full-length IL-1 beta pr omoter connected to a luciferase reporter gene were submitted to trans cription assays, Northern blotting, and DNA electrophoresis mobility g el shift assays. These studies revealed that Calphostin C inhibited th e nuclear translocation of the transcription factor activator protein- 1 (AP-I) which is considered necessary for FN induction of IL-1 beta g ene transcription, and prevented the transcription of the IL-1 beta ge ne. In contrast, Staurosporine alone induced AP-I translocation and st imulation of the gene. Overall, our data indicate that Calphostin C pr events the transcription of the IL-I beta gene thereby inhibiting prot ein synthesis. Based on the high specificity of this compound for PKC, we conclude that PKC is necessary for FN-induced IL-1 beta protein pr oduction. In contrast, Staurosporine prevented secretion of IL-I beta by unknown mechanisms.