Role of Protein Kinase C in Basal and Hydrogen Peroxide-Stimulated NF-κB Activation in the Murine Macrophage J774A.1 Cell Line

In macrophages, hydrogen peroxide appears to be a physiological activator of the transcription factor, nuclear factor kappa B (NF-κB); however, the molecular basis of H2O2-stimulated NF-κB activation is not well defined. The observations that NF-κB can be activated in cells by phorbol 12-myristate 13-acetate andin vitroby addition of protein kinase C (PKC) are suggestive of a role of PKC in NF-κB activation, which was investigated in the J774A.1 murine macrophage cell line. Basal NF-κB DNA-binding activity and nuclear localization were decreased by PKC inhibitors. Although PKC activity was modified by H2O2with a similar time course as H2O2activation of NF-κB, the H2O2-stimulated increase in NF-κB DNA binding and translocation to the nucleus was unaffected by PKC inhibitors. Furthermore, PKC down-regulation (through preincubation with phorbol esters) also affected only baseline NF-κB DNA binding but not H2O2-stimulated NF-κB activation. Buffering of changes in intracellular free calcium concentration also had no effect upon H2O2-stimulated NF-κB activation. Thus, classical PKC activity may modulate basal NF-κB activity but does not participate in H2O2-stimulated NF-κB activation.