Mice of diverse genetic backgrounds may be classified as being either resistant or susceptible to infection with Mycobacteria. These phenotypes appear to be determined by a single gene on chromosome 1, the Bcg gene, and are expressed at the level of the macrophage in vitro. When compared to macrophages from mice of the susceptible phenotype (Bcgs), macrophages from mice of the resistant phenotype (Bcgr) show enhanced functional properties including increased expression of MHC class II molecules, increased nitric oxide production, and greater capacity to inhibit the growth of several intracellular pathogens. The bacteriostatic activity of B10R and B10S macrophages correlated with the amount of nitric oxide produced by the macrophages. Since protein kinase C (PKC) has been shown to be involved in the induction of a range of macrophage functional activities, experiments were conducted to examine the possibility that phenotypic differences between Bcgr and Bcgs macrophages may be related to differences in PKC-dependent signalling. Macrophage cell lines were derived from mice congenic at the Bcg locus that are either resistant (B10R) or susceptible (B10S) to infection with Mycobacteria. In the basal state, PKC-specific activity was significantly increased in the cytosolic fractions of B10R cells when compared to B10S cells. Following phorbol myristate acetate (PMA) treatment and following the stimulation with Mycobacteria bovis BCG, PKC-specific activity increased significantly in membrane fractions of both B10R and B10S cells, but the absolute level was significantly greater in particulate fractions from B10R macrophages. Furthermore, B10R cells had a superior ability to phosphorylate endogenous substrates compared to B10S macrophages. Scatchard analysis of phorbol ester receptors revealed no differences between B10R and B10S cells. In contrast, the sensitivity of partially purified PKC from B10S cells to activation in vitro by diacylglycerol was decreased by approximately 50 % when compared to enzyme from B10R cells. Western-blotting analysis using antibodies specific for PKC isoforms (α, β, δ, ε, ζ and η) showed similar levels of PKC isoforms present in B10R and B10S cells. To examine whether differences in PKC activity of B10R and B10S cells had functional consequences, the induction of c-fos gene expression was compared in the two cell lines. In response either to infection with M. bovis BCG or to stimulation with PMA, c-fos mRNA levels in B10R macrophages were increased 2−4-fold in comparison to B10S macrophages. Since we have previously found that the bacteriostatic activity of B10R and B10S macrophages correlated with the amount of nitric oxide produced by the macrophages, we have tested if the enhancement of PKC activity in these macrophages affects their ability to produce nitric oxide. We have found that interferon-γ-(IFNγ)-induced secretion of nitric oxide by B10R macrophages could be augmented a few fold by the activation of PKC whereas, in B10S macrophages stimulated with IFNγ, nitric oxide release could be augmented by only about 10−20 %. These results indicate that the differences in PKC activity between B10R and B10S macrophages may contribute to altered responsiveness to IFNγ that results in different production of effector molecules crucial for bacteriostatic activity against M. bovis BCG.