AP-1 activity is negatively regulated by cannabinol through inhibition of its protein components, c-fos and c-jun

Abstract

Regulation of the activator protein-1 (AP-1) complex is very intricate because it involves phosphorylation state, protein-protein, and protein-DNA interactions. In these studies, the regulation of AP-1 activity, with emphasis on c-fos and c-jun regulation, was investigated using cannabinol (CBN) in primary mouse splenocytes in vitro. Cannabinoid compounds exhibit immunosuppressive actions that are putatively mediated through Gi-protein coupled receptors that negatively regulate adenylate cyclase. However, recent studies suggest that cannabinoids modulate other signaling cascades. Indeed, we demonstrate that CBN inhibited binding to AP-1-containing sites from the interleukin-2 promoter. This inhibition of binding was, in part, due to decreased nuclear expression of c-fos and c-jun. We further determined that the effects of CBN were due to posttranslational modifications of these phosphoproteins and showed that CBN inhibited the activation of ERK MAP kinases. Thus, cannabinoidinduced immunosuppression involves disruption of the ERK signaling cascade. J. Leukoc. Biol. 67: 259-266; 2000.

Evidence for Cannabinoid Receptor-Dependent and -Independent Mechanisms of Action in Leukocytes

“…It has also been reported that cannabinol exhibited a 10-fold higher affinity for the CB2 receptor, the predominant cannabinoid receptor expressed in the immune system (Bouaboula et al, 1993;Galiegue et al, 1995;Schatz et al, 1997). Indeed, cannabinol exhibits immunomodulating effects (Condie et al, 1996;Herring et al, 1998;Faubert and Kaminski, 2000;Yea et al, 2000), but, as demonstrated here, so did cannabidiol and WIN 55212-3, both of which exhibit low binding affinity for CB1 and CB2.…”

“…1, interleukin-2 was readily induced by PMA/Io treatment and was inhibited by cannabinol (1-20 M) in a concentration-dependent manner. These studies were conducted with micromolar concentrations of cannabinoid compounds because it has been previously demonstrated that these concentrations are required in vitro due to both the lipophilicity of the cannabinoids and the serum concentration dependence for many of the cannabinoid effects (Faubert and Kaminski, 2000). Pretreatment of splenocytes with a combination of antagonists (0.5 M/0.5 M, 2.5 M/2.5 M, and 5 M/5 M SR144528/SR141716A) did not significantly attenuate the cannabinol-mediated inhibition of interleukin-2 production at any cannabinol concentration tested.…”

“…With the demonstration that CB1 and CB2 were not involved in cannabinoid-induced inhibition of PMA/Io-stimulated interleukin-2, other potential T cell targets were examined. A major target of cannabinoids in T cells is nuclear factor of activated T cells (Faubert and Kaminski, 2000;Yea et al, 2000), a critical transcription factor in the regulation of interleukin-2 that is tightly controlled by changes in [Ca 2ϩ ] i (reviewed in (Cantrell, 1996). Therefore, generation of the calcium signal seemed a likely point of inhibition by cannabinoid compounds.…”

“…Using an optimum stimulatory concentration of PMA/ Io, cannabinol treatment inhibited interleukin-2 protein secretion by splenic T cells (Condie et al, 1996). This inhibition correlated with a decrease in expression of phosphorylated extracellular signal-regulated kinase mitogen-activated protein kinase (Faubert and Kaminski, 2000). Furthermore, it was determined that cannabinol inhibited transcription factor binding to the proximal activator protein-1 and distal nuclear factor of activated T cells motifs in the interleukin-2 promoter (Condie et al, 1996;Faubert and Kaminski, 2000;Yea et al, 2000), which might also contribute to the inhibition of interleukin-2.…”

“…This inhibition correlated with a decrease in expression of phosphorylated extracellular signal-regulated kinase mitogen-activated protein kinase (Faubert and Kaminski, 2000). Furthermore, it was determined that cannabinol inhibited transcription factor binding to the proximal activator protein-1 and distal nuclear factor of activated T cells motifs in the interleukin-2 promoter (Condie et al, 1996;Faubert and Kaminski, 2000;Yea et al, 2000), which might also contribute to the inhibition of interleukin-2. Thus, although the mechanism of cannabinoid-induced inhibition of interleukin-2 in PMA/Io-stimulated splenocyte is understood to occur via inhibition of transcriptional activation of interleukin-2 (Yea et al, 2000), the upstream signals contributing to inhibition of transcription have not been completely elucidated.…”