Abstract
The production of cAMP from Bacillus anthracis edema toxin (ET) activates gene expression in macrophages through a complex array of signaling pathways, most of which remain poorly defined. In this study, the tumor suppressor protein adenomatous polyposis coli (APC) was found to be important for the up-regulation of previously defined ET-stimulated genes (Vegfa, Ptgs2, Arg2, Cxcl2, Sdc1, and Cebpb). A reduction in the expression of these genes after ET exposure was observed when APC was disrupted in macrophages using siRNA or in bone marrow-derived macrophages obtained from C57BL/6J-Apc(Min) mice, which are heterozygous for a truncated form of APC. In line with this observation, ET increased the expression of APC at the transcriptional level, leading to increased amounts of APC in the nucleus. The mechanism utilized by APC to increase ET-induced gene expression was determined to depend on the ability of APC to interact with C/EBP β, which is a transcription factor activated by cAMP. Coimmunoprecipitation experiments found that APC associated with C/EBP β and that levels of this complex increase after ET exposure. A further connection was uncovered when silencing APC was determined to reduce the ET-induced phosphorylation of C/EBP β at Thr-188. This ET-mediated phosphorylation of C/EBP β was blocked by glycogen synthase kinase 3 (GSK-3) inhibitors, suggesting that GSK-3 is involved in the activation of C/EBP β and supporting the idea of APC helping direct interactions between GSK-3 and C/EBP β. These results indicate that ET stimulates gene expression by promoting the formation of an inducible protein complex consisting of APC and C/EBP β.