NF-κB- and C/EBPβ-driven interleukin-1β gene expression and PAK1-mediated caspase-1 activation play essential roles in interleukin-1β release from Helicobacter pylori lipopolysaccharide-stimulated macrophages

Abstract

Helicobacter pylori is a Gram-negative microaerophilic bacterium that causes chronic gastritis, peptic ulcer, and gastric carcinoma. Interleukin-1β (IL-1β) is one of the potent proinflammatory cytokines elicited by H. pylori infection. We have evaluated the role of H. pylori lipopolysaccharide (LPS) as one of the mediators of IL-1β release and dissected the signaling pathways leading to LPS-induced IL-1β secretion. We demonstrate that both the NF-κB and the C/EBPβ-binding elements of the IL-1β promoter drive LPS-induced IL-1β gene expression. NF-κB activation requires the classical TLR4-initiated signaling cascade leading to IκB phosphorylation as well as PI-3K/Rac1/p21-activated kinase (PAK) 1 signaling, whereas C/EBPβ activation requires PI-3K/Akt/p38 mitogen-activated protein (MAP) kinase signaling. We observed a direct interaction between activated p38 MAP kinase and C/EBPβ, suggesting that p38 MAPK is the immediate upstream kinase responsible for activating C/EBPβ. Most important, we observed a role of Rac1/PAK1 signaling in activation of caspase-1, which is necessary for maturation of pro-IL-1β. H. pylori LPS induced direct interaction between PAK1 and caspase-1, which was inhibited in cells transfected with dominant-negative Rac1. PAK1 immunoprecipitated from lysates of H. pylori LPS-challenged cells was able to phosphorylate recombinant caspase-1, but not its S376A mutant. LPS-induced caspase-1 activation was abrogated in cells transfected with caspase-1(S376A). Taken together, these results suggested a role of PAK1-induced phosphorylation of caspase-1 at Ser³⁷⁶ in activation of caspase-1. To the best of our knowledge our studies show for the first time that LPS-induced Rac1/PAK1 signaling leading to caspase-1 phosphorylation is crucial for caspase-1 activation. These studies also provide detailed insight into the regulation of IL-1β gene expression by H. pylori LPS and are particularly important in the light of the observations that IL-1β gene polymorphisms are associated with increased risk of H. pylori-associated gastric cancer.