Inhibition of Proinflammatory RANTES Expression by TGF-β1 Is Mediated by Glycogen Synthase Kinase-3β-dependent β-Catenin Signaling

TGF-beta 1 is a pleiotropic cytokine with potent anti-inflammation property. However, the mechanisms underlying TGF-beta 1 suppression of inflammation remain largely unexplored. In this study, we demonstrated that TGF-beta 1 inhibited TNF-alpha-or IL-1-induced RANTES expression in human kidney tubular epithelial cells (HKC-8). To delineate the mechanism by which TGF-beta 1 inhibits RANTES expression, we examined the potential signal pathway activated by TGF-beta 1 in suppressing NF-kappa B signaling. TGF-beta 1 affected neither TNF-beta-induced I kappa B alpha phosphorylation and subsequent degradation, nor p65 NF-kappa B phosphorylation and its nuclear translocation. However, TGF-beta 1 could inhibit p65 and p50 binding to the kappa B site in human RANTES promoter as revealed by chromatin immunoprecipitation assay and protein-DNA binding assay. We found that TGF-beta 1 induced glycogen synthase kinase-3 beta (GSK-3 beta) phosphorylation on Ser-9 in HKC-8 cells, leading to its inactivation. Knockdown of GSK-3 beta mimicked TGF-beta 1 and inhibited RANTES induction, whereas overexpression of GSK-3 beta abolished the inhibitory effect of TGF-beta 1 and completely restored RANTES expression. Furthermore, TGF-beta 1 induced the dephosphorylation and activation of beta-catenin, a major downstream target of GSK-3 beta. Ectopic expression of constitutively active beta-catenin mimicked the TGF-beta 1 effect and completely suppressed RANTES expression induced by TNF-alpha. Interestingly, TGF-beta 1 induced a physical interaction between beta-catenin and p65 NF-kappa B, which prevented p65 binding to the kappa B site, sequestered its trans-activating activity, and repressed p65-mediated gene transcription. We conclude that TGF-beta 1 inhibition of proinflammatory RANTES expression is mediated by beta-catenin-triggered blockade of NF-kappa B signaling.