Interleukin-6 regulation of transforming growth factor (TGF)-β receptor compartmentalization and turnover enhances TGF-β1 signaling

Abstract

Transforming growth factor (TGF)-β1 is a key cytokine involved in the pathogenesis of fibrosis in many organs, whereas interleukin (IL)-6 plays an important role in the regulation of inflammation. Recent reports demonstrate interaction between the two cytokines in disease states. We have assessed the effect of IL-6 on TGF-β1 signaling and defined the mechanism by which this occurred. Stimulation of Smad-responsive promoter (SBE)4-Lux activity by TGF-β1 was significantly greater in the presence of IL-6 than that induced by TGF-β1 alone. Augmented TGF-β1 signaling following the addition of IL-6 appeared to be mediated through binding to the cognate IL-6 receptor, the presence of which was confirmed by fluorescence-activated cell sorting and Stat-specific signaling. TGF-β1 receptors internalize by both caveolin-1 (Cav-1) lipid raft and early endosome antigen 1 (EEA-1) non-lipid raft pathways, with non-lipid raft-associated internalization increasing TGF-β1 signaling. Affinity labeling of TGF-β1 receptors demonstrated that IL-6 stimulation resulted in increased partitioning of TGF-β receptors to the non-lipid raft fraction. There was no change in expression of Cav-1; however, following IL-6 stimulation, co-immunoprecipitation demonstrated decreased association of IL-6 receptor with Cav-1. Increased TGF-β1-dependent Smad signaling by IL-6 was significantly attenuated by inhibition of clathrin-mediated endocytosis and augmented by depletion of membrane cholesterol. These results indicate that IL-6 increased trafficking of TGF-β1 receptors to non-lipid raft-associated pools results in augmented TGF-β1 Smad signaling. © 2005 by The American Society for Biochemistry and Molecular Biology, Inc.