Negative regulation of NADPH oxidase-4 by hydrogen peroxide clone-5

Hydrogen peroxide-inducible clone-5 (Hic5) is a focal adhesion adaptor protein induced by the pro-fibrotic cytokine, transforming growth factor-β1 (TGF-β1). We have previously demonstrated that TGF-β1 induces myofibroblast differentiation and lung fibrosis by activation of the reactive oxygen species (ROS)-generating enzyme, NADPH oxidase 4 (Nox4). Here, we investigated a potential role for Hic-5 in regulating Nox4, myofibroblast differentiation and senescence. In normal human diploid fibroblasts, TGF-β1 induces Hic-5 expression in a delayed manner relative to induction of Nox4 and myofibroblast differentiation. Hic-5 silencing induced constitutive Nox4 expression and enhanced TGF-β1-inducible Nox4 levels. Induction of constitutive Nox4 protein in Hic-5 silenced cells was independent of transcription and translation, and controlled by the ubiquitinproteasomal system (UPS). Hic-5 associates with the ubiquitin ligase, Cbl-c, and the ubiquitin-binding protein, heat shock protein 27 (HSP27). The interaction of these proteins is required for the ubiquitination of Nox4 and for maintaining low basal levels of this ROSgenerating enzyme. Our model suggests that TGF-β1-induced Hic-5 functions as a negative feedback mechanism to limit myofibroblast differentiation and senescence by promoting UPS-mediated degradation of Nox4. Together, these studies indicate that endogenous Hic-5 suppresses senescence and pro-fibrotic activities of myofibroblasts by down-regulating Nox4 protein expression. Additionally, these are the first studies, to our http://www.jbc.org/cgi/doi/10.1074/jbc.M114.562249 The latest version is at JBC Papers in Press. Published on May 15, 2014 as Manuscript M114.562249 Copyright 2014 by The American Society for Biochemistry and Molecular Biology, Inc. by gest on N ovem er 7, 2017 hp://w w w .jb.org/ D ow nladed from