Endosomal Nox2 Facilitates Redox-Dependent Induction of NF-κB by TNF-α

Abstract

Growing evidence suggests that NADPH oxidase (Nox)-derived reactive oxygen species (ROS) play important roles in regulating cytokine signaling. We have explored how TNF-a induction of Nox-dependent ROS influences NF-kB activation. Cellular stimulation by TNF-a induced NADPH-dependent superoxide production in the endosomal compartment, and this ROS was required for IKK-mediated activation of NF-kB. Inhibiting endocytosis reduced the ability of TNF-a to induce both NADPH-dependent endosomal superoxide and NF-kB, supporting the notion that redox-dependent signaling of the receptor occurs in the endosome. Molecular analyses demonstrated that endosomal H 2 O 2 was critical for the recruitment of TRAF2 to the TNFR1=TRADD complex after endocytosis. Studies using both Nox2 siRNA and Nox2-knockout primary fibroblasts indicated that Nox2 was critical for TNF-a-mediated induction of endosomal superoxide. Redox-active endosomes that form after TNF-a or IL-1b induction recruit several common proteins (Rac1, Nox2, p67 phox , SOD1), while also retaining specificity for ligand-activated receptor effectors. Our studies suggest that TNF-a and IL-1b signaling pathways both can use Nox2 to facilitate redox activation of their respective receptors at the endosomal level by promoting the redox-dependent recruitment of TRAFs. These studies help to explain how cellular compartmentalization of redox signals can be used to direct receptor activation from the plasma membrane.

Redox Modifier Genes and Pathways in Amyotrophic Lateral Sclerosis

“…In this context, Rac1 appears to play a critical role, as it is required for bringing both IL-1R and Nox2 from the plasma membrane into the endosome in a mammary epithelial cell line (82). Similar processes likely control TNFR1 recruitment into redoxosomes in this cell type (83). In smooth muscle cells, Nox1 appears to be capable of replacing Nox2 function in redoxosomes following both IL-1b and TNFa stimulation (97), suggesting that Rac1-dependent NADPH oxidases (i.e., Nox1 and Nox2) perform cell type-specific redoxosomal signaling.…”

“…More recently, the redox-dependent processes that control NFkB activation by these cytokines have been shown to also involve NADPH oxidases (specifically Nox1 and Nox2). In this context, endosomal activation of Nox1 and=or Nox2 has been shown to control redox-dependent activation of both the IL-1b and TNFa receptors (82,83,97,100). Although these receptors have unique effector pathways responsible for activating NFkB, they share a similar dependence on redox-active signaling endosomes for their activation cascades.…”

“…Redoxosomes are signaling endosomes that generate O 2 À in their lumen in response to stimuli such IL-1b (82,97) and TNFa (83,97,138) (Fig. 2).…”

“…ROS generated by redoxosomes act as second messengers to facilitate the redox-dependent activation of the IL-1b receptor (IL-1R) and TNFa receptor (TNFR1). An important feature of redoxosomal biology is its dependence on endocytosis of the ligand-activated receptor, together with NADPH oxidases from the plasma membrane; inhibiting receptor endocytosis prevents the NADPHdependent ROS production in redoxosomes that is required for activation of the proinflammatory transcription factor NFkB (82,83). In this context, Rac1 appears to play a critical role, as it is required for bringing both IL-1R and Nox2 from the plasma membrane into the endosome in a mammary epithelial cell line (82).…”

Regulation of CuZnSOD and Its Redox Signaling Potential: Implications for Amyotrophic Lateral Sclerosis

“…A majority of inducible SOD1 expression is facilitated by the binding of nuclear factor kappa B (NFjB), nuclear factor (erythroid-derived 2)-like 2 (NRF2), aryl hydrocarbon receptor/aryl hydrocarbon receptor nuclear translocator (AHR/ ARNT), and CAATT enhancer binding proteins (C/EBP) transcription factors to enhancer regions of SOD1 (38,40,47). For example, NFjB binding the human SOD1 promoter increases after exposure to cytokines, oxidative stress, and hydrogen peroxide (H 2 O 2 ) (32,40). Likewise, cellular stress leads to the swift localization of NRF2 to the nucleus to increase the expression of genes such as SOD1.…”