Abstract
Reactive oxygen species (ROS) convey signals essential for proliferation, maintenance, and senescence of a growing list of cell types. Compartmentalization of these signals is integral to cell viability as well as the signaling pathways ROS direct. Redox-active endosomes (redoxosomes) are formed downstream of several ligand-activated receptors. NADPH oxidase (NOX) is a main component of redoxosomes, which recruits multiple proteins (Rac1, NOX2, p67phox, SOD1). Isolation of redoxosomes and evaluation of how superoxide (O2˙−) production directs receptor signaling at the level of the endosome have enabled a better understanding of biologic processes controlled by ROS. In this chapter, we will first review the major signaling pathways that utilize redoxosomes and components that control its redox-dependent functions. We will then outline biochemical and biophysical methods for the isolation and characterization of redoxosome properties.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Oakley FD, Abbott D, Li Q, Engelhardt JF (2009) Signaling components of redox active endosomes: the redoxosomes. Antioxid Redox Signal 11(6):1313–1333. https://doi.org/10.1089/ARS.2008.2363
Klomsiri C, Rogers LC, Soito L, McCauley AK, King SB, Nelson KJ, Poole LB, Daniel LW (2014) Endosomal H2O2 production leads to localized cysteine sulfenic acid formation on proteins during lysophosphatidic acid-mediated cell signaling. Free Radic Biol Med 71:49–60. https://doi.org/10.1016/j.freeradbiomed.2014.03.017
Li Q, Harraz MM, Zhou W, Zhang LN, Ding W, Zhang Y, Eggleston T, Yeaman C, Banfi B, Engelhardt JF (2006) Nox2 and Rac1 regulate H2O2-dependent recruitment of TRAF6 to endosomal interleukin-1 receptor complexes. Mol Cell Biol 26(1):140–154. https://doi.org/10.1128/MCB.26.1.140-154.2006
Li Q, Spencer NY, Oakley FD, Buettner GR, Engelhardt JF (2009) Endosomal Nox2 facilitates redox-dependent induction of NF-kappaB by TNF-alpha. Antioxid Redox Signal 11(6):1249–1263. https://doi.org/10.1089/ARS.2008.2407
Oakley FD, Smith RL, Engelhardt JF (2009) Lipid rafts and caveolin-1 coordinate interleukin-1beta (IL-1beta)-dependent activation of NFkappaB by controlling endocytosis of Nox2 and IL-1beta receptor 1 from the plasma membrane. J Biol Chem 284(48):33,255–33,264. https://doi.org/10.1074/jbc.M109.042127
Miller FJ Jr, Filali M, Huss GJ, Stanic B, Chamseddine A, Barna TJ, Lamb FS (2007) Cytokine activation of nuclear factor kappa B in vascular smooth muscle cells requires signaling endosomes containing Nox1 and ClC-3. Circ Res 101(7):663–671. https://doi.org/10.1161/CIRCRESAHA.107.151076
Li Q, Zhang Y, Marden JJ, Banfi B, Engelhardt JF (2008) Endosomal NADPH oxidase regulates c-Src activation following hypoxia/reoxygenation injury. Biochem J 411(3):531–541. https://doi.org/10.1042/BJ20071534
Banfi B, Clark RA, Steger K, Krause KH (2003) Two novel proteins activate superoxide generation by the NADPH oxidase NOX1. J Biol Chem 278(6):3510–3513. https://doi.org/10.1074/jbc.C200613200
Lambeth JD (2004) NOX enzymes and the biology of reactive oxygen. Nat Rev Immunol 4(3):181–189. https://doi.org/10.1038/nri1312
Mumbengegwi DR, Li Q, Li C, Bear CE, Engelhardt JF (2008) Evidence for a superoxide permeability pathway in endosomal membranes. Mol Cell Biol 28(11):3700–3712. https://doi.org/10.1128/MCB.02038-07
Lynch RE, Fridovich I (1978) Permeation of the erythrocyte stroma by superoxide radical. J Biol Chem 253(13):4697–4699
Salvador A, Sousa J, Pinto RE (2001) Hydroperoxyl, superoxide and pH gradients in the mitochondrial matrix: a theoretical assessment. Free Radic Biol Med 31(10):1208–1215
Harraz MM, Marden JJ, Zhou W, Zhang Y, Williams A, Sharov VS, Nelson K, Luo M, Paulson H, Schoneich C, Engelhardt JF (2008) SOD1 mutations disrupt redox-sensitive Rac regulation of NADPH oxidase in a familial ALS model. J Clin Invest 118(2):659–670. https://doi.org/10.1172/JCI34060
Mizuno T, Kaibuchi K, Ando S, Musha T, Hiraoka K, Takaishi K, Asada M, Nunoi H, Matsuda I, Takai Y (1992) Regulation of the superoxide-generating NADPH oxidase by a small GTP-binding protein and its stimulatory and inhibitory GDP/GTP exchange proteins. J Biol Chem 267(15):10,215–10,218
Kopani M, Celec P, Danisovic L, Michalka P, Biro C (2006) Oxidative stress and electron spin resonance. Clin Chim Acta 364(1-2):61–66. https://doi.org/10.1016/j.cca.2005.05.016
Acknowledgments
This work was supported by NIH grant R24 DK096518 (to J.F.E.).
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2019 Springer Science+Business Media, LLC, part of Springer Nature
About this protocol
Cite this protocol
Shahin, W.S., Engelhardt, J.F. (2019). Isolation of Redox-Active Endosomes (Redoxosomes) and Assessment of NOX Activity. In: Knaus, U., Leto, T. (eds) NADPH Oxidases. Methods in Molecular Biology, vol 1982. Humana, New York, NY. https://doi.org/10.1007/978-1-4939-9424-3_27
Download citation
DOI: https://doi.org/10.1007/978-1-4939-9424-3_27
Published:
Publisher Name: Humana, New York, NY
Print ISBN: 978-1-4939-9423-6
Online ISBN: 978-1-4939-9424-3
eBook Packages: Springer Protocols