SOD1 mutations disrupt redox-sensitive Rac regulation of NADPH oxidase in a familial ALS model
Maged M. Harraz, … , Christian Schöneich, John F. Engelhardt
Maged M. Harraz, … , Christian Schöneich, John F. Engelhardt
Published January 24, 2008
Citation Information: J Clin Invest. 2008;118(2):659-670. https://doi.org/10.1172/JCI34060.
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Research Article

SOD1 mutations disrupt redox-sensitive Rac regulation of NADPH oxidase in a familial ALS model

Abstract

Neurodegeneration in familial amyotrophic lateral sclerosis (ALS) is associated with enhanced redox stress caused by dominant mutations in superoxide dismutase–1 (SOD1). SOD1 is a cytosolic enzyme that facilitates the conversion of superoxide (O2•–) to H2O2. Here we demonstrate that SOD1 is not just a catabolic enzyme, but can also directly regulate NADPH oxidase–dependent (Nox-dependent) O2•– production by binding Rac1 and inhibiting its GTPase activity. Oxidation of Rac1 by H2O2 uncoupled SOD1 binding in a reversible fashion, producing a self-regulating redox sensor for Nox-derived O2•– production. This process of redox-sensitive uncoupling of SOD1 from Rac1 was defective in SOD1 ALS mutants, leading to enhanced Rac1/Nox activation in transgenic mouse tissues and cell lines expressing ALS SOD1 mutants. Glial cell toxicity associated with expression of SOD1 mutants in culture was significantly attenuated by treatment with the Nox inhibitor apocynin. Treatment of ALS mice with apocynin also significantly increased their average life span. This redox sensor mechanism may explain the gain-of-function seen with certain SOD1 mutations associated with ALS and defines new therapeutic targets.

Authors

Maged M. Harraz, Jennifer J. Marden, Weihong Zhou, Yulong Zhang, Aislinn Williams, Victor S. Sharov, Kathryn Nelson, Meihui Luo, Henry Paulson, Christian Schöneich, John F. Engelhardt

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Figure 1

ALS-associated SOD1 mutations activate cellular Nox activity.

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ALS-associated SOD1 mutations activate cellular Nox activity. (A) Rate o...
(A) Rate of NADPH-dependent O2•– production by total endomembranes isolated from the brain, spinal cord, and liver of nontransgenic or transgenic mice overexpressing SOD1WT or SOD1G93A (mean ± SEM; n = 3 per group). (B) DHE fluorescent detection of O2•– in lumbar spinal cord sections from nontransgenic and transgenic mice overexpressing SOD1WT or SOD1G93A. DAPI staining shows cell nuclei in each section. (C) Rate of NADPH-dependent O2•– production in total endomembranes isolated from SH-SY (neuronal) or MO59J (glial) cells at 48 hours following infection with adenoviral vectors expressing LacZ, SOD1WT, SOD1L8Q, or SOD1G93A. (D) Cell death was quantified in SH-SY and MO59J cells using trypan-blue exclusion at 72 hours after infection with the indicated adenoviral vectors. (E) Using conditions specified in C and D, the rate of NADPH-dependent O2•– production and cell death was assessed in the presence or absence of the Nox inhibitor apocynin (100 μM). Values are mean ± SEM (n = 6 per group). (F) Assessment of GTP-bound Rac1 (activated form) in spinal cord lysates from 2 nontransgenic or 3 SOD1G93A transgenic mice (120 days old) and from MO59J cells overexpressing WT or mutant SOD1 proteins (at 36 hours after adenoviral infection). The 2 left lanes are controls for the Rac activation assay, in which nontransgenic spinal cord lysates were preincubated with the indicated non-hydrolysable guanine nucleotide analogs.