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256 Amyotrophic Lateral Sclerosis

deficient in SOD1 have been reported to be resistant to acetaminophen toxicity (Lei et al., 2006). Even though SOD1 as a cytosolic antioxidant protects against mitochondrial dysfunction in a mouse model of transient focal cerebral ischemia (Fujimura et al., 2000), SOD1 deficiency, rather than overexpression, is associated with enhanced recovery and

This apparent discrepancy concerning the role of SOD1 in cellular injury can be explained by the model introduced, showing that increased SOD1 activity in the IMS paradoxically produces peroxides which are converted to highly toxic ROS. This view is further supported by an observation in mouse model of genetic disorder ataxia-telangiectasia, where elevated levels of SOD1 exacerbate the phenotype of neurodegeneration (Peter et al., 2001). It is also of interest that SOD1 overexpression and high tissue dismutase activity may potentiate atherogenesis in fat-fed atherosclerosis-susceptible mice (Tribble et al., 1997). The evidence about deleterious role of increased SOD1 expression has been most recently complemented by studies demonstrating that overexpression of SOD1 in retina leads to increased

The key component for the SOD1-derived hydroperoxide toxicity in IMS is cytochrome c. Previous studies, including electron paramagnetic resonance (EPR) studies (Barr et al., 1996; Svistunenko, 2005; Belikova et al., 2006; Basova et al., 2007) have demonstrated that the reaction of cytochrome c with hydroperoxide results in formation of oxoferryl cytochrome c (peroxidase compound I-type intermediate) and corresponding protein-derived tyrosyl radical, which is highly reactive and has a potential to oxidize proteins, DNA, and lipids, as well as endogenous antioxidants such as glutathione, NADH, and ascorbate (Lawrence et al., 2003) (Fig. 3). In particular, oxidation of cardiolipin, a phospholipid which is in complex with cytochrome c on the surface of the inner mitochondrial membrane, causes the release of proapoptotic factors from mitochondria (Kagan et al., 2005; Belikova et al., 2006). This leads to a scenario where the hydroperoxide produced by increased SOD1 activity in the IMS, would thus serve as a substrate for cardiolipin-bound cytochrome c and consequently switch on very early proapoptotic processes, inducing consecutive programmed cell death. Additionally, upon increased hydroperoxide levels cytochrome c peroxidase activity may cause NADH oxidation producing a radical, which in turn donates an electron to oxygen

The toxicity based on the dismutase activity of mutant SOD1 in the IMS might also be true even for dismutase inactive mutant SOD1 proteins. In human FALS SOD1 mutations are dominantly inherited resulting in the presence of both wild type and mutant SOD1 subunits in each cell. Thus, dismutase activity lacking G85R-SOD1 can form active heterodimers with wt SOD1 molecules. In mice the co-expression of human mutant and wt SOD1 accelerated disease (Jaarsma et al., 2000; Fukada et al., 2001; Deng et al., 2006). Importantly, unaffected A4V-SOD1 mutant mice developed the disease only when mated with human wt SOD1 overexpressing mice (Deng et al., 2006). It was also shown recently that the toxicity of mutant SOD1 dimers is not correlated with their capacity to form protein aggregates but

Until now, several pathological mechanisms have been demonstrated how mutant SOD1 induces mitochondrial dysfunction in FALS models. Among them, the emerging evidence indicates that the SOD1-dependant hydroperoxide production in mitochondrial IMS may

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oxidative metabolism in exercising muscle from ALS patients. *J Neurol Sci*, Vol.191,

evidence for altered calcium in motor nerve terminals in amyotropic lateral

(2007). Overexpression of CCS in G93A-SOD1 mice leads to accelerated neurological deficits with severe mitochondrial pathology. *Proc Natl Acad Sci U S A*,

Lepock, J.R. & Meiering, E.M. (2003). Cu/Zn superoxide dismutase mutants associated with amyotrophic lateral sclerosis show enhanced formation of aggregates in vitro. *Proc Natl Acad Sci U S A*, Vol.100, No.12, (June 2003), pp. 7021- 7026.


**11** 

**Folding and Aggregation of** 

Elizabeth M. Meiering

*University of Waterloo* 

*Canada* 

**Cu, Zn-Superoxide Dismutase** 

Helen R. Broom, Heather A. Primmer, Jessica A.O. Rumfeldt, Peter B. Stathopulos, Kenrick A. Vassall, Young-Mi Hwang and

*Guelph-Waterloo Centre for Graduate Work in Chemistry and Biochemistry,* 

In 1993, a genetic link was established between amyotrophic lateral sclerosis (ALS) and mutant forms of Cu,Zn superoxide dismutase (SOD1) (Deng et al. 1993; Rosen et al. 1993), an antioxidant enzyme that catalyzes the dismutation of the damaging free radical superoxide anion (O2-) to hydrogen peroxide (H2O2) and diatomic oxygen (O2) via cyclic reduction and oxidation of a protein-bound Cu ion (Valentine et al. 2005). Today, over 150, predominantly missense mutations have been identified at ~75 sites spread throughout the protein (http://alsod.iop.kcl.ac.uk/). SOD1 mutations are found in ~15- 20% of inherited or familial ALS (fALS) cases and in a small percentage of sporadic ALS (sALS) cases (Rosen et al. 1993; Kato et al. 2000; Liu et al. 2009; Forsberg et al. 2011). fALS accounts for ~10% of all ALS cases and so SOD1 mutations comprise ~1.5-2% of all ALS cases, but nevertheless represent a major known cause of the disease. The clinical symptoms of fALS and sALS are similar, yet fALS patients with SOD1 mutations have an earlier age of disease onset than sALS (by ~10 years) (Wijesekera and Leigh 2009). Furthermore, while the age of disease onset has not been identified as statistically different between different SOD1 mutations, disease duration for each mutation is often different, ranging from shorter (e.g. ~1 year for A4V, the most common mutation in North America) than the typical 3-5 years to longer (e.g. ~18 years for H46R) (Cudkowicz et al. 1997; Valentine et al. 2005; Wang et al. 2008). In humans and murine models of ALS, mutations in the gene encoding SOD1 are typically autosomal dominant and are associated with a toxic gain of function. Despite extensive research, the molecular basis for mutant SOD1 toxicity remains unclear (Valentine et al. 2005; Boillee et al. 2006; Ilieva et al. 2009). Extensive research has been conducted on SOD1-linked fALS, as understanding and treatment of this disease may be relevant to ALS in general. While ALS patients share many clinical symptoms, numerous genes have been linked to ALS, and there is evidence for differences in pathology related to both genetic and environmental factors; hence, ALS is a syndrome and not a single disease with unique

**1. Introduction 1.1 ALS and SOD1** 

pathology (Cozzolino et al. 2008).

