**8. Conclusion**

306 Amyotrophic Lateral Sclerosis

al., 2003), which may be components of aggregates in mitochondria (Deng et al., 2006; Ferri et al., 2006) and harmful to the mitochondrial function. However, intermolecular disulfide bonds mediated at free cysteines (Cys6 and Cys111) can also be components of the detergent-insoluble SOD1 aggregates (Banci et al., 2007; Niwa et al., 2007). In either case, apo SOD1 is more prone to the disulfide-linked oxidative aggregation than holo SOD1 (Banci et al., 2007; Furukawa & O'Halloran, 2005). That is in concert with the notion that immature SOD1 is the pathogenic species in familial ALS (Seetharaman et al.,

It is still controversial whether the cysteine-mediated misfolding or aggregation of mutant SOD1 is the origin of the protein's toxicity. Removal of free cysteines, especially of Cys111, strongly reduced the ability of mutant SOD1 to form disulfide crosslinking and aggregates, and improved cell viability in cultured cells (Cozzolino et al., 2008; Niwa et al., 2007). Moreover, glutaredoxins, which specifically catalyze the reduction of protein-SSG-mixed disulfides, significantly increased the solubility of mutant SOD1 and protected neuronal cells (Cozzolino et al., 2008; Ferri et al., 2010). On the other hand, the intermolecular disulfide binding at cysteines is shown to have a limited effect on the aggregation of mutant SOD1 (Karch & Borchelt, 2008). Even in this case, Cys111-modified mutant SOD1 may cause neuronal toxicity independently of the aggregation, by oxidative stress such as thiol oxidase

In sporadic ALS, there had been no direct evidence that SOD1 was involved in the pathogenesis of the disease, except that some mutations of *SOD1* gene expressed familial ALS in a low penetration rate with seemingly 'sporadic' cases. The link between SOD1 and sporadic ALS was first introduced by the detection of SOD1 specifically modifiable with a chemical compound commonly in familial and sporadic ALS, although the molecular basis for it has not been determined in detail (Gruzman et al., 2007). It indicates that a similar conformational change in mutant and wild type SOD1 can trigger the phenotype of familial and sporadic ALS in common. In *in vitro* study, wild type SOD1 acquires toxic properties of mutant SOD1 through oxidation by hydrogen peroxide (Ezzi et al., 2007), implying that cysteine-oxidized wild type SOD1 may be a contributor to motor neuronal death in sporadic

Recently, a conformation-specific antibody generated against misfolded mutant SOD1 has been shown to recognize wild type SOD1 only when the protein was sulfonylated (-SO3H) at Cys111, and the antibody immunostained motor neurons in the spinal cords of sporadic ALS patients, but not of SOD1-unrelated familial ALS patients or controls (Bosco et al., 2010). Chemically oxidized or purified wild type SOD1 from sporadic ALS spinal cords inhibited kinesin-based fast axonal transport as did mutant SOD1, supposing that Cys111 mediated conformational change or misfolding of SOD1 is a shared pathological denominator of familial and sporadic ALS. Interestingly, most of the sporadic ALS-derived toxic SOD1 was soluble and non-aggregated, meaning that misfolding or monomerization is sufficient for SOD1 to gain the toxic property such as oxidative insult we have shown in mutant and wild type SOD1 (Kishigami et al., 2010). Further studies *in vivo* will be required to clarify the detailed mechanism of SOD1 toxicity mediated by oxidation of cysteine

2009).

ALS.

residues including Cys111.

activity we have shown (Kishigami et al., 2010).

**7. Role of cysteine-oxidized wild type SOD1 in sporadic ALS** 

The findings mentioned above indicate that oxidative modification of SOD1 at cysteine residues is a critical factor to contribute to the oxidative stress, inclusion pathology and degeneration of motor neurons commonly to familial and sporadic ALS. Based on them, steric inhibition of cysteine oxidation, monomerization or exposure of the dimer interface can be the first-line treatment strategy of this incurable disease.
