Mutant Copper-Zinc Superoxide Dismutase Binds to and Destabilizes Human Low Molecular Weight Neurofilament mRNA

Abstract

The mechanism by which mutated copper-zinc superoxide dismutase (SOD1) causes familial amyotrophic lateral sclerosis is believed to involve an adverse gain of function, independent of the physiological antioxidant enzymatic properties of SOD1. In this study, we have observed that mutant SOD1 (G41S, G85A, and G93A) but not the wild type significantly reduced the stability of the low molecular weight neurofilament mRNA in a dosage-dependent manner. We have also demonstrated that mutant SOD1 but not the wild type bound directly to the neurofilament mRNA 3-untranslated region and that the binding was necessary to induce mRNA destabilization. These observations provide an explanation for a novel gain of function in which mutant SOD1 expression in motor neurons alters an intermediate filament protein expression. Copper-zinc superoxide dismutase (SOD1)1 is a small and abundant intracellular metalloenzyme that catalyzes the conversion of the superoxide anion to hydrogen peroxide (1, 2). The demonstration of a genetic linkage between mutations in the SOD1 gene and familial amyotrophic lateral sclerosis aroused interest in the role of SOD1 in motor neuron death (3). The mechanisms by which mutations in the SOD1 gene contribute to the pathogenesis of ALS remain to be defined crisply. A gain of function to the SOD1 enzyme, conferred by the mutation, remains a potential mechanism for this toxicity. Supporting this concept are the observations that many SOD1 mutants maintain normal enzymatic function (4), that SOD1 knock-out mice do not develop an ALS-like phenotype (5), and that the over-expression of wild type (WT) SOD1 in mutant SOD1 transgenic mouse models does not modify the disease state (5). A mechanism reported recently (6) for the underlying mutant SOD1 toxicity proposes that the formation of the monomeric mutant SOD1 protein is the result of oxidative stress, which leads to microaggregate formation. It was also shown that SOD1 G93A but not WT co-localizes with neurofilamentous aggregates induced by low molecular weight neurofilament (NFL) overexpression in transfected Neuro2A cells (7). Neuropathological hallmarks of ALS include the degeneration of the descending corticospinal tracts and spinal, bulbar, and cortical motor neurons (8). In the latter nerves, degenerating neurons characteristically contain a variety of inclusions, including neurofilamentous aggregates. Such neurofilamentous aggregates have been observed in a number of transgenic mouse models of motor neuron degeneration in which the stoichiometry of expression of the individual neurofilament (NF) subunits has been altered. These include alterations in the level of expression of each of the neurofilament subunit proteins (NFL and middle and high molecular weight neurofilaments) (9 -11) and related intermediate filament proteins (12). It has been shown previously (13-15) by both in situ hybridization and single cell RT-PCR that the level of expression of NFL mRNA is reduced selectively in degenerating spinal motor neurons in ALS, suggesting that the alte...

Mutant Cu/Zn-Superoxide Dismutase Associated with Amyotrophic Lateral Sclerosis Destabilizes Vascular Endothelial Growth Factor mRNA and Downregulates Its Expression

“…The altered mobility shift and the presence of mutant SOD1 in the RNP complex indicate a change in either RBP-VEGF RNA interactions, protein-protein interactions, or both. Strong and colleagues have shown that mutant SOD1 can destabilize low-molecular-weight NFL (neurofilament) mRNA by binding directly to the 3Ј-UTR (Ge et al, 2005). We did not observe direct binding of mutant SOD1 to the VEGF 3Ј-UTR by UV cross-linking.…”

Aldolases A and C Are Ribonucleolytic Components of a Neuronal Complex That Regulates the Stability of the Light-Neurofilament mRNA

“…A defective neurocytoskeletal network, resulting from unproductive aldolase/NF-mRNA interactions could negatively affect the local concentrations of general translation factors or impair recycling of translational components (Decker and Parker, 1995;Mangus et al, 2003;Amrani et al, 2004). This, in turn, might have effects in trans with deleterious consequences for the normal expression of other gene products (Lin et al, 2004;Ge et al, 2005). The latter becomes critical for large neurons with extraordinary high metabolic demands at specific sites (Wu et al, 1997).…”

DNA cleavage mediated by copper superoxide dismutase via two pathways

“…The important substrates for the peroxidase activity include its end product hydrogen peroxide, the SOD protein itself and nucleic acids. However, the conclusion remains elusive as to the direct link between the aberrant properties and the injurious effect of Cu 2 Zn 2 SOD, implying the presence of other toxic pathways [25].…”