Subunit-destabilizing mutations in Drosophila copper/zinc superoxide dismutase: neuropathology and a model of dimer dysequilibrium.

Abstract

Mutations in Cu/Zn superoxide dismutase (SOD), a hallmark of familial amyotrophic lateral sclerosis (FALS) in humans, are shown here to confer striking neuropathology in Drosophila. Heterozygotes with one wild-type and one deleted SOD allele retain the expected 50Y% of normal activity for this dimeric enzyme. However, heterozygotes with one wild-type and one missense SOD allele show lesser SOD activities, ranging from 37% for a heterozygote carrying a missense mutation predicted from structural models to destabilize the dimer interface, to an average of 13% for several heterozygotes carrying missense mutations predicted to destabilize the subunit fold. Genetic and biochemical evidence suggests a model of dimer dysequilibrium whereby SOD activity in missense heterozygotes is reduced through entrapment of wild-type subunits into unstable or enzymatically inactive heterodimers. This dramatic impairment of the activity ofwild-type subunits in vivo has implications for our understanding of FALS and for possible therapeutic strategies.Mutations in Cu/Zn superoxide dismutase (SOD, EC 1.15.1.1) have been identified in the etiology of familial amyotrophic lateral sclerosis (FALS), a syndrome commonly known as Lou Gehrig's disease (1, 2). Some FALS-affected individuals are carriers of missense mutations that appear to alter the stability (3) and/or the function of this critical oxygen radicalmetabolizing enzyme (2). The dominant phenotype associated with FALS mutations in SOD could result from loss of enzyme function, consistent with the reduced SOD activity in FALS patients (2, 4-7), or from the gain of a deleterious function, such as enhanced reactivity with peroxynitrite leading to elevated protein-tyrosine nitration (8, 9), consistent with the motor neuron damage found in transgenic mice overexpressing FALS-type mutant SOD (10, 11).A direct test of the relationship between SOD subunit function and neurodegenerative disease would be to induce and select mutations in SOD in an organism amenable to genetic analysis and then characterize the biochemical and neurological consequences conferred by these mutations. We have previously shown that null mutation for SOD in Drosophila melanogaster confers a syndrome including reduced adult lifespan, infertility, toxic hypersensitivity to a variety of oxygen stress conditions, and lethality in combination with mutations conferring defects in other oxygen radical scavengers (12-16). Overt symptoms of the SOD-null syndrome have been described only in homozygotes and are ameliorated by P element-vectored SOD transgenes (ref. 17; T. Parkes, A.J.H., and J.P.P., unpublished data).Here we show that mutations in SOD in Drosophila, as in humans, cause neuropathology. We also show that missense mutations expected to destabilize SOD subunits and dimer assembly significantly impair the activity of normal subunits in heterozygotes for a wild-type allele, resulting in significantly lower enzyme activity than expected in heterozygotes. These findings are important to our understanding of...

Phenotypic effects of familial amyotrophic lateral sclerosis mutant Sod alleles in transgenic Drosophila

“…The relationships among SOD activity, oxidative stress, and longevity have been studied extensively in the fruit fly, Drosophila melanogaster (10)(11)(12), but only one study examined the effects of a human FALS Sod mutant allele in Drosophila (13). Expression of this allele led to extension of life span, which paralleled the results of human wild-type SOD expression in Drosophila motor neurons, but did not suggest a plausible model for the shortening of life span that occurs in ALS patients.…”

Compartment-specific Protection of Iron-Sulfur Proteins by Superoxide Dismutase

“…within the compartment of its origin along with the SOD specific to that compartment is a central feature of reactive oxygen homeostasis in Drosophila. How this homeostatic mechanism might relate to the severe debilitating phenotypes that characterize Sod1 Ϫ/Ϫ mutants of Drosophila (25,26) as compared with the relatively benign phenotype of the corresponding Sod1 Ϫ/Ϫ mutant in mice (42) is unclear at present. In principle, the liberation of intracellular iron resulting from the reaction of unscavenged O 2 .…”

“…Sod1 n108 and Sod1 x39 represent null-activity alleles of the Sod1 gene (25,26). For RNA interference studies, UAS-* This work was supported by grants from the Natural Sciences and Engineering Research Council of Canada (NSERC) (to J. P. P. and A. J. H.) and by the Intramural program of the National Institute of Child Health and Human Development (NICHD).…”

Protein-coding genes are epigenetically regulated in Arabidopsis polyploids

“…Although the role of developmental regulation in silencing duplicate genes is speculative, this type of regulation has advantages for selection and adaptation of the newly formed polyploid plants. For instance, there is evidence that some protein heterodimers may not function as well as homodimers or vice versa (40,41). Thus, a silencing strategy could balance the advantage and disadvantage of having multiple copies of orthologous genes or gene products (e.g., transcriptional factors) in a polyploid cell.…”