The long-term objective of the proposed studies is to elucidate the role of oxidative stress in the aging process (or senescence) in animals. Endogenously-generated reactive oxygen species have been widely hypothesized to be a major causal factor in the aging process. It is postulated that the age-associated loss of functional capacity is substantially due to the accrual of molecular oxidative damage. Although the oxidative stress hypothesis of aging has evoked a considerable amount of interest, few studies have been conducted to directly test its validity. The purpose of the studies proposed here is to test one essential aspect of this hypothesis, namely, the role of mitochondria in the causation of oxidative stress and senescence. Mitochondria are the foremost intracellular generators of superoxide anion radicals and hydrogen peroxide, which are the progenitors of various other reactive oxygen metabolites. Mitochondria also exhibit an age-related increase in molecular oxidative damage and a decline in oxidative phosphorylation. Specifically, using Drosophila melanogaster as a model system, the proposed study will test the hypothesis that """"""""an elevation in the intramitochondrial activities of Mn-superoxide dismutase (SOD) and catalase, which remove superoxide anion radicals (O2-) and hydrogen peroxide, respectively, would reduce the rate of mitochondrial release of H2O2, and attenuate the senescent changes in mitochondria as well as the organism."""""""" Elevation in the activities of Mn-SOD and catalase in mitochondria will be achieved by introducing extra copies of Mn-SOD and (modified) catalase genes into the germ line of Drosophila melanogaster. A key feature of this proposal will be the use of an appropriate pre-sequence to target the catalase peptide to the mitochondrion. The effects of increased intramitochondrial activities of Mn-SOD and catalase on mitochondrial and organismic senescence will be studied. Tests of mitochondrial senescence will include: rates of O2- and H2O2 production, protein and DNA oxidative damage, respiratory rates and selected mitochondrial functions. Whether the transgenic flies exhibit a retardation of the overall aging process will be determined on the basis of mortality data and comparisons of age-related biochemical and physiological changes. It is suggested that the results of this study should critically test one of the central assumptions of the oxidative stress hypothesis of aging, namely, that mitochondrially generated reactive oxygen species constitute a major causal factor in mitochondrial and organismic senescence. This study is intended to provide new knowledge about transgenic strategies to lower the level of oxidative stress, which can then be adapted to mammalian systems. Such knowledge may also contribute to the explorations of the feasibility of genetic interventions in the aging process.
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