The Oxidative Stress Hypothesis of Aging postulates that the imbalance between oxidative stress and antioxidant defenses results in the accumulation of oxidative stress/damage with age and thus serves as a limiting factor on longevity. Glutathione reductase and glutathione peroxidase represent an important enzymatic component of the antioxidant system in most organisms (including mammals). In Drosophila melanogaster and several other insects, these enzymes are absent. There is however a second group of enzymes, thioredoxin reductase and thioredoxin peroxidases, which may in part replace the glutathione-dependent system in insects. These enzymes then may play an important role in insect aging.

The general purpose of this study is to test the oxidative stress hypothesis of aging by manipulating levels of Dpx 5037, a thioredoxin peroxidase normally found in the mitochondria, also referred to as mtDPX. mtDPX is highly expressed in adult flies. To test the effects of over-expression of the gene on lifespan and resistance to oxidative stress, transgenic methods were used. A mtDPX (DPX5037) cDNA-pUAST construct was cloned, sequenced, and then micro-injected into embryos along with the delta 2-3 helper plasmid. Additional lines were established by P element mobilization. The successful transgenic lines were confirmed by southern blot using an mtDPX (DPX5037) probe and chromosome positions were inferred by patterns of segregation with respect to dominant markers. All the transgenic lines used for experiments have exactly one extra copy of mtDPX (DPX5037) on either the second or third chromosome. The transgenic lines were backcrossed to a reference yellow white background for at least six generations to ensure a relatively homogeneous genetic background. To overexpress mtDPX (Dpx5037), the transgenic mtDPX(DPX5037)—pUAST was crossed with different tissue-specific GAL4 lines. The levels of mtDPX (DPX5037) protein in overexpressed lines were 2.3 to 5.7 times higher than that of the controls as determined by western blot using anti-mtDPX (DPX5037) anti-sera. The lifespan results showed that when mtDPX (Dpx5037) was overexpressed in Drosophila melanogaster globally by means of the tublin-GAL4 driver, the mean lifespan was increased by up to 30 percent. When mtDPX (Dpx5037) was overexpressed exclusively in the neuronal system using the pan-neuronal driver, Appl-GAL4, greater resistance to paraquat was observed. No effects on mean lifespan were observed when mtDPX (Dpx5037) was over-expressed globally by the daughterless-GAL4 driver, in the neuronal system by pan-neuronal drivers, Elav-GAL4, Apple-GAL4, or the motor neuron-specific driver D42-GAL4, or in muscle tissue by the 2703-GAL4 driver. Also no beneficial effects in response to hydrogen peroxide and hyperoxia when mtDPX (Dpx5037) was overexpressed by any of the driver set mentioned above. These results suggest that: (1) mtDPX (Dpx5037) does play some role on lifespan and oxidative stress, when over-expressed in certain tissues, although the complexity of the effects observed suggest that elements of tissue-specific redox regulation may need to be examined more closely; (2) the results provide some support for the oxidative stress hypothesis of aging. (3) the correlation between lifespan and oxidative stress resistance is neither confirmed nor invalidated by this study.