Efficient measurement of H2O2 resistance in Drosophila using an activity monitor.
We have established a method for the efficient measurement of oxidative stress resistance in Drosophila melanogaster, using a commercially available activity monitor. Conditions under which flies in glass tubes placed in the monitor can survive over one month at 25 degrees C were optimized. The active periods of flies were reduced by administration of H(2)O(2) into the media in a dose-dependent manner. Although we used only eight flies per assay, far fewer individuals than in conventional methods, it was possible to detect the effects of H(2)O(2) at a statistically significant level. Increased levels of H(2)O(2) resistance were confirmed in transgenic flies overexpressing antioxidant enzymes, catalase or Cu/Zn superoxide dismutase. We applied the method to determine oxidative stress resistance in fly lines bearing insertions of a gene misexpression vector. H(2)O(2) resistance in these flies varied considerably depending on the insertion, and positively correlated with previously determined longevity. We identified one insertion that conferred a significantly higher level of resistance to H(2)O(2) compared to controls. Molecular analysis of the insertion revealed that a misexpressed transcript matched an expressed sequence tag, and suggested that its full-length product was overproduced upon GAL4 activation. Our method should be applicable to the systematic screening for genes involved in the antioxidant mechanism in Drosophila.
Kaneuchi
Taro
T
Department of Biological Science, Tokyo Metropolitan University, 1-1 Minami-osawa, Hachioji-shi, Tokyo, 192-0397, Japan.
Togawa
Toru
T
Matsuo
Takashi
T
Fuyama
Yoshiaki
Y
Aigaki
Toshiro
T
D000375:Aging / Q000235:genetics / Q000378:metabolism; D000818:Animals; D030801:Animals, Genetically Modified; D000975:Antioxidants / Q000378:metabolism; D001522:Behavior, Animal; D002374:Catalase / Q000235:genetics; D004331:Drosophila melanogaster / Q000235:genetics* / Q000378:metabolism; D006861:Hydrogen Peroxide / Q000494:pharmacology*; D009154:Mutation; D016877:Oxidants / Q000494:pharmacology*; D018384:Oxidative Stress / Q000187:drug effects / Q000235:genetics*; D013482:Superoxide Dismutase / Q000235:genetics
D000975:Antioxidants; D016877:Oxidants; D006861:Hydrogen Peroxide; D002374:Catalase; D013482:Superoxide Dismutase