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Production of reactive oxygen species by the mitochondrial electron transport chain in Drosophila melanogaster

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Abstract

Mitochondrial free radicals and in particular mitochondrial Reactive Oxygen Species (mtROS) are considered to be totally or partially responsible for several different diseases including Parkinson, diabetes or cancer. Even more importantly, mtROS have also been proposed as the main driving force behind the aging process. Thus, in the last decade, there has been a growing interest in the role of free radicals as signalling molecules. Collectively this makes understanding mechanisms controlling free radical production extremely important. There is extensive published literature on mammalian models (essentially rat, mouse and guinea pig) however; this is not the case in Drosophila melanogaster. Drosophila is an excellent model to study different physiological and pathological processes. Additionally a robust method to study mtROS is extremely useful. In the present article, we describe a simple—but extremely sensitive—method to study mtROS production in Drosophila. We have performed various experiments to determine which specific respiratory complexes produce free radicals in the electron transport chain of Drosophila melanogaster. Complex I is the main generator of ROS in Drosophila mitochondria, leaking electrons either in the forward or reverse direction. The production of ROS during reverse electron transport can be prevented either by rotenone or by the oxidation of NADH by complex I. These results clearly show that Drosophila mitochondria function in a very similar way to mammalian mitochondria, and therefore are a very relevant experimental model for biochemical studies related to ageing.

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References

  • Ballard JW, Melvin RG, Miller JT, Katewa SD (2007) Sex differences in survival and mitochondrial bioenergetics during aging in Drosophila. Aging Cell 6:699–708

    Article  CAS  Google Scholar 

  • Brown GC, Borutaite V (2008) Regulation of apoptosis by the redox state of cytochrome c. Biochim Biophys Acta 1777:877–881

    Article  CAS  Google Scholar 

  • Chen Q, Moghaddas S, Hoppel CL, Lesnefsky EJ (2008) Ischemic defects in the electron transport chain increase the production of reactive oxygen species from isolated rat heart mitochondria. Am J Physiol Cell Physiol 294:C460–C466

    Article  CAS  Google Scholar 

  • Das N, Levine RL, Orr WC, Sohal RS (2001) Selectivity of protein oxidative damage during aging in Drosophila melanogaster. Biochem J 360:209–216

    Article  CAS  Google Scholar 

  • Giorgio M, Migliaccio E, Orsini F, Paolucci D, Moroni M, Contursi C, Pelliccia G, Luzi L, Minucci S, Marcaccio M, Pinton P, Rizzuto R, Bernardi P, Paolucci F, Pelicci PG (2005) Electron transfer between cytochrome c and p66Shc generates reactive oxygen species that trigger mitochondrial apoptosis. Cell 122:221–233

    Article  CAS  Google Scholar 

  • Giulivi C, Ross-Inta C, Horton AA, Luckhart S (2008) Metabolic pathways in Anopheles stephensi mitochondria. Biochem J 415:309–316

    Article  CAS  Google Scholar 

  • Gredilla R, Sanz A, Lopez-Torres M, Barja G (2001) Caloric restriction decreases mitochondrial free radical generation at complex I and lowers oxidative damage to mitochondrial DNA in the rat heart. FASEB J 15:1589–1591

    CAS  Google Scholar 

  • Ishikawa K, Takenaga K, Akimoto M, Koshikawa N, Yamaguchi A, Imanishi H, Nakda K, Honma Y, Hayashi J (2008) ROS-generating mitochondrial DNA mutations can regulate tumor cell metastasis. Science 320:661–664

    Article  CAS  Google Scholar 

  • Katewa SD, Ballard JW (2007) Sympatric Drosophila simulans flies with distinct mtDNA show age related differences in mitochondrial metabolism. Insec Biochem Mol Biol 37:323–932

    Google Scholar 

  • Kushnareva Y, Murphy AN, Andreyev A (2002) Complex I-mediated reactive oxygen species generation: modulation by cytochrome c and NAD(P) + oxidation-reduction state. Biochem J 368:545–553

    Article  CAS  Google Scholar 

  • Lambert AJ, Merry BJ (2005) Lack of effect of caloric restriction on bioenergetics and reactive oxygen species production in intact rat hepatocytes. J Gerontol A Biol Sci Med Sci 60:175–180

    Google Scholar 

  • Lambert AJ, Boysen HM, Buckingham JA, Yang T, Podlutsky A, Austad SN, Kunz TH, Buffenstein R, Brand MD (2007) Low rates of hydrogen peroxide production by isolated heart mitochondria associate with long maximum lifespan in vertebrate homeotherms. Aging Cell 6:607–618

    Article  CAS  Google Scholar 

  • Miwa S, Brand MD (2005) The topology of superoxide production by complex III and glycerol 3-phosphate dehydrogenase in Drosophila mitochondria. Biochim Biophys Acta 1709:214–219

    Article  CAS  Google Scholar 

  • Miwa S, St-Pierre J, Partridge L, Brand MD (2003) Superoxide and hydrogen peroxide production by Drosophila mitochondria. Free Rad Biol Med 35:938–948

    Article  CAS  Google Scholar 

  • Muller FL, Liu Y, Van Remmen H (2004) Complex III releases superoxide to both sides of the inner mitochondrial membrane. J Biol Chem 279:49064–49073

    Article  CAS  Google Scholar 

  • Muller FL, Liu Y, Abdul-Ghani MA, Lustgarten MS, Bhattacharya A, Jang YC, Van Remmen H (2008) High rates of superoxide production in skeletal-muscle mitochondria respiring on both complex I and complex II-linked substrates. Biochem J 409:491–499

    Article  CAS  Google Scholar 

  • Murphy MP (2009) How mitochondria produce reactive oxygen species. Biochem J 417:1–13

    Article  CAS  Google Scholar 

  • Mracek T, Pecinova A, Vrbacky M, Drahota Z, Houstek J (2009) High efficiency of ROS production by glycerophosphate dehydronase in mammalian mitochondria. Arch Biochem Biophys 481:30–36

    Article  CAS  Google Scholar 

  • Ruch W, Cooper PH, Baggiolini M (1983) Assay of H2O2 release by macrophages and neutrophils with homovanillic acid and horse-radish peroxidase. J Immunol Meth 63:347–357

    Article  CAS  Google Scholar 

  • Sanz A, Barja G (2006) Estimation of the rate of production of oxygen radicals at mitochondria. In Handbook of models for human aging, Academic Press, 16,183–189

  • Sanz A, Pamplona R, Barja G (2006a) Is the mitochondrial free radical theory of aging intact? Antiox Redox Signal 8:582–599

    Article  CAS  Google Scholar 

  • Sanz A, Caro P, Ayala V, Portero-Otin M, Pamplona R, Barja G (2006b) Methionine restriction decreases mitochondrial oxygen radical generation and leak as well as oxidative damage to mitochondrial DNA and proteins. FASEB J 20:1064–1073

    Article  CAS  Google Scholar 

  • Sanz A, Fernandez-Ayala DJ, Stefanatos R, Jacobs HT (2009) Mitochondrial free radical production does not control life span in Drosophila melanogaster (submitted)

  • Schofeld P, Wojtczak L (2007) Fatty acids decrease mitochondrial generation of reactive oxygen species at reverse electron transport but increase it at the forward transport. Biochim Biophys Acta 1767:1032–1040

    Article  CAS  Google Scholar 

  • Shokolenko I, Venediktova N, Bochkareva A, Wilson GL, Alexeyev MF (2009) Oxidative stress induces degradation of mitochondrial DNA. Nucleic Acid Res 38:2539–2548

    Article  CAS  Google Scholar 

  • Starkov AA, Fiskum G, Chinopoulos C, Lorenzo BJ, Browne SE, Patel MS, Beal MF (2004) Mitochondrial alpha-ketoglutarate dehydrogenase complex generates reactive oxygen species. J Neurosci 24:7779–7788

    Article  CAS  Google Scholar 

  • Suter M, Richter C (1999) Fragmented mitochondrial DNA is the predominant carrier of oxidized DNA bases. Biochemistry 38:459–464

    Article  CAS  Google Scholar 

  • Tong JJ, Schriner SE, McCleary D, Day BJ, Wallace DC (2007) Life extension through neurofibromin mitochondrial regulation and antioxidant therapy for neurofibromatosis-1 in Drosophila melanogaster. Nat Genet 39:476–485

    Article  CAS  Google Scholar 

  • Tretter L, Takacs K, Hegedus V, Adam-Vizi V (2007) Characteristics of alpha-glycerophosphate-evoked H2O2 generation in brain mitochondria. J Neurochem 100:650–663

    Article  CAS  Google Scholar 

  • Van Remmen H, Ikeno Y, Hamilton M, Pahlavani M, Wolf N, Thorpe SR, Alderson NL, Baynes JW, Epstein CJ, Huang TT, Nelson J, Strong R, Richardson A (2003) Life-long reduction in MnSOD activity results in increased DNA damage and higher incidence of cancer but does not accelerate aging. Physiol Genomics 16:29–37

    Article  CAS  Google Scholar 

  • Ventura B, Genova ML, Bovina C, Formiggini G, Lenaz G (2002) Control of oxidative phosphorylation by complex I in rat liver mitochondria: implication for aging. Biochim Biophys Acta 1553:249–260

    Article  CAS  Google Scholar 

  • Wen JJ, Garg NJ (2008) Mitochondrial generation of reactive oxygen species is enhanced at the Q(o) site of the complex III in the myocardium of Trypanosoma cruzi-infected mice: beneficial effects of an antioxidant. J Bioenerg Biomembr 40:587–598

    Article  CAS  Google Scholar 

  • Zoccarato F, Cavallini L, Bortolami S, Alendre A (2007) Succinate modulation of H2O2 release at NADH:ubiquinone oxidoreductase (Complex I) in brain mitochondria. Biochem J 406: 25–129

    Article  CAS  Google Scholar 

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Correspondence to Alberto Sanz.

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Sanz, A., Stefanatos, R. & McIlroy, G. Production of reactive oxygen species by the mitochondrial electron transport chain in Drosophila melanogaster . J Bioenerg Biomembr 42, 135–142 (2010). https://doi.org/10.1007/s10863-010-9281-z

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  • DOI: https://doi.org/10.1007/s10863-010-9281-z

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