title: Over-expression of the catalytic core of mitochondrial DNA (mtDNA) polymerase in the nervous system of Drosophila melanogaster reduces median life span by inducing mtDNA depletion.
abstract: DNA polymerase gamma (pol gamma) is the sole DNA polymerase devoted to mitochondrial DNA (mtDNA) replication. We have characterized the molecular and physiological effects of over-expression of the catalytic subunit of pol gamma, pol gamma-alpha, in the nervous system of Drosophila melanogaster using the upstream activation sequence (UAS)/yeast transcriptional activator by binding to UAS (GAL4) system. Tissue-specific over-expression of pol gamma-alpha was confirmed by immunoblot analysis, whereas the very low levels of endogenous protein are undetectable in UAS or GAL4 control lines. The transgenic flies over-expressing pol gamma-alpha in the nervous system showed a moderate increase in pupal lethality, and a significant decrease in the median life span of adult flies. Moreover, these flies displayed a decrease in the rate of synthesis of mtDNA, which is accompanied by a significant mtDNA depletion, and a corresponding decrease in the levels of mitochondrial transcription factor A (mtTFA). Biochemical analysis showed an oxidative phosphorylation (OXPHOS) defect in transgenic flies, which were more susceptible to oxidative stress. Although we did not detect apoptosis in the nervous system of adult transgenic flies, brains of larvae over-expressing pol gamma-alpha showed evidence of increased cell death that correlates with the observed phenotypes. Our data establish an animal model that mimics some of the features of human mtDNA depletion syndromes.
authors.affiliation: Departamento de Bioquímica, Instituto de Investigaciones Biomédicas Alberto Sols CSIC-UAM, Facultad de Medicina CIBERER ISCIII, Universidad Autónoma de Madrid, Madrid, Spain. fmartinez@cbm.uam.es
authors.forename: Francisco
authors.initials: F
authors.lastname: Martínez-Azorín
authors.forename: Manuel
authors.initials: M
authors.lastname: Calleja
authors.forename: Rosana
authors.initials: R
authors.lastname: Hernández-Sierra
authors.forename: Carol L
authors.initials: CL
authors.lastname: Farr
authors.forename: Laurie S
authors.initials: LS
authors.lastname: Kaguni
authors.forename: Rafael
authors.initials: R
authors.lastname: Garesse
mesh_terms: D000818:Animals; D030801:Animals, Genetically Modified; D020134:Catalytic Domain / Q000502:physiology*; D000074002:DNA Polymerase gamma; D004272:DNA, Mitochondrial / Q000737:chemistry* / Q000235:genetics; D004259:DNA-Directed DNA Polymerase / Q000378:metabolism; D029721:Drosophila Proteins / Q000235:genetics / Q000378:metabolism; D004331:Drosophila melanogaster; D015870:Gene Expression / Q000502:physiology*; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D008297:Male; D008928:Mitochondria; D028361:Mitochondrial Diseases / Q000503:physiopathology*; D009245:NADH Dehydrogenase / Q000378:metabolism; D009420:Nervous System / Q000378:metabolism*; D018384:Oxidative Stress / Q000502:physiology; D016019:Survival Analysis; D013997:Time Factors
chemical_list: D004272:DNA, Mitochondrial; D029721:Drosophila Proteins; D049452:Green Fluorescent Proteins; D009245:NADH Dehydrogenase; D000074002:DNA Polymerase gamma; D004259:DNA-Directed DNA Polymerase