title: Screen for mitochondrial DNA copy number maintenance genes reveals essential role for ATP synthase.
abstract: The machinery of mitochondrial DNA (mtDNA) maintenance is only partially characterized and is of wide interest due to its involvement in disease. To identify novel components of this machinery, plus other cellular pathways required for mtDNA viability, we implemented a genome-wide RNAi screen in Drosophila S2 cells, assaying for loss of fluorescence of mtDNA nucleoids stained with the DNA-intercalating agent PicoGreen. In addition to previously characterized components of the mtDNA replication and transcription machineries, positives included many proteins of the cytosolic proteasome and ribosome (but not the mitoribosome), three proteins involved in vesicle transport, some other factors involved in mitochondrial biogenesis or nuclear gene expression, > 30 mainly uncharacterized proteins and most subunits of ATP synthase (but no other OXPHOS complex). ATP synthase knockdown precipitated a burst of mitochondrial ROS production, followed by copy number depletion involving increased mitochondrial turnover, not dependent on the canonical autophagy machinery. Our findings will inform future studies of the apparatus and regulation of mtDNA maintenance, and the role of mitochondrial bioenergetics and signaling in modulating mtDNA copy number.
authors: BioMediTech and Tampere University Hospital, University of Tampere, Tampere, Finland Department of Clinical Chemistry and Laboratory Medicine, Kyushu University Graduate school of Medical Sciences, Fukuoka, Japan Department of Medical Laboratory Science, Junshin Gakuen University, Fukuoka, Japan.
authors: Atsushi
authors: A
authors: Fukuoh
authors: BioMediTech and Tampere University Hospital, University of Tampere, Tampere, Finland.
authors: Giuseppe
authors: G
authors: Cannino
authors: BioMediTech and Tampere University Hospital, University of Tampere, Tampere, Finland.
authors: Mike
authors: M
authors: Gerards
authors: BioMediTech and Tampere University Hospital, University of Tampere, Tampere, Finland.
authors: Suzanne
authors: S
authors: Buckley
authors: BioMediTech and Tampere University Hospital, University of Tampere, Tampere, Finland.
authors: Selena
authors: S
authors: Kazancioglu
authors: BioMediTech and Tampere University Hospital, University of Tampere, Tampere, Finland.
authors: Filippo
authors: F
authors: Scialo
authors: Department of Signal Processing, Tampere University of Technology, Tampere, Finland.
authors: Eero
authors: E
authors: Lihavainen
authors: Department of Signal Processing, Tampere University of Technology, Tampere, Finland.
authors: Andre
authors: A
authors: Ribeiro
authors: BioMediTech and Tampere University Hospital, University of Tampere, Tampere, Finland.
authors: Eric
authors: E
authors: Dufour
authors: BioMediTech and Tampere University Hospital, University of Tampere, Tampere, Finland Research Program of Molecular Neurology, University of Helsinki, Helsinki, Finland howard.t.jacobs@uta.fi.
authors: Howard T
authors: HT
authors: Jacobs
mesh_terms: D000818:Animals; D001343:Autophagy; D002460:Cell Line; D004272:DNA, Mitochondrial / Q000032:analysis* / Q000737:chemistry; D029721:Drosophila Proteins / Q000235:genetics* / Q000378:metabolism; D004331:Drosophila melanogaster / Q000235:genetics / Q000378:metabolism*; D018628:Gene Dosage; D005786:Gene Expression Regulation; D015723:Gene Library; D020043:Genes, Essential; D016678:Genome; D025261:Mitochondrial Proton-Translocating ATPases / Q000235:genetics* / Q000378:metabolism; D034622:RNA Interference; D017382:Reactive Oxygen Species / Q000378:metabolism
chemical_list: D004272:DNA, Mitochondrial; D029721:Drosophila Proteins; D017382:Reactive Oxygen Species; D025261:Mitochondrial Proton-Translocating ATPases
keywords: DNA replication; complex V; mitochondrial DNA; mitochondrial biogenesis; mitophagy; nuclease; nucleoid; reactive oxygen species