Endonuclease G promotes mitochondrial genome cleavage and replication
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Rahel Stefanie Wiehe1, Boris Gole1,2, Laurent Chatre3,4, Paul Walther5, Enrico Calzia6, Miria Ricchetti3,4 and Lisa Wiesmüller1
1Department of Obstetrics and Gynecology, Ulm University, Ulm, 89075, Germany
2Present address: Centre for Human Molecular Genetics and Pharmacogenomics, Medical Faculty, University of Maribor, Maribor, SI-2000, Slovenia
3Department of Developmental and Stem Cell Biology, Institute Pasteur, Stem Cells and Development, 75724 Cedex 15, Paris, France
4Team Stability of Nuclear and Mitochondrial DNA, Unit of Stem Cells and Development, CNRS UMR 3738, 75724 Cedex 15, Paris, France
5Central Facility for Electron Microscopy, Ulm University, Ulm, 89081, Germany
6Institute of Anesthesiological Pathophysiology and Process Engineering, Ulm University Hospital, Ulm, 89081, Germany
Lisa Wiesmüller, email: [email protected]
Keywords: endonuclease G; oxidative damage; base excision repair; mitochondrial DNA degradation
Received: January 12, 2018 Accepted: March 12, 2018 Published: April 06, 2018
Endonuclease G (EndoG) is a nuclear-encoded endonuclease, mostly localised in mitochondria. In the nucleus EndoG participates in site-specific cleavage during replication stress and genome-wide DNA degradation during apoptosis. However, the impact of EndoG on mitochondrial DNA (mtDNA) metabolism is poorly understood. Here, we investigated whether EndoG is involved in the regulation of mtDNA replication and removal of aberrant copies. We applied the single-cell mitochondrial Transcription and Replication Imaging Protocol (mTRIP) and PCR-based strategies on human cells after knockdown/knockout and re-expression of EndoG. Our analysis revealed that EndoG stimulates both mtDNA replication initiation and mtDNA depletion, the two events being interlinked and dependent on EndoG’s nuclease activity. Stimulation of mtDNA replication by EndoG was independent of 7S DNA processing at the replication origin. Importantly, both mtDNA-directed activities of EndoG were promoted by oxidative stress. Inhibition of base excision repair (BER) that repairs oxidative stress-induced DNA damage unveiled a pronounced effect of EndoG on mtDNA removal, reminiscent of recently discovered links between EndoG and BER in the nucleus. Altogether with the downstream effects on mitochondrial transcription, protein expression, redox status and morphology, this study demonstrates that removal of damaged mtDNA by EndoG and compensatory replication play a critical role in mitochondria homeostasis.
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