Mitochondrial Topoisomerase I is Critical for Mitochondrial Integrity and Cellular Energy Metabolism

Abstract

BackgroundMitochondria contain their own DNA genome (mtDNA), as well as specific DNA replication and protein synthesis machineries. Relaxation of the circular, double-stranded mtDNA relies on the presence of topoisomerase activity. Three different topoisomerases have been identified in mitochondria: Top1mt, Top3α and a truncated form of Top2β.Methodology/Principal FindingsThe present study shows the importance of Top1mt in mitochondrial homeostasis. Here we show that Top1mt−/− murine embryonic fibroblasts (MEF) exhibit dysfunctional mitochondrial respiration, which leads decreased ATP production and compensation by increased glycolysis and fatty acid oxidation. ROS production in Top1mt−/− MEF cells is involved in nuclear DNA damage and induction of autophagy. Lack of Top1mt also triggers oxidative stress and DNA damage associated with lipid peroxidation and mitophagy in Top1mt−/− mice.Conclusion/SignificanceTogether, our data implicate Top1mt for mitochondrial integrity and energy metabolism. The compensation mechanism described here contributes to the survival of Top1mt−/− cells and mice despite alterations of mitochondrial functions and metabolism. Therefore, this study supports a novel model for cellular adaptation to mitochondrial damage.

Lack of mitochondrial topoisomerase I ( TOP1mt ) impairs liver regeneration

“…In addition to its roles in regulating mtDNA replication (8), transcription (9), and mtDNA integrity (10), Top1mt participates in mitochondrial functions. Indeed, murine embryonic fibroblasts (MEFs) generated from TOP1mt KO mice have mitochondrial defects, including excessive reactive oxygen species (ROS) production and altered polarization of mitochondrial membranes (11).…”

“…To address the role of mtDNA in liver regeneration, we took advantage of the fact that mice lacking TOP1mt are viable (5,11,17). We suspected that Top1mt might play a role in liver regeneration because we recently uncovered a critical role for TOP1mt in maintaining mtDNA homeostasis and mitochondrial function in adaptive response to doxorubicin-induced cardiotoxicity (17).…”

Poisoning of Mitochondrial Topoisomerase I by Lamellarin D

“…Some drugs have been reported to deplete mitochondrial DNA by interfering with mitochondrial DNA polymerase gamma activity (Rowe et al, 2001;Neuzil et al, 2013). Top1mt is also important for mitochondrial homeostasis (Douarre et al, 2012), mtDNA replication (Zhang and Pommier, 2008), mtDNA transcription (Sobek et al, 2013), and mtDNA integrity (Medikayala et al, 2011). Yet, there are no drugs known to specifically target Top1mt.…”

“…At least 100 ml of DNA solution were applied to polyvinylidene difluoride membranes with a slot blot apparatus. After 1 hour blocking with 5% milk in PBST (phosphatebuffered saline, Tween-20 0.1%), membranes were incubated overnight with Top1mt antibody (Douarre et al, 2012) or (nuclear) Top1 antibody (#556597; BD Biosciences, San Jose, CA) (Antony et al, 2007). After three washes in PBST, membranes were incubated with horseradish peroxidase-conjugated goat anti-mouse (1:5000 dilution) antibody (Amersham Biosciences, Piscataway, NJ) for 1 hour and washed three times.…”

“…In addition to studying Top1mt's roles in regulating mtDNA replication (Zhang and Pommier, 2008), transcription (Sobek et al, 2013), and mtDNA integrity (Medikayala et al, 2011), it has become possible to examine Top1mt functions genetically. Murine embryonic fibroblasts generated from TOP1mt knockout mice exhibit mitochondrial defects with marked increase in reactive oxygen species production, calcium signaling, hyperpolarization of mitochondrial membranes, and increased mitophagy (Douarre et al, 2012).…”