Zelenka J 2013 Abstract MiP2013
|Zelenka J, Alan L, Jezek P(2013) Response of cancer cells to mitochondrial DNA damage. Mitochondr Physiol Network 18.08.|
Human mitochondrial DNA (mtDNA) is present in each cell in hundreds to thousands of copies representing the autonomous hereditary unit of our genome and encoding 13 core polypeptides of the oxidative phosphorylation complexes together with 22 tRNAs and 2 rRNAs. It is stored in the form of DNA-protein bodies called nucleoids which are regularly distributed in the mitochondrial reticulum. This redundancy together with the proximity to free radical-producing respiratory complexes makes mtDNA highly prone to accumulation of mutations which eventually lead to cellular bioenergetics failure. Therefore, mtDNA quality and quantity is under tight control of multiple independent pathways, e.g. macroautophagy (mitophagy).
This study aimed at investigating the role of mitophagy in the response of cancer cells to DNA intercalating agents, which are routinely used for cancer treatment and are known to preferentially bind to mtDNA .
Three different cancer cell lines (HepG2, HeLa, Flp) were treated with mtDNA intercalators ethidium bromide (EtBr). The composition and distribution of nucleoids was followed with SIM and FPALM nanoscopy or regular confocal microscopy. Moreover, mitochondrial bioenergetics was assessed using Oroboros Oxygraph-2k. mtDNA copy number and gene expression were analyzed with Q-PCR and mitochondrial fluorescence in situ hybridization .
EtBr treatment resulted in inhibition of mtDNA transcription and replication and in a significant drop of cellular respiratory capacity. Moreover, it caused reorganization of nucleoids into large clusters stored in separate mitochondrial cisternae and relatively depleted of mtSSB protein. Interestingly, the mitophagy marker LC3BII was significantly upregulated while mtDNA copy number per cell decreased dramatically. However, subsequent studies with autophagy inhibitors wortmannin and chloroquin revealed that nucleoid clusters were protected from mitophagy. These results suggest that cancer cells respond to global mtDNA damage by forming novel structures – nucleoid clusters, which apparently protect cells against complete loss of mtDNA under conditions of extreme genotoxic stress. The relevance of this process for human physiology and medicine will be discussed.
• O2k-Network Lab: CZ Prague Jezek P
Labels: MiParea: Respiration, mt-Medicine Pathology: Cancer Stress:Ischemia-reperfusion
Tissue;cell: Other cell lines
Regulation: Aerobic glycolysis, ATP, Coupling efficiency;uncoupling, Ion;substrate transport Coupling state: ROUTINE, ET
Institute of Physiology, Academy of Sciences, Prague, Czech Republic. - Email: firstname.lastname@example.org
Supported by grants no. P305/12/P388 from Czech Science Foundation and by GAUK 418811 from Grant Agency of Charles University.
- Ashley N, Poulton J (2009) Anticancer DNA intercalators cause p53-dependent mitochondrial DNA nucleoid re-modelling. Oncogene 28: 3880–3891.
- Alan L, Zelenka J, Jezek J, Dlaskova A, Jezek P (2010) Fluorescent in situ hybridization of mitochondrial DNA and RNA. Acta Biochim Pol 57: 403-408.