SPECIFIC INHIBITION BY ETHIDIUM BROMIDE OF MITOCHONDRIAL DNA SYNTHESIS IN <i>PHYSARUM POLYCEPHALUM </i>

Horwitz, Henry B.; Holt, Charles E.

doi:10.1083/jcb.49.2.546

Cited by 29 publications

(6 citation statements)

References 18 publications

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“…After 14 days of treatment, mtDNA copy number in cells and lactate levels in cell culture media were measured relative to the mock-treated cells. ddC, a known inhibitor of DNA polymerase ␥ (24), and ethidium bromide (EtBr), a mitochondrial replication inhibitor (25), were used as positive controls. Data from these experiments are summarized in Tables 3 and 4.…”

Section: Resultsmentioning

confidence: 99%

Analysis of the Potential for N ⁴ -Hydroxycytidine To Inhibit Mitochondrial Replication and Function

Sticher

Mitchell

et al. 2020

Antimicrob Agents Chemother

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N 4 -Hydroxycytidine (NHC) is an antiviral ribonucleoside analog that acts as a competitive alternative substrate for virally encoded RNA-dependent RNA polymerases. It exhibits measurable levels of cytotoxicity, with 50% cytotoxic concentration values ranging from 7.5 M in CEM cells and up to Ͼ100 M in other cell lines. The mitochondrial DNA-dependent RNA polymerase (POLRMT) has been shown to incorporate some nucleotide analogs into mitochondrial RNAs, resulting in substantial mitochondrial toxicity. NHC was tested in multiple assays intended to determine its potential to cause mitochondrial toxicity. NHC showed similar cytotoxicity in HepG2 cells incubated in a glucose-free and glucose-containing media, suggesting that NHC does not impair mitochondrial function in this cell line based on the Crabtree effect. We demonstrate that the 5=-triphosphate of NHC can be used by POLRMT for incorporation into nascent RNA chain but does not cause immediate chain termination. In PC-3 cells treated with NHC, the 50% inhibitory concentrations of mitochondrial protein expression inhibition were 2.7-fold lower than those for nuclear-encoded protein expression, but this effect did not result in selective mitochondrial toxicity. A 14-day incubation of HepG2 cells with NHC had no effect on mitochondrial DNA copy number or extracellular lactate levels. In CEM cells treated with NHC at 10 M, a slight decrease (by ϳ20%) in mitochondrial DNA copy number and a corresponding slight increase in extracellular lactate levels were detected, but these effects were not enhanced by an increase in NHC treatment concentration. In summary, the results indicate that mitochondrial impairment by NHC is not the main contributor to the compound's observed cytotoxicity in these cell lines.

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Section: Resultsmentioning

confidence: 99%

Analysis of the Potential for N ⁴ -Hydroxycytidine To Inhibit Mitochondrial Replication and Function

Sticher

Mitchell

et al. 2020

Antimicrob Agents Chemother

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“…To investigate the causal role of low mtDNA copy number in promoting EMT, we used two alternative models: one in which mtDNA content is selectively reduced by treatment with low doses of ethidium bromide (EtBr), which does not affect nuclear DNA replication, 24,25 and second in which mtDNA is depleted by genetic manipulation of TFAM. To delineate the contribution of reduced mtDNA copy number in tumor initiation and metastatic progression through EMT, we selected human mammary epithelial cells of non-carcinoma (MCF10A) and carcinoma (MCF7) origin.…”

Section: Introductionmentioning

confidence: 99%

Mitochondrial retrograde signaling induces epithelial–mesenchymal transition and generates breast cancer stem cells

et al. 2013

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Metastatic breast tumors undergo epithelial-to-mesenchymal transition (EMT), which renders them resistant to therapies targeted to the primary cancers. The mechanistic link between mtDNA (mitochondrial DNA) reduction, often seen in breast cancer patients, and EMT is unknown. We demonstrate that reducing mtDNA content in human mammary epithelial cells (hMECs) activates Calcineurin (Cn)-dependent mitochondrial retrograde signaling pathway, which induces EMT-like reprogramming to fibroblastic morphology, loss of cell polarity, contact inhibition and acquired migratory and invasive phenotype. Notably, mtDNA reduction generates breast cancer stem cells. In addition to retrograde signaling markers, there is an induction of mesenchymal genes but loss of epithelial markers in these cells. The changes are reversed by either restoring the mtDNA content or knockdown of CnAα mRNA, indicating the causal role of retrograde signaling in EMT. Our results point to a new therapeutic strategy for metastatic breast cancers targeted to the mitochondrial retrograde signaling pathway for abrogating EMT and attenuating cancer stem cells, which evade conventional therapies. We report a novel regulatory mechanism by which low mtDNA content generates EMT and cancer stem cells in hMECs.

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“…Furthermore, in C. elegans , EtBr has been shown to activate the mtUPR as evidenced by the increase expression of hsp-60 , the mitochondrial chaperone that responds to mitochondrial dysfunction upon EtBr exposure [ 15 , 16 ]. Although it is possible that EtBr has some effect on nuclear DNA, it is negligible compared to its effect on the mitochondria [ 17 ].…”

Section: Resultsmentioning

confidence: 99%

Loss of hif-1 promotes resistance to the exogenous mitochondrial stressor ethidium bromide in Caenorhabditis elegans

2016

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BackgroundMitochondrial dysfunction is one of the leading causes of neurological disorders in humans. Mitochondrial perturbations lead to adaptive mechanisms that include HIF-1 stabilization, though the consequences of increased levels of HIF-1 following mitochondrial stress remain poorly understood.ResultsUsing Caenorhabditis elegans, we show that a hif-1 loss-of-function mutation confers resistance towards the mitochondrial toxin ethidium bromide (EtBr) and suppresses EtBr-induced production of ROS. In mammals, the PD-related gene DJ-1 is known to act as a redox sensor to confer protection against antioxidants and mitochondrial inhibitors. A deletion mutant of the C. elegans homolog djr-1.1 also showed increased resistance to EtBr. Furthermore, our data implicates p38 MAP kinase as an indispensable factor for survival against mitochondrial stress in both hif-1 and djr-1.1 mutants. ConclusionsWe propose that EtBr-induced HIF-1 activates pathways that are antagonistic in conferring protection against EtBr toxicity and that blocking HIF-1 activity may promote survival in cells with compromised mitochondrial function.

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SPECIFIC INHIBITION BY ETHIDIUM BROMIDE OF MITOCHONDRIAL DNA SYNTHESIS IN PHYSARUM POLYCEPHALUM

Cited by 29 publications

References 18 publications

Analysis of the Potential for N ⁴ -Hydroxycytidine To Inhibit Mitochondrial Replication and Function

Analysis of the Potential for N ⁴ -Hydroxycytidine To Inhibit Mitochondrial Replication and Function

Mitochondrial retrograde signaling induces epithelial–mesenchymal transition and generates breast cancer stem cells

Loss of hif-1 promotes resistance to the exogenous mitochondrial stressor ethidium bromide in Caenorhabditis elegans

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SPECIFIC INHIBITION BY ETHIDIUM BROMIDE OF MITOCHONDRIAL DNA SYNTHESIS IN PHYSARUM POLYCEPHALUM

Cited by 29 publications

References 18 publications

Analysis of the Potential for N 4 -Hydroxycytidine To Inhibit Mitochondrial Replication and Function

Analysis of the Potential for N 4 -Hydroxycytidine To Inhibit Mitochondrial Replication and Function

Mitochondrial retrograde signaling induces epithelial–mesenchymal transition and generates breast cancer stem cells

Loss of hif-1 promotes resistance to the exogenous mitochondrial stressor ethidium bromide in Caenorhabditis elegans

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Analysis of the Potential for N ⁴ -Hydroxycytidine To Inhibit Mitochondrial Replication and Function

Analysis of the Potential for N ⁴ -Hydroxycytidine To Inhibit Mitochondrial Replication and Function