Novel insights into the mode of action of 1,4-dioxane using a systems screening approach

Abstract1,4-Dioxane (1,4-DX) is an environmental contaminant found in drinking water throughout the United States (US). While it is a suspected liver carcinogen, there is no federal or state maximum contaminant level for 1,4-DX in drinking water. Very little is known about the mechanisms by which this chemical elicits liver carcinogenicity. In the present study, female BDF-1 mice were exposed to 1,4-DX (0, 50, 500 and 5,000 mg/L) in their drinking water for one or four weeks, to explore the toxic effects. Histopathological studies and a multi-omics approach (transcriptomics and metabolomics) were performed to investigate potential mechanisms of toxicity. Immunohistochemical analysis of the liver revealed increased H2AXγ-positive hepatocytes (a marker of DNA double strand breaks), and an expansion of precholangiocytes (reflecting both DNA damage and repair mechanisms) after exposure. Liver transcriptomics revealed 1,4-DX-induced perturbations in signaling pathways predicted to impact the oxidative stress response, detoxification, and DNA damage. Liver, kidney, feces and urine metabolomic profiling revealed no effect of 1,4-DX exposure, and bile acid quantification in liver and feces similarly showed no effect of exposure. We speculate that the results may be reflective of DNA damage being counterbalanced by the repair response, with the net result being a null overall effect on the systemic biochemistry of the exposed mice. Our results show a novel approach for the investigation of environmental chemicals that do not elicit cell death but have activated the repair systems in response to 1,4-DX exposure.

Download Full-text

Identification of dose-dependent DNA damage and repair responses from subchronic exposure to 1,4-dioxane in mice using a systems analysis approach

Toxicological Sciences ◽

10.1093/toxsci/kfab030 ◽

2021 ◽

Author(s):

Georgia Charkoftaki ◽

Jaya Prakash Golla ◽

Alvaro Santos-Neto ◽

David J Orlicky ◽

Rolando Garcia-Milian ◽

...

Keyword(s):

Dna Damage ◽

Drinking Water ◽

Immunohistochemical Analysis ◽

The United States ◽

Environmental Chemicals ◽

Dna Double Strand Breaks ◽

Dna Damage And Repair ◽

Mechanisms Of Toxicity ◽

Novel Approach ◽

Histopathological Studies

Abstract 1,4-Dioxane (1,4-DX) is an environmental contaminant found in drinking water throughout the United States (US). While it is a suspected liver carcinogen, there is no federal or state maximum contaminant level for 1,4-DX in drinking water. Very little is known about the mechanisms by which this chemical elicits liver carcinogenicity. In the present study, female BDF-1 mice were exposed to 1,4-DX (0, 50, 500 and 5,000 mg/L) in their drinking water for one or four weeks, to explore the toxic effects. Histopathological studies and a multi-omics approach (transcriptomics and metabolomics) were performed to investigate potential mechanisms of toxicity. Immunohistochemical analysis of the liver revealed increased H2AXγ-positive hepatocytes (a marker of DNA double strand breaks), and an expansion of precholangiocytes (reflecting both DNA damage and repair mechanisms) after exposure. Liver transcriptomics revealed 1,4-DX-induced perturbations in signaling pathways predicted to impact the oxidative stress response, detoxification, and DNA damage. Liver, kidney, feces and urine metabolomic profiling revealed no effect of 1,4-DX exposure, and bile acid quantification in liver and feces similarly showed no effect of exposure. We speculate that the results may be reflective of DNA damage being counterbalanced by the repair response, with the net result being a null overall effect on the systemic biochemistry of the exposed mice. Our results show a novel approach for the investigation of environmental chemicals that do not elicit cell death but have activated the repair systems in response to 1,4-DX exposure.

Download Full-text

Arsenic Biotransformation as a Cancer Promoting Factor by Inducing DNA Damage and Disruption of Repair Mechanisms

Molecular Biology International ◽

10.4061/2011/718974 ◽

2011 ◽

Vol 2011 ◽

pp. 1-11 ◽

Cited By ~ 32

Author(s):

Victor D. Martinez ◽

Emily A. Vucic ◽

Marta Adonis ◽

Lionel Gil ◽

Wan L. Lam

Keyword(s):

Dna Damage ◽

Drinking Water ◽

Repair Process ◽

Dna Methyltransferases ◽

Health Concern ◽

Dna Damage And Repair ◽

Repair Mechanisms ◽

Contaminated Drinking Water ◽

High Concentrations ◽

Epigenetic Patterns

Chronic exposure to arsenic in drinking water poses a major global health concern. Populations exposed to high concentrations of arsenic-contaminated drinking water suffer serious health consequences, including alarming cancer incidence and death rates. Arsenic is biotransformed through sequential addition of methyl groups, acquired from s-adenosylmethionine (SAM). Metabolism of arsenic generates a variety of genotoxic and cytotoxic species, damaging DNA directly and indirectly, through the generation of reactive oxidative species and induction of DNA adducts, strand breaks and cross links, and inhibition of the DNA repair process itself. Since SAM is the methyl group donor used by DNA methyltransferases to maintain normal epigenetic patterns in all human cells, arsenic is also postulated to affect maintenance of normal DNA methylation patterns, chromatin structure, and genomic stability. The biological processes underlying the cancer promoting factors of arsenic metabolism, related to DNA damage and repair, will be discussed here.

Download Full-text

Mammalian Cell DNA Damage and Repair Kinetics of Monohaloacetic Acid Drinking Water Disinfection By-Products

Environmental Science & Technology ◽

10.1021/es901852z ◽

2009 ◽

Vol 43 (21) ◽

pp. 8437-8442 ◽

Cited By ~ 33

Author(s):

Yukako Komaki ◽

Justin Pals ◽

Elizabeth D. Wagner ◽

Benito J. Mariñas ◽

Michael J. Plewa

Keyword(s):

Dna Damage ◽

Drinking Water ◽

Mammalian Cell ◽

Water Disinfection ◽

Dna Damage And Repair ◽

Drinking Water Disinfection ◽

Kinetics Of ◽

By Products

Download Full-text

DNA damage alters nuclear mechanics through chromatin reorganisation

10.1101/2020.07.10.197517 ◽

2020 ◽

Author(s):

Ália dos Santos ◽

Alexander W. Cook ◽

Rosemarie E Gough ◽

Martin Schilling ◽

Nora Aleida Olszok ◽

...

Keyword(s):

Dna Damage ◽

Genomic Instability ◽

Molecular Diffusion ◽

Biomechanical Properties ◽

Protective Role ◽

Dna Lesions ◽

Dna Double Strand Breaks ◽

Dna Damage And Repair ◽

Nuclear Mechanics ◽

Dna Damage Signalling

ABSTRACTDNA double-strand breaks (DSBs) drive genomic instability. For efficient and accurate repair of these DNA lesions, the cell activates DNA damage repair pathways. However, it remains unknown how these processes may affect the biomechanical properties of the nucleus and what role nuclear mechanics play in DNA damage and repair efficiency.Here, we used Atomic Force Microscopy (AFM) to investigate nuclear mechanical changes, arising from externally induced DNA damage. We found that nuclear stiffness is significantly reduced after cisplatin treatment, as a consequence of DNA damage signalling. This softening was linked to global chromatin decondensation, which improves molecular diffusion within the organelle. We propose that this can increase recruitment for repair factors. Interestingly, we also found that reduction of nuclear tension, through cytoskeletal relaxation, has a protective role to the cell and reduces accumulation of DNA damage. Overall, these changes protect against further genomic instability and promote DNA repair. We propose that these processes may underpin the development of drug resistance.

Download Full-text

γ-H2AX Foci Persistence at Chromosome Break Suggests Slow and Faithful Repair Phases Restoring Chromosome Integrity

Cancers ◽

10.3390/cancers11091397 ◽

2019 ◽

Vol 11 (9) ◽

pp. 1397 ◽

Cited By ~ 2

Author(s):

Michelle Ricoul ◽

Tamizh Selvan Gnana Sekaran ◽

Patricia Brochard ◽

Cecile Herate ◽

Laure Sabatier

Keyword(s):

Dna Damage ◽

Chromosome Rearrangement ◽

Chromosome Break ◽

Dna Double Strand Breaks ◽

Chromosome Fusion ◽

Post Irradiation ◽

Dna Lesion ◽

Novel Approach ◽

Chromosome Integrity ◽

Kinetics Of

Many toxic agents can cause DNA double strand breaks (DSBs), which are in most cases quickly repaired by the cellular machinery. Using ionising radiation, we explored the kinetics of DNA lesion signaling and structural chromosome aberration formation at the intra- and inter-chromosomal level. Using a novel approach, the classic Premature Chromosome Condensation (PCC) was combined with γ-H2AX immunofluorescence staining in order to unravel the kinetics of DNA damage signalisation and chromosome repair. We identified an early mechanism of DNA DSB joining that occurs within the first three hours post-irradiation, when dicentric chromosomes and chromosome exchanges are formed. The slower and significant decrease of ”deleted chromosomes” and 1 acentric telomere fragments observed until 24 h post-irradiation, leads to the conclusion that a second and error-free repair mechanism occurs. In parallel, we revealed remaining signalling of γ-H2AX foci at the site of chromosome fusion long after the chromosome rearrangement formation. Moreover there is important signalling of foci on the site of telomere and sub-telomere sequences suggesting either a different function of γ-H2AX signalling in these regions or an extreme sensibility of the telomere sequences to DNA damage that remains unrepaired 24 h post-irradiation. In conclusion, chromosome repair happens in two steps, including a last and hardly detectable one because of restoration of the chromosome integrity.

Download Full-text

DNA damage alters nuclear mechanics through chromatin reorganization

Nucleic Acids Research ◽

10.1093/nar/gkaa1202 ◽

2020 ◽

Author(s):

Ália dos Santos ◽

Alexander W Cook ◽

Rosemarie E Gough ◽

Martin Schilling ◽

Nora A Olszok ◽

...

Keyword(s):

Dna Damage ◽

Genomic Instability ◽

Molecular Diffusion ◽

Biomechanical Properties ◽

Protective Role ◽

Dna Double Strand Breaks ◽

Dna Damage And Repair ◽

Nuclear Mechanics ◽

Repair Efficiency ◽

Dna Damage Signalling

Abstract DNA double-strand breaks drive genomic instability. However, it remains unknown how these processes may affect the biomechanical properties of the nucleus and what role nuclear mechanics play in DNA damage and repair efficiency. Here, we have used Atomic Force Microscopy to investigate nuclear mechanical changes, arising from externally induced DNA damage. We found that nuclear stiffness is significantly reduced after cisplatin treatment, as a consequence of DNA damage signalling. This softening was linked to global chromatin decondensation, which improves molecular diffusion within the organelle. We propose that this can increase recruitment for repair factors. Interestingly, we also found that reduction of nuclear tension, through cytoskeletal relaxation, has a protective role to the cell and reduces accumulation of DNA damage. Overall, these changes protect against further genomic instability and promote DNA repair. We propose that these processes may underpin the development of drug resistance.

Download Full-text

Differential Response to DNA Damage May Explain Different Cancer Susceptibility Between Small and Large Intestine

Experimental Biology and Medicine ◽

10.1177/153537020523000704 ◽

2005 ◽

Vol 230 (7) ◽

pp. 464-471 ◽

Cited By ~ 28

Author(s):

Mee Young Hong ◽

Nancy D. Turner ◽

Raymond J. Carroll ◽

Robert S. Chapkin ◽

Joanne R. Lupton

Keyword(s):

Dna Damage ◽

Large Intestine ◽

Dna Adducts ◽

Oxidative Dna Damage ◽

Cancer Susceptibility ◽

Immunohistochemical Analysis ◽

The United States ◽

Treatment Groups ◽

The Difference ◽

Small And Large Intestine

Although large intestine (LI) cancer is the second-leading cause of cancer-related deaths in the United States, small intestine (SI) cancer is relatively rare. Because oxidative DNA damage is one possible initiator of tumorigenesis, we investigated if the SI is protected against cancer because of a more appropriate response to oxidative DNA damage compared with the LI. Sixty rats were allocated to three treatment groups: 3% dextran sodium sulfate (DSS, a DNA-oxidizing agent) for 48 hrs, withdrawal (DSS for 48 hrs + DSS withdrawal for 48 hrs), or control (no DSS). The SI, compared with the LI, showed greater oxidative DNA damage (P < 0.001) as determined using a quantitative immunohistochemical analysis of 8-oxodeoxyguanosine (8-oxodG). The response to the DNA adducts in the SI was greater than in the LI. The increase of TdT–mediated dUTP-biotin nick end labeling (TUNEL)-positive apoptosis after DSS treatment was greater in the SI compared with the LI (P < 0.001), and there was a positive correlation (P = 0.031) between DNA damage and apoptosis in the SI. Morphologically, DSS caused an extensive loss of crypt structure shown in lower crypt height (P = 0.006) and the number of intact crypts (P = 0.0001) in the LI, but not in the SI. These data suggest that the SI may be more protected against cancer by having a more dynamic response to oxidative damage that maintains crypt morphology, whereas the response of the LI makes it more susceptible to loss of crypt architecture. These differential responses to oxidative DNA damage may contribute to the difference in cancer susceptibility between these two anatomic sites of the intestine.

Download Full-text

Mutational impact and signature of ionizing radiation

10.1101/2021.01.12.426324 ◽

2021 ◽

Author(s):

Jeonghwan Youk ◽

Hyun Woo Kwon ◽

Joonoh Lim ◽

Eunji Kim ◽

Ryul Kim ◽

...

Keyword(s):

Dna Damage ◽

Ionizing Radiation ◽

Mammalian Cells ◽

Single Cells ◽

Dna Double Strand Breaks ◽

Structural Variations ◽

Dna Damage And Repair ◽

Repair Processes ◽

Massive Number ◽

Chaotic Nature

AbstractWhole-genome sequencing (WGS) of human tumors and normal cells exposed to various carcinogens has revealed distinct mutational patterns that provide deep insights into the DNA damage and repair processes. Although ionizing radiation (IR) is conventionally known as a strong carcinogen, its genome-wide mutational impacts have not been comprehensively investigated at the single-nucleotide level. Here, we explored the mutational landscape of normal single-cells after exposure to the various levels of IR. On average, 1 Gy of IR exposure generated ∼16 mutational events with a spectrum consisting of predominantly small nucleotide deletions and a few characteristic structural variations. In ∼30% of the post-irradiated cells, complex genomic rearrangements, such as chromoplexy, chromothripsis, and breakage-fusion-bridge cycles, were resulted, indicating the stochastic and chaotic nature of DNA repair in the presence of the massive number of concurrent DNA double-strand breaks. These mutational signatures were confirmed in the genomes of 22 IR-induced secondary malignancies. With high-resolution genomic snapshots of irradiated cells, our findings provide deep insights into how IR-induced DNA damage and subsequent repair processes operate in mammalian cells.

Download Full-text

HTS-Compatible CometChip Enables Genetic Screening for Modulators of Apoptosis and DNA Double-Strand Break Repair

SLAS DISCOVERY Advancing Life Sciences ◽

10.1177/2472555220918367 ◽

2020 ◽

Vol 25 (8) ◽

pp. 906-922

Author(s):

Ian J. Tay ◽

James J. H. Park ◽

Anna L. Price ◽

Bevin P. Engelward ◽

Scott R. Floyd

Keyword(s):

Dna Damage ◽

Dna Damage Response ◽

High Throughput ◽

High Throughput Screening ◽

Strand Break ◽

Indirect Detection ◽

Dna Double Strand Breaks ◽

Physical Damage ◽

Dna Damage And Repair ◽

Damage Response

Dysfunction of apoptosis and DNA damage response pathways often drive cancer, and so a better understanding of these pathways can contribute to new cancer therapeutic strategies. Diverse discovery approaches have identified many apoptosis regulators, DNA damage response, and DNA damage repair proteins; however, many of these approaches rely on indirect detection of DNA damage. Here, we describe a novel discovery platform based on the comet assay that leverages previous technical advances in assay precision by incorporating high-throughput robotics. The high-throughput screening (HTS) CometChip is the first high-throughput-compatible assay that can directly detect physical damage in DNA. We focused on DNA double-strand breaks (DSBs) and utilized our HTS CometChip technology to perform a first-of-its-kind screen using an shRNA library targeting 2564 cancer-relevant genes. Conditions of the assay enable detection of DNA fragmentation from both exogenous (ionizing radiation) and endogenous (apoptosis) sources. Using this approach, we identified LATS2 as a novel DNA repair factor as well as a modulator of apoptosis. We conclude that the HTS CometChip is an effective assay for HTS to identify modulators of physical DNA damage and repair.

Download Full-text