-
Environmental and Molecular Mutagenesis Feb 2020DNA adducts of carcinogenic polycyclic aromatic hydrocarbons (PAHs) play a critical role in the etiology of gastrointestinal tract cancers in humans and other species...
DNA adducts of carcinogenic polycyclic aromatic hydrocarbons (PAHs) play a critical role in the etiology of gastrointestinal tract cancers in humans and other species orally exposed to PAHs. Yet, the precise localization of PAH-DNA adducts in the gastrointestinal tract, and the long-term postmortem PAH-DNA adduct stability are unknown. To address these issues, the following experiment was performed. Mice were injected intraperitoneally with the PAH carcinogen benzo[a]pyrene (BP) and euthanized at 24 h. Tissues were harvested either at euthanasia (0 time), or after 4, 8, 12, 24, 48, and 168 hr (7 days) of storage at 4°C. Portions of mouse tissues were formalin-fixed, paraffin-embedded, and immunohistochemically (IHC) evaluated by incubation with r7,t8-dihydroxy-t-9,10-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene (BPDE)-DNA antiserum and H-scoring. The remaining tissues were frozen, and DNA was extracted and assayed for the r7,t8,t9-trihydroxy-c-10-(N -deoxyguanosyl)-7,8,9,10-tetrahydrobenzo[a]pyrene (BPdG) adduct using two quantitative assays, the BPDE-DNA chemiluminescence immunoassay (CIA), and high-performance liquid chromatography electrospray ionization tandem mass spectrometry (HPLC-ES-MS/MS). By IHC, which required intact nuclei, BPdG adducts were visualized in forestomach basal cells, which included gastric stem cells, for up to 7 days. In proximal small intestine villus epithelium BPdG adducts were visualized for up to 12 hr. By BPDE-DNA CIA and HPLC-ES-MS/MS, both of which used DNA for analysis and correlated well (P= 0.0001), BPdG adducts were unchanged in small intestine, forestomach, and lung stored at 4°C for up to 7 days postmortem. In addition to localization of BPdG adducts, this study reveals the feasibility of examining PAH-DNA adduct formation in wildlife species living in colder climates. Environ. Mol. Mutagen. 61:216-223, 2020. © 2019 Wiley Periodicals, Inc.
Topics: Animals; Benzo(a)pyrene; Carcinogens, Environmental; Chromatography, High Pressure Liquid; DNA Adducts; Intestine, Small; Luminescent Measurements; Male; Mice; Stomach; Tandem Mass Spectrometry; Tissue Distribution
PubMed: 31569280
DOI: 10.1002/em.22337 -
Chemical Research in Toxicology Aug 2020The human intestine is host to a vast microbial community: the gut microbiome (GM). The GM has been considered as a key modulator of human health in the past decade. In...
The human intestine is host to a vast microbial community: the gut microbiome (GM). The GM has been considered as a key modulator of human health in the past decade. In particular, several studies have supported that altered GM is associated with cancer, such as colorectal cancer, adenocarcinoma, and pancreatic cancer. The formation of a DNA adduct is one of the key events in carcinogenesis, and whether GM can influence DNA adducts has yet to be examined. This study analyzed 10 DNA adducts (N-Me-dG, N-Me-dA, N-Et-dG, OH-Me-dG, OH-Me-dA, N-EtD-dG, O-Me-dG, 1,N-ε-dG, 8-oxo-dG, and 5-Cl-dC), attributed to various endogenous processes and physiological stressors, using highly sensitive LC-MS/MS in germ-free (GF) and conventionally raised (CONV-R) mice. Our results showed that significant differences in specific DNA adducts appeared in liver, colon, and small intestine samples between GF and CONV-R mice. The differences in adduct levels may indicate that GM can locally or systemically regulate endogenous processes including neutrophil bactericidal activity (represented by 5-Cl-dC), lipid peroxidation (1,N-ε-dG), oxidative stress generation (8-oxo-dG), and endogenous aldehyde metabolism (OH-Me-dA). Further studies are warranted to elucidate how the GM influences endogenous process, DNA damage, and the risks of developing cancer.
Topics: Animals; Carcinogenesis; Carcinogens; DNA Adducts; DNA Damage; Female; Gastrointestinal Microbiome; Mice; Mice, Inbred C57BL; Molecular Structure
PubMed: 32677427
DOI: 10.1021/acs.chemrestox.0c00142 -
Journal of Agricultural and Food... Aug 2017Ochratoxin A (OTA) is a fungal toxin that is considered to be a potent kidney carcinogen in rodent models. The toxin produces double strand breaks and has a propensity... (Review)
Review
Ochratoxin A (OTA) is a fungal toxin that is considered to be a potent kidney carcinogen in rodent models. The toxin produces double strand breaks and has a propensity for deletions, single-base substitutions, and insertions. The toxin reacts covalently with DNA to afford a C8-2'-deoxyguanosine carbon-linked adduct (OT-dG) as the major lesion in animal tissues. Incorporation of model C-linked C8-aryl-dG adducts into the G3 site of the NarI sequence demonstrates a tendency to induce base substitutions and deletion mutations in primer extension assays using model polymerases. The degree of misincorporation induced by the C-linked C8-dG adducts correlates with an ability to adopt the promutagenic syn conformation within the NarI duplex as predicted by molecular dynamics (MD) simulations. MD simulations of the OT-dG adduct within the NarI duplex predict an even greater degree of conformational flexibility, suggesting enhanced in vitro mutagenicity compared to the simpler model C-linked C8-dG adducts. Together these findings support the role of OT-dG in promoting OTA-mediated mutagenicity and carcinogenicity in animal studies.
Topics: Animals; DNA Adducts; Deoxyguanosine; Humans; Mutagenesis; Mutagens; Nucleic Acid Conformation; Ochratoxins
PubMed: 28830149
DOI: 10.1021/acs.jafc.6b03897 -
Food and Chemical Toxicology : An... Jun 2020As one of the most widespread environmental pollutants, benzo[α]pyrene is metabolized to diol epoxides and then covalently breaks the initial DNA base pairs, which has...
As one of the most widespread environmental pollutants, benzo[α]pyrene is metabolized to diol epoxides and then covalently breaks the initial DNA base pairs, which has been closely related to the occurrence and development of many human cancers. High fidelity DNA polymerases play an extremely important role in maintaining the reliability or fidelity of nucleic acid replication, which is generally blocked by BP adducts. To reveal the blocking mechanism of BP, two comparative molecular dynamics simulations were performed for the thermophilic Bacillus stearothermophilus DNA polymerase I large fragment (BF) complexes with normal and BP-bound DNA duplexes. The results of global conformational changes and molecular interactions show that the association of BP leads to the rearrangement of intramolecular hydrogen bonds, impairing the molecular recognition between the polymerase and the DNA duplex. It is also found that the conformation of DNA duplex is distorted, accompanied by an increase in molecular overall rigidity. In terms of possible blocking mechanisms, the BP moiety perfectly integrates itself into the base-paired environment in a special vertical conformation and occupies the space required for the incoming nucleotide. This work provides useful dynamics and structural information for understanding the toxic effect of BP on DNA replication at atomic level.
Topics: Benzo(a)pyrene; DNA; DNA Adducts; Deoxyguanosine; Humans; Hydrogen Bonding; Reproducibility of Results
PubMed: 32272200
DOI: 10.1016/j.fct.2020.111325 -
Talanta May 2018Bisphenol A (BPA) is a widely used additive in the plastic industry and has been reported to have genotoxicity. A hypothesis that BPA may enhance breast cancer risk...
Bisphenol A (BPA) is a widely used additive in the plastic industry and has been reported to have genotoxicity. A hypothesis that BPA may enhance breast cancer risk through the formation of its metabolic intermediate or DNA adduct has been proposed. In this study, breast cancer cell MCF-7 was cultured and the cellular DNA was extracted from the cells. The adducts of bisphenol A 3,4-quinone (BPAQ) with 2'-deoxyguanosine (dG), calf thymus DNA and MCF-7 cell DNA were investigated. DNA adducts were characterized by using electrospray ionization Orbitrap high-resolution mass spectrometry and tandem mass spectrometry. The BPA-DNA adducts of BPAQ with dG, calf thymus and MCF-7 cell DNA were identified as 3-hydroxy-bisphenol A-N7-guanine (3-OH-BPA-N7Gua). The MS/MS fragmentation pathway of 3-OH-BPA-N7Gua was proposed based on obtained accurate mass data. BPA quinone metabolites can react with MCF-7 cell DNA in vitro. The findings provide evidence that BPA might covalently bind to DNA in MCF-7 cells mediated by quinone metabolites, which may increase our understanding of health risk associated with BPA exposure.
Topics: Animals; Benzhydryl Compounds; Benzoquinones; Cattle; DNA; DNA Adducts; DNA, Neoplasm; Deoxyguanosine; Endocrine Disruptors; Humans; MCF-7 Cells; Phenols; Spectrometry, Mass, Electrospray Ionization; Tandem Mass Spectrometry
PubMed: 29501196
DOI: 10.1016/j.talanta.2018.02.037 -
Analytical Chemistry Aug 2022DNA methylation can occur naturally or be induced by various environmental and chemotherapeutic agents. The regioisomeric - and -methyldeoxyadenosine (1mdA and 6mdA,...
DNA methylation can occur naturally or be induced by various environmental and chemotherapeutic agents. The regioisomeric - and -methyldeoxyadenosine (1mdA and 6mdA, respectively) represent an important class of methylated DNA adducts. In this study, we developed a shuttle vector- and next-generation sequencing-based assay to quantitatively assess the effects of 1mdA and 6mdA on the accuracy and efficiency of DNA transcription. Our results revealed that 1mdA can induce multiple types of mutant transcripts and strongly inhibit DNA transcription, whereas 6mdA is a nonmutagenic DNA adduct that can exhibit a subtle but significant inhibitory effect on DNA transcription in vitro and in human cells. Moreover, our results demonstrated that the transcription-coupled nucleotide excision repair pathway is dispensable for the removal of 1mdA and 6mdA from the template DNA strand in human cells. These findings provided new important insights into the functional interplay between DNA methylation modifications and transcription in mammalian cells.
Topics: Animals; DNA; DNA Adducts; DNA Repair; Deoxyadenosines; High-Throughput Nucleotide Sequencing; Humans; Mammals; Transcription, Genetic
PubMed: 35924299
DOI: 10.1021/acs.analchem.2c01764 -
The Journal of Biological Chemistry 2021Etheno (ε)-adducts, e.g., 1,N-ε-guanine (1,N-ε-G) and 1,N-ε-adenine (1,N-ε-A), are formed through the reaction of DNA with metabolites of vinyl compounds or with...
Etheno (ε)-adducts, e.g., 1,N-ε-guanine (1,N-ε-G) and 1,N-ε-adenine (1,N-ε-A), are formed through the reaction of DNA with metabolites of vinyl compounds or with lipid peroxidation products. These lesions are known to be mutagenic, but it is unknown how they lead to errors in DNA replication that are bypassed by DNA polymerases. Here we report the structural basis of misincorporation frequencies across from 1,N-ε-G by human DNA polymerase (hpol) η. In single-nucleotide insertions opposite the adduct 1,N-ε-G, hpol η preferentially inserted dGTP, followed by dATP, dTTP, and dCTP. This preference for purines was also seen in the first extension step. Analysis of full-length extension products by LC-MS/MS revealed that G accounted for 85% of nucleotides inserted opposite 1,N-ε-G in single base insertion, and 63% of bases inserted in the first extension step. Extension from the correct nucleotide pair (C) was not observed, but the primer with A paired opposite 1,N-ε-G was readily extended. Crystal structures of ternary hpol η insertion-stage complexes with nonhydrolyzable nucleotides dAMPnPP or dCMPnPP showed a syn orientation of the adduct, with the incoming A staggered between adducted base and the 5'-adjacent T, while the incoming C and adducted base were roughly coplanar. The formation of a bifurcated H-bond between incoming dAMPnPP and 1,N-ε-G and T, compared with the single H-bond formed between incoming dCMPnPP and 1,N-ε-G, may account for the observed facilitated insertion of dGTP and dATP. Thus, preferential insertion of purines by hpol η across from etheno adducts contributes to distinct outcomes in error-prone DNA replication.
Topics: Chromatography, Liquid; Crystallography, X-Ray; DNA Adducts; DNA-Directed DNA Polymerase; Deoxyguanosine; Humans; Tandem Mass Spectrometry
PubMed: 33839151
DOI: 10.1016/j.jbc.2021.100642 -
The Journal of Biological Chemistry Jun 2019Aside from abasic sites and ribonucleotides, the DNA adduct -methyl deoxyguanosine ( -CH dG) is one of the most abundant lesions in mammalian DNA. Because -CH dG is...
Aside from abasic sites and ribonucleotides, the DNA adduct -methyl deoxyguanosine ( -CH dG) is one of the most abundant lesions in mammalian DNA. Because -CH dG is unstable, leading to deglycosylation and ring-opening, its miscoding potential is not well-understood. Here, we employed a 2'-fluoro isostere approach to synthesize an oligonucleotide containing an analog of this lesion ( -CH 2'-F dG) and examined its miscoding potential with four Y-family translesion synthesis DNA polymerases (pols): human pol (hpol) η, hpol κ, and hpol ι and Dpo4 from the archaeal thermophile We found that hpol η and Dpo4 can bypass the -CH 2'-F dG adduct, albeit with some stalling, but hpol κ is strongly blocked at this lesion site, whereas hpol ι showed no distinction with the lesion and the control templates. hpol η yielded the highest level of misincorporation opposite the adduct by inserting dATP or dTTP. Moreover, hpol η did not extend well past an -CH 2'-F dG:dT mispair. MS-based sequence analysis confirmed that hpol η catalyzes mainly error-free incorporation of dC, with misincorporation of dA and dG in 5-10% of products. We conclude that -CH 2'-F dG and, by inference, -CH dG have miscoding and mutagenic potential. The level of misincorporation arising from this abundant adduct can be considered as potentially mutagenic as a highly miscoding but rare lesion.
Topics: DNA Adducts; DNA Damage; DNA Repair; DNA Replication; DNA-Directed DNA Polymerase; Deoxyguanosine; Humans; Molecular Structure
PubMed: 31101656
DOI: 10.1074/jbc.RA119.008986 -
Free Radical Biology & Medicine Nov 2019Human individuals are continually exposed to various exogenous and endogenous reactive electrophiles, which readily react with nucleophilic biomacromolecules, such as... (Review)
Review
Human individuals are continually exposed to various exogenous and endogenous reactive electrophiles, which readily react with nucleophilic biomacromolecules, such as protein, and form a variety of covalent adducts. The covalent modifications of protein are thought to be involved in various physiological and pathological processes. Recently, the "adductome", a new concept that represents the totality of covalent adducts bound to nucleophilic biomolecules, has been offered as a useful technique for characterizing essentially all reactive electrophilic compounds in biological samples. The primary advantage of this approach is that non-targeted comprehensive analysis can readily be extended to investigate covalent adduct pattern of different situation of exposure and thereby makes it possible to detect/identify not only known but also unknown adducts. In this review, we provide a summary of the concept and methodology of protein adductomics, especially focusing on redox protein adductomics.
Topics: Aldehydes; Cell Membrane; DNA Adducts; Eukaryotic Cells; Hemoglobins; Humans; Lipids; Mass Spectrometry; Oxidation-Reduction; Oxidative Stress; Protein Interaction Mapping; Protein Processing, Post-Translational; Serum Albumin, Human; Signal Transduction
PubMed: 30853395
DOI: 10.1016/j.freeradbiomed.2019.02.034 -
Bulky DNA adduct levels in normal-appearing colon mucosa, and the prevalence of colorectal adenomas.Biomarkers : Biochemical Indicators of... Dec 2018Examine the association between bulky DNA adduct levels in colon mucosa and colorectal adenoma prevalence, and explore the correlation between adduct levels in...
PURPOSE
Examine the association between bulky DNA adduct levels in colon mucosa and colorectal adenoma prevalence, and explore the correlation between adduct levels in leukocytes and colon tissue.
METHODS
Bulky DNA adduct levels were measured using P-postlabelling in biopsies of normal-appearing colon tissue and blood donated by 202 patients. Multivariable logistic regression was used to examine associations between DNA adducts, and interactions of DNA adduct-DNA repair polymorphisms, with the prevalence of colorectal adenomas. Correlation between blood and tissue levels of DNA adducts was evaluated using Spearman's correlation coefficient.
RESULTS
An interaction between bulky DNA adduct levels and XPA rs1800975 on prevalence of colorectal adenoma was observed. Among individuals with lower DNA repair activity, increased DNA adduct levels were associated with increased colorectal adenoma prevalence (OR = 1.41 per SD increase, 95%CI: 0.92-2.18). Conversely, among individuals with normal DNA activity, an inverse association was observed (OR = 0.60 per SD increase, 95%CI: 0.34-1.07). Blood and colon DNA adduct levels were inversely correlated (ρ = -0.20).
CONCLUSIONS
Among genetically susceptible individuals, higher bulky DNA adducts in the colon was associated with the prevalence of colorectal adenomas. The inverse correlation between blood and colon tissue measures demonstrates the importance of quantifying biomarkers in target tissues.
Topics: Adenoma; Adult; Colorectal Neoplasms; DNA Adducts; Female; Genetic Predisposition to Disease; Humans; Intestinal Mucosa; Male; Middle Aged; Polymorphism, Genetic; Prevalence; Xeroderma Pigmentosum Group A Protein
PubMed: 29871536
DOI: 10.1080/1354750X.2018.1485055