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1.
BMC Bioinformatics ; 22(1): 450, 2021 Sep 21.
Artigo em Inglês | MEDLINE | ID: mdl-34548010

RESUMO

BACKGROUND: The liver plays a major role in the metabolic activation of xenobiotics (drugs, chemicals such as pollutants, pesticides, food additives...). Among environmental contaminants of concern, heterocyclic aromatic amines (HAA) are xenobiotics classified by IARC as possible or probable carcinogens (2A or 2B). There exist little information about the effect of these HAA in humans. While HAA is a family of more than thirty identified chemicals, the metabolic activation and possible DNA adduct formation have been fully characterized in human liver for only a few of them (MeIQx, PhIP, A[Formula: see text]C). RESULTS: We have developed a modeling approach in order to predict all the possible metabolites of a xenobiotic and enzymatic profiles that are linked to the production of metabolites able to bind DNA. Our prediction of metabolites approach relies on the construction of an enriched and annotated map of metabolites from an input metabolite.The pipeline assembles reaction prediction tools (SyGMa), sites of metabolism prediction tools (Way2Drug, SOMP and Fame 3), a tool to estimate the ability of a xenobotics to form DNA adducts (XenoSite Reactivity V1), and a filtering procedure based on Bayesian framework. This prediction pipeline was evaluated using caffeine and then applied to HAA. The method was applied to determine enzymes profiles associated with the maximization of metabolites derived from each HAA which are able to bind to DNA. The classification of HAA according to enzymatic profiles was consistent with their chemical structures. CONCLUSIONS: Overall, a predictive toxicological model based on an in silico systems biology approach opens perspectives to estimate the genotoxicity of various chemical classes of environmental contaminants. Moreover, our approach based on enzymes profile determination opens the possibility of predicting various xenobiotics metabolites susceptible to bind to DNA in both normal and physiopathological situations.


Assuntos
Adutos de DNA , Xenobióticos , Aminas , Teorema de Bayes , Carcinógenos , Humanos
2.
Int J Mol Sci ; 22(15)2021 Jul 28.
Artigo em Inglês | MEDLINE | ID: mdl-34360828

RESUMO

The environmental pollutant benzo[a]pyrene (BaP) is a human carcinogen that reacts with DNA after metabolic activation catalysed by cytochromes P450 (CYP) 1A1 and 1B1 together with microsomal epoxide hydrolase. The azo dye Sudan I is a potent inducer of CYP1A1/2. Here, Wistar rats were either treated with single doses of BaP (150 mg/kg bw) or Sudan I (50 mg/kg bw) alone or with both compounds in combination to explore BaP-derived DNA adduct formation in vivo. Using 32P-postlabelling, DNA adducts generated by BaP-7,8-dihydrodiol-9,10-epoxide were found in livers of rats treated with BaP alone or co-exposed to Sudan I. During co-exposure to Sudan I prior to BaP treatment, BaP-DNA adduct levels increased 2.1-fold in comparison to BaP treatment alone. Similarly, hepatic microsomes isolated from rats exposed to Sudan I prior to BaP treatment were also the most effective in generating DNA adducts in vitro with the activated metabolites BaP-7,8-dihydrodiol or BaP-9-ol as intermediates. DNA adduct formation correlated with changes in the expression and/or enzyme activities of CYP1A1, 1A2 and 1B1 in hepatic microsomes. Thus, BaP genotoxicity in rats in vivo appears to be related to the enhanced expression and/or activity of hepatic CYP1A1/2 and 1B1 caused by exposure of rats to the studied compounds. Our results indicate that the industrially employed azo dye Sudan I potentiates the genotoxicity of the human carcinogen BaP, and exposure to both substances at the same time seems to be hazardous to humans.


Assuntos
Benzo(a)pireno/toxicidade , Citocromo P-450 CYP1A1/metabolismo , Adutos de DNA/toxicidade , Fígado/efeitos dos fármacos , Microssomos Hepáticos/efeitos dos fármacos , Naftóis/toxicidade , Animais , Carcinógenos Ambientais/toxicidade , Corantes/toxicidade , Masculino , Ratos , Ratos Wistar
3.
Int J Mol Sci ; 22(15)2021 Jul 30.
Artigo em Inglês | MEDLINE | ID: mdl-34361004

RESUMO

This article reviews evidence suggesting that a common mechanism of initiation leads to the development of many prevalent types of cancer. Endogenous estrogens, in the form of catechol estrogen-3,4-quinones, play a central role in this pathway of cancer initiation. The catechol estrogen-3,4-quinones react with specific purine bases in DNA to form depurinating estrogen-DNA adducts that generate apurinic sites. The apurinic sites can then lead to cancer-causing mutations. The process of cancer initiation has been demonstrated using results from test tube reactions, cultured mammalian cells, and human subjects. Increased amounts of estrogen-DNA adducts are found not only in people with several different types of cancer but also in women at high risk for breast cancer, indicating that the formation of adducts is on the pathway to cancer initiation. Two compounds, resveratrol, and N-acetylcysteine, are particularly good at preventing the formation of estrogen-DNA adducts in humans and are, thus, potential cancer-prevention compounds.


Assuntos
Acetilcisteína/farmacologia , Carcinogênese/efeitos dos fármacos , Estradiol/farmacologia , Estrona/farmacologia , Quinonas/farmacologia , Resveratrol/farmacologia , Animais , Antioxidantes/farmacologia , Carcinogênese/genética , Adutos de DNA , Estradiol/toxicidade , Estrogênios/farmacologia , Estrogênios/toxicidade , Estrona/toxicidade , Humanos , Quinonas/toxicidade
4.
Toxicol Lett ; 351: 53-64, 2021 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-34454013

RESUMO

Benzo[a]pyrene(B[a]P) is a known human carcinogen. The ability of B[a]P to form stable DNA adducts has been repeatedly demonstrated. However, the relationship between DNA adduct formation and cell damage and its underlying molecular mechanisms are less well understood. In this study, we determined the cytotoxicity of benzo[a]pyrenediolepoxide, a metabolite of B[a]P, in human bronchial epithelial cells (BEAS-2B). The formation of BPDE-DNA adducts was quantified using a dot blot. DNA damage resulting from the formation of BPDE-DNA adducts was detected by chromatin immuneprecipitation sequencing (ChIP-Seq), with minor modifications, using specific antibodies against BPDE. In total, 1846 differentially expressed gene loci were detected between the treatment and control groups. The distribution of the BPDE-bound regions indicated that BPDE could covalently bind with both coding and non-coding regions to cause DNA damage. However, the majority of binding occurred at protein-coding genes. Furthermore, among the BPDE-bound genes, we found 16 protein-coding genes related to DNA damage repair. We explored the response to BPDE exposure at the transcriptional level using qRT-PCR and observed a strong inhibition of EIF4A3. We then established an EIF4A3 overexpression cell model and performed comet assays, which revealed that the levels of DNA damage in EIF4A3-overexpressing cells were lower than those in normal cells following BPDE exposure. This suggests that the BPDE-DNA adduct-induced reduction in EIF4A3 expression contributed to the DNA damage induced by BPDE exposure in BEAS-2B cells. These novel findings indicate that ChIP-Seq combined with BPDE specific antibody may be used for exploring the underlying mechanism of DNA adduct-induced genomic damage.


Assuntos
Benzo(a)pireno/toxicidade , RNA Helicases DEAD-box/metabolismo , Adutos de DNA , Dano ao DNA/efeitos dos fármacos , Células Epiteliais/efeitos dos fármacos , Fator de Iniciação 4A em Eucariotos/metabolismo , Linhagem Celular , Clonagem Molecular , RNA Helicases DEAD-box/genética , Fator de Iniciação 4A em Eucariotos/genética , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Mucosa Respiratória/citologia
5.
Mar Environ Res ; 170: 105434, 2021 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-34333338

RESUMO

Studies that aim to characterise the susceptibility of the ecologically relevant and non-model fish polar cod (Boreogadus saida) to model carcinogens are required. Polar cod were exposed under laboratory conditions for six months to control, 0.03 µg BaP/g fish/week and 0.3 µg BaP/g fish/week dietary benzo(a)pyrene (BaP), a reference carcinogen. The concentrations of the 3-OH-BaP bile metabolite and transcriptional responses of genes involved in DNA adduct recognition (xpc), helicase activity (xpd), DNA repair (xpf, rad51) and tumour suppression (tp53) were assessed after 0, 1, 3 and 6 months of exposure, alongside body condition indexes (gonadosomatic index, hepatosomatic index and condition factor). Micronuclei and nuclear abnormalities in blood and spleen, and liver histopathological endpoints were assessed at the end of the experiment. Fish grew steadily over the whole experiment and no mortality was recorded. The concentrations of 3-OH-BaP increased significantly after 1 month of exposure to the highest BaP concentration and after 6 months of exposure to all BaP concentrations showing the biotransformation of the mother compound. Nevertheless, no significant induction of gene transcripts involved in DNA damage repair or tumour suppression were observed at the selected sampling times. These results together with the absence of chromosomal damage in blood and spleen cells, the subtle increase in nuclear abnormalities observed in spleen cells and the low occurrence of foci of cellular alteration suggested that the exposure was below the threshold of observable effects. Taken together, the results showed that polar cod was not susceptible to carcinogenesis using the BaP exposure regime employed herein.


Assuntos
Carcinógenos , Gadiformes , Animais , Benzo(a)pireno/toxicidade , Bile , Adutos de DNA/farmacologia
6.
Chem Biol Interact ; 346: 109580, 2021 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-34280354

RESUMO

Dichloromethane (DCM), a widely used chlorinated solvent, is classified by IARC (2017) as probably carcinogenic to humans. Exposure to DCM has been associated with increased incidence of cholangiocarcinoma (CCA) in humans. This study aimed to investigate how DCM could contribute to CCA development by investigating the effects of DCM on DNA damage and cell transformation in cholangiocytes (MMNK-1) and on metastatic potential as measured by invasion and cell migration in malignant CCA cell lines (HuCCA-1 and RMCCA-1). MMNK-1 cells treated with the non-cytotoxic concentration of DCM (25 µM, 24 h) significantly increased the levels of mutagenic DNA adducts including 8-hydroxydeoxyguanosine, 8-OHdG, (1.84-fold, p < 0.01) and 8-nitroguanine (1.96-fold, p < 0.01) and enhanced cell transformation by 1.47-fold (p < 0.01). In addition, the expression of various genes involved in carcinogenesis, namely, NFE2L2 (antioxidative response), CXCL8 (inflammation), CDH1 (cell adhesion), MMP9 (tissue remodeling) and MKI67 (cell proliferation) were altered in cholangiocytes treated with DCM. When MMNK-1 cells were transformed by DCM, the expression of all the aforementioned genes was also increased. In malignant cell lines (HuCCA-1 and RMCCA-1), DCM treatment resulted in increased CXCL8 and MMP9 transcription and decreased CDH1 transcription accompanied by increased invasion and migration capabilities of these cells. Taken together, this study demonstrated that DCM exposure could be linked to the development of CCA.


Assuntos
Transformação Celular Neoplásica/efeitos dos fármacos , Dano ao DNA/efeitos dos fármacos , Cloreto de Metileno/toxicidade , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Colangiocarcinoma/metabolismo , Colangiocarcinoma/patologia , Adutos de DNA/análise , Adutos de DNA/metabolismo , Expressão Gênica/efeitos dos fármacos , Humanos , Interleucina-8/genética , Interleucina-8/metabolismo , Metaloproteinase 9 da Matriz/genética , Metaloproteinase 9 da Matriz/metabolismo , Cloreto de Metileno/química , Fator 2 Relacionado a NF-E2/genética , Fator 2 Relacionado a NF-E2/metabolismo , RNA Mensageiro/metabolismo
7.
Spectrochim Acta A Mol Biomol Spectrosc ; 262: 120096, 2021 Dec 05.
Artigo em Inglês | MEDLINE | ID: mdl-34214741

RESUMO

Pt(II) complex cis-[Pt(PEA)(OH2)2] X2, C-2 (where, PEA = 2-Pyridylethylamine and X  = ClO4- or NO3-) was synthesized by hydrolysis of cis-[Pt(PEA)Cl2] C-1. Glutathione (GSH) and DL-penicilamine (DL-pen) substituted complexes cis-[Pt(PEA)(GSH)],C-3 and cis-[Pt(PEA)DL-pen)]X C-4 were synthesized and characterized by spectroscopic methods. Kinetic studies were traced on complex C-2 with the thiols, GSH and DL-pen. Pt(II)-Sulfur adduct formation mechanisms of the substituted products C-3 and C-4 were established from the kinetic investigation. At pH 4.0, C-2 - thiols interactions follow two consecutive steps: the first step is dependent, and the second is independent of [thiol]. The association equilibrium constant (KE), substitution rate constants for both steps (k1 & k2), and activation parameters (ΔH‡ and ΔS‡) have been assessed to propose the mechanism. Agarose gel electrophoresis mobilization pattern of DNA with complexes was performed to visualize the interaction nature. CT-DNA and BSA binding activities of the complexes have been executed by electronic, fluorescence spectroscopy, and viscometric titration methods. Evaluation of thermodynamic parameters (ΔH0, ΔS0, and ΔG0) from BSA binding constants was executed to propose the driving forces of interaction between these species. A molecular docking study was performed to evaluate the binding mode of complexes with BDNA strands. Anticancer activity of the complexes C-1 to C-4 was explored on both A549 and HEp-2 cell lines, compared with approved anticancer drugs cisplatin, carboplatin, and oxaliplatin. All these complexes were tested by NBT assay on normal cell line skeletal muscle cells (L6 myotubes) to observe the adverse effects compared to recognized anticancer medications. The ultimate aim is to explore the role of anticancer agents on cell death mechanism, which has been performed by flow-cytometer on HEp-2 cell lines.


Assuntos
Antineoplásicos , Adutos de DNA , Antineoplásicos/farmacologia , Morte Celular , Cisplatino , Cinética , Simulação de Acoplamento Molecular
8.
J Chem Phys ; 154(18): 184101, 2021 May 14.
Artigo em Inglês | MEDLINE | ID: mdl-34241037

RESUMO

We examine the use of the truncated singular value decomposition and Tikhonov regularization in standard form to address ill-posed least squares problems Ax = b that frequently arise in molecular mechanics force field parameter optimization. We illustrate these approaches by applying them to dihedral parameter optimization of genotoxic polycyclic aromatic hydrocarbon-DNA adducts that are of interest in the study of chemical carcinogenesis. Utilizing the discrete Picard condition and/or a well-defined gap in the singular value spectrum when A has a well-determined numerical rank, we are able to systematically determine truncation and in turn regularization parameters that are correspondingly used to produce truncated and regularized solutions to the ill-posed least squares problem at hand. These solutions in turn result in optimized force field dihedral terms that accurately parameterize the torsional energy landscape. As the solutions produced by this approach are unique, it has the advantage of avoiding the multiple iterations and guess and check work often required to optimize molecular mechanics force field parameters.


Assuntos
Adutos de DNA/química , Análise dos Mínimos Quadrados , Hidrocarbonetos Policíclicos Aromáticos/química , Algoritmos
9.
J Chem Phys ; 154(17): 175102, 2021 May 07.
Artigo em Inglês | MEDLINE | ID: mdl-34241046

RESUMO

Polycyclic aromatic hydrocarbons (PAHs) are widely distributed in environments, and some of them are causative agents of human cancer. Previous studies concluded that benzo[a]pyrene-7,8-dione (BPQ), which is one kind of carcinogenic PAH metabolites, forms covalently bonded adducts with DNA, and the major adduct formed is a deoxyguanosine adduct. In this work, we investigate the interactions between BPQ and DNA molecules via first-principles calculations. We identify six possible DNA adducts with BPQ. In addition to the four adducts forming covalent bonds, there are two adducts bound purely by van der Waals (vdW) interactions. Remarkably, the two vdW-bound adducts have comparable, if not larger, binding energies as the covalent adducts. The results may help us gain more understanding of the interactions between PAH metabolites and DNA.


Assuntos
Benzopirenos/química , Adutos de DNA/química , Teoria da Densidade Funcional , Simulação de Dinâmica Molecular , Benzopirenos/metabolismo , Adutos de DNA/metabolismo , Estrutura Molecular
10.
Int J Mol Sci ; 22(11)2021 Jun 02.
Artigo em Inglês | MEDLINE | ID: mdl-34199457

RESUMO

Polycyclic aromatic hydrocarbons (PAHs) are chemical compounds comprised of carbon and hydrogen molecules in a cyclic arrangement. PAHs are associated with risks to human health, especially carcinogenesis. One form of exposure to these compounds is through ingestion of contaminated food, which can occur during preparation and processing involving high temperatures (e.g., grilling, smoking, toasting, roasting, and frying) as well as through PAHs present in the soil, air, and water (i.e., environmental pollution). Differently from changes caused by microbiological characteristics and lipid oxidation, consumers cannot sensorially perceive PAH contamination in food products, thereby hindering their ability to reject these foods. Herein, the occurrence and biological effects of PAHs were comprehensively explored, as well as analytical methods to monitor their levels, legislations, and strategies to reduce their generation in food products. This review updates the current knowledge and addresses recent regulation changes concerning the widespread PAHs contamination in several types of food, often surpassing the concentration limits deemed acceptable by current legislations. Therefore, effective measures involving different food processing strategies are needed to prevent and reduce PAHs contamination, thereby decreasing human exposure and detrimental health effects. Furthermore, gaps in literature have been addressed to provide a basis for future studies.


Assuntos
Carcinogênese/efeitos dos fármacos , Poluição Ambiental/efeitos adversos , Alimentos/efeitos adversos , Hidrocarbonetos Policíclicos Aromáticos/efeitos adversos , Benzopirenos/efeitos adversos , Carcinogênese/genética , Carvão Vegetal/efeitos adversos , Culinária , Adutos de DNA/efeitos adversos , Análise de Alimentos , Manipulação de Alimentos , Humanos
11.
Int J Radiat Biol ; 97(9): 1166-1180, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34259614

RESUMO

PURPOSE: DNA, the hereditary material of a human cell generally exists as Watson-Crick base paired double-stranded B-DNA. Studies suggest that DNA can also exist in non-B forms, such as four stranded G-quadruplexes (G4 DNA). Recently, our studies revealed that the regions of DNA that can fold into G-quadruplex structures are less sensitive to ionizing radiation (IR) compared to B-DNA. Importantly, we reported that the planar G-quartet of a G4 structure is shielded from radiation induced DNA breaks, while the single- and double-stranded DNA regions remained susceptible. Thus, in the present study, we investigate whether telomeric repeat DNA present at the end of telomere, known to fold into G4 DNA can protect from radiation induced damages including strand breaks, oxidation of purines and bulky adduct formation on DNA. MATERIALS AND METHODS: For plasmid irradiation assay, plasmids containing human telomeric repeat DNA sequence TTAGGG (0.8 kb or 1.8 kb) were irradiated with increasing doses of IR along with appropriate control plasmids and products were resolved on 1% agarose gel. Radioprotection was evaluated based on extent of conversion of supercoiled to nicked or linear forms of the DNA following irradiation. Formation of G-quadruplex structure on supercoiled DNA was evaluated based on circular dichroism (CD) spectroscopy studies. Cleavage of radiation induced oxidative damage and extent of formation of nicks was further evaluated using base and nucleotide excision repair proteins. RESULTS: Results from CD studies showed that the plasmid DNA harboring human telomeric repeats (TTAGGG) can fold into G-quadruplex DNA structures. Further, results showed that human telomeric repeat sequence when present on a plasmid can protect the plasmid DNA against IR induced DNA strand breaks, unlike control plasmids bearing random DNA sequence. CONCLUSIONS: Human telomeric repeat sequence when present on plasmids can fold into G-quadruplex DNA structures, and can protect the DNA against IR induced DNA strand breaks and oxidative damage. These results in conjunction with our previous studies suggest that telomeric repeat sequence imparts less sensitivity to IR and thus telomeres of chromosomes are protected from radiation.


Assuntos
Adutos de DNA/genética , Adutos de DNA/efeitos da radiação , Quadruplex G/efeitos da radiação , Raios gama/efeitos adversos , Estresse Oxidativo/genética , Estresse Oxidativo/efeitos da radiação , Telômero/genética , Sequência de Bases , Humanos , Telômero/efeitos da radiação
12.
Chemistry ; 27(57): 14263-14272, 2021 Oct 13.
Artigo em Inglês | MEDLINE | ID: mdl-34319608

RESUMO

Mitomycin C, (MC), an antitumor drug, is a DNA alkylating agent currently used in the clinics. Inert in its native form, MC is reduced to reactive mitosenes, which undergo nucleophilic attack by guanine or adenine bases in DNA to form monoadducts as well as interstrand crosslinks (ICLs). Although ICLs are considered the most cytotoxic lesions, the role of each individual adduct in the drug's cytotoxicity is still not fully understood. Synthetic routes have been developed to access modified oligonucleotides containing dG MC-monoadducts and dG-MC-dG ICL at a single position of their base sequences to investigate the biological effects of these adducts. However, until now, oligonucleotides containing monoadducts formed by MC at the adenine base had not been available, thus preventing the examination of the role played by these lesions in the toxicity of MC. Here, we present a route to access these substrates. Structural proof of the adducted oligonucleotides were provided by enzymatic digestion to nucleosides and high-resolution mass spectral analysis. Additionally, parent oligonucleotides containing a dG monoadduct and a dG-MC-dG ICL were also produced. The stability and physical properties of all substrates were compared via CD spectroscopy and UV melting temperature studies. Finally, virtual models were created to explore the conformational space and structural features of these MC-DNA complexes.


Assuntos
Adutos de DNA , Mitomicina , Adenina , Guanina , Oligonucleotídeos
13.
Nat Commun ; 12(1): 3338, 2021 06 07.
Artigo em Inglês | MEDLINE | ID: mdl-34099686

RESUMO

The versatile nucleotide excision repair (NER) pathway initiates as the XPC-RAD23B-CETN2 complex first recognizes DNA lesions from the genomic DNA and recruits the general transcription factor complex, TFIIH, for subsequent lesion verification. Here, we present a cryo-EM structure of an NER initiation complex containing Rad4-Rad23-Rad33 (yeast homologue of XPC-RAD23B-CETN2) and 7-subunit coreTFIIH assembled on a carcinogen-DNA adduct lesion at 3.9-9.2 Å resolution. A ~30-bp DNA duplex could be mapped as it straddles between Rad4 and the Ssl2 (XPB) subunit of TFIIH on the 3' and 5' side of the lesion, respectively. The simultaneous binding with Rad4 and TFIIH was permitted by an unwinding of DNA at the lesion. Translocation coupled with torque generation by Ssl2 and Rad4 would extend the DNA unwinding at the lesion and deliver the damaged strand to Rad3 (XPD) in an open form suitable for subsequent lesion scanning and verification.


Assuntos
Microscopia Crioeletrônica , Dano ao DNA , Reparo do DNA , Proteínas de Ligação a DNA/química , DNA/química , Proteínas de Saccharomyces cerevisiae/química , Saccharomyces cerevisiae/metabolismo , Fator de Transcrição TFIIH/química , Adutos de DNA/metabolismo , DNA Helicases/química , DNA Helicases/genética , Proteínas de Ligação a DNA/genética , Proteínas de Saccharomyces cerevisiae/genética , Fator de Transcrição TFIIH/genética
14.
J Biol Chem ; 297(1): 100868, 2021 07.
Artigo em Inglês | MEDLINE | ID: mdl-34119520

RESUMO

In a previous study, we showed that replication through the N1-methyl-deoxyadenosine (1-MeA) adduct in human cells is mediated via three different Polι/Polθ, Polη, and Polζ-dependent pathways. Based on biochemical studies with these Pols, in the Polι/Polθ pathway, we inferred a role for Polι in the insertion of a nucleotide (nt) opposite 1-MeA and of Polθ in extension of synthesis from the inserted nt; in the Polη pathway, we inferred that this Pol alone would replicate through 1-MeA; in the Polζ pathway, however, the Pol required for inserting an nt opposite 1-MeA had remained unidentified. In this study, we provide biochemical and genetic evidence for a role for Polλ in inserting the correct nt T opposite 1-MeA, from which Polζ would extend synthesis. The high proficiency of purified Polλ for inserting a T opposite 1-MeA implicates a role for Polλ-which normally uses W-C base pairing for DNA synthesis-in accommodating 1-MeA in a syn confirmation and forming a Hoogsteen base pair with T. The potential of Polλ to replicate through DNA lesions by Hoogsteen base pairing adds another novel aspect to Polλ's role in translesion synthesis in addition to its role as a scaffolding component of Polζ. We discuss how the action mechanisms of Polλ and Polζ could be restrained to inserting a T opposite 1-MeA and extending synthesis thereafter, respectively.


Assuntos
Monofosfato de Adenosina/análogos & derivados , DNA Polimerase beta/metabolismo , Replicação do DNA , Monofosfato de Adenosina/metabolismo , Pareamento de Bases , Linhagem Celular , Adutos de DNA/metabolismo , DNA Polimerase Dirigida por DNA/metabolismo , Humanos , Mutação
15.
Nucleic Acids Res ; 49(11): 6331-6346, 2021 06 21.
Artigo em Inglês | MEDLINE | ID: mdl-34096589

RESUMO

Cockayne syndrome (CS) is an autosomal recessive genetic disorder characterized by photosensitivity, developmental defects, neurological abnormalities, and premature aging. Mutations in CSA (ERCC8), CSB (ERCC6), XPB, XPD, XPG, XPF (ERCC4) and ERCC1 can give rise to clinical phenotypes resembling classic CS. Using a yeast two-hybrid (Y2H) screening approach, we identified LEO1 (Phe381-Ser568 region) as an interacting protein partner of full-length and C-terminal (Pro1010-Cys1493) CSB in two independent screens. LEO1 is a member of the RNA polymerase associated factor 1 complex (PAF1C) with roles in transcription elongation and chromatin modification. Supportive of the Y2H results, purified, recombinant LEO1 and CSB directly interact in vitro, and the two proteins exist in a common complex within human cells. In addition, fluorescently tagged LEO1 and CSB are both recruited to localized DNA damage sites in human cells. Cell fractionation experiments revealed a transcription-dependent, coordinated association of LEO1 and CSB to chromatin following either UVC irradiation or cisplatin treatment of HEK293T cells, whereas the response to menadione was distinct, suggesting that this collaboration occurs mainly in the context of bulky transcription-blocking lesions. Consistent with a coordinated interaction in DNA repair, LEO1 knockdown or knockout resulted in reduced CSB recruitment to chromatin, increased sensitivity to UVC light and cisplatin damage, and reduced RNA synthesis recovery and slower excision of cyclobutane pyrimidine dimers following UVC irradiation; the absence of CSB resulted in diminished LEO1 recruitment. Our data indicate a reciprocal communication between CSB and LEO1 in the context of transcription-associated DNA repair and RNA transcription recovery.


Assuntos
DNA Helicases/metabolismo , Enzimas Reparadoras do DNA/metabolismo , Reparo do DNA , Proteínas de Ligação a Poli-ADP-Ribose/metabolismo , Fatores de Transcrição/metabolismo , Cromatina/metabolismo , Adutos de DNA , Dano ao DNA , Células HEK293 , Células HeLa , Humanos , Mutagênicos/toxicidade , RNA/biossíntese , Fatores de Transcrição/química , Transcrição Genética
16.
Environ Sci Technol ; 55(13): 9024-9032, 2021 07 06.
Artigo em Inglês | MEDLINE | ID: mdl-34125507

RESUMO

Balkan endemic nephropathy (BEN) is a chronic tubulointerstitial nephropathy affecting residents of rural farming areas in many Balkan countries. Although it is generally believed that BEN is an environmental disease caused by multiple geochemical factors with much attention on aristolochic acids (AAs), its etiology remains controversial. In this study, we tested the hypothesis that environmental contamination and subsequent food contamination by polycyclic aromatic hydrocarbons (PAHs) and phthalate esters are AA toxicity factors and important to BEN development. We identified significantly higher concentrations of phenanthrene, anthracene, diethyl phthalate (DEP), dibutyl phthalate (DBP), and benzyl butyl phthalate (BBP) in both maize and wheat grain samples collected from endemic villages than from nonendemic villages. Other PAHs and phthalate esters were also detected at higher concentrations in the soil samples from endemic villages. Subsequent genotoxicity testing of cultured human kidney cells showed an alarming phenomenon that phenanthrene, DEP, BBP, and DBP can interact synergistically with AAs to form elevated levels of AA-DNA adducts, which are associated with both the nephrotoxicity and carcinogenicity of AAs, further increasing their disease risks. This study provides direct evidence that prolonged coexposure to these environmental contaminants via dietary intake may lead to greater toxicity and accelerated development of BEN.


Assuntos
Ácidos Aristolóquicos , Nefropatia dos Bálcãs , Hidrocarbonetos Policíclicos Aromáticos , Ácidos Aristolóquicos/análise , Ácidos Aristolóquicos/toxicidade , Nefropatia dos Bálcãs/induzido quimicamente , Nefropatia dos Bálcãs/epidemiologia , Península Balcânica , Adutos de DNA , Ésteres , Humanos , Ácidos Ftálicos , Hidrocarbonetos Policíclicos Aromáticos/toxicidade , Solo
17.
Zhonghua Yu Fang Yi Xue Za Zhi ; 55(5): 653-659, 2021 May 06.
Artigo em Chinês | MEDLINE | ID: mdl-34034407

RESUMO

Objective: To investigate the dose-response relationship between serum polycyclic aromatic hydrocarbon adducts and serum complement C3 and C4 levels among children from a city in East China. Methods: In September 2016, two boarding schools in the air pollution exposure area and the control area (beyond the upwind of 30 km in the air pollution exposure area) in a city in East China were selected as the research site, and the eligible school-age children were recruited as the research objects. A total of 273 children were included, including 163 in the exposure group and 110 in the control group. The annual air pollutant data (PM2.5, PM10 and NO2) of the two regions during the study period were collected. The exposure level of tobacco was evaluated by cotinine in urine. The levels of serum complement C3 and C4 were determined by automatic biochemical analyzer. The serum anti-7, 8, -dihydrodiol-9, 10-epoxide benzo[a]pyrene (BPDE)-albumin adduct levels were detected by ELISA. Linear regression model was used to explore the dose-response relationship between BPDE-albumin adducts and serum complement C3 and C4. Results: The age of 273 subjects was (13.67±0.37) years old, including 165 boys (60.4%). The average annual exposure levels of PM2.5, PM10 and NO2 and the level of serum BPDE-albumin adducts in the exposure group were higher than those in the control group (P<0.05). The results of linear regression model analysis showed that after adjusting age, sex, BMI z-score and urinary cotinine level, when the serum BPDE-albumin adduct level increased by 10%, the serum complement C4 level decreased by 1.2% (P=0.017). After adjusting age, BMI z-score and urinary cotinine level, for every 10% increase in serum BPDE-albumin adduct level in boys, the serum complement C4 level decreased by 1.68% (P=0.024). After adjusting age, sex and BMI z-score, the levels of serum complement C3 and C4 decreased by 1.31% and 3.57% respectively for every 10% increase in serum BPDE-albumin adducts among children in the urinary cotinine detection group (P<0.05). Conclusion: There is a significant dose-response relationship between serum BPDE-albumin adducts and the complement C4 among children.


Assuntos
Poluentes Atmosféricos , Hidrocarbonetos Policíclicos Aromáticos , Adolescente , Benzo(a)pireno , China , Adutos de DNA , Feminino , Humanos , Masculino , Albumina Sérica/análise
18.
J Chem Inf Model ; 61(5): 2313-2327, 2021 05 24.
Artigo em Inglês | MEDLINE | ID: mdl-33977716

RESUMO

Human exposure to aromatic amines (AAs) can result in carcinogenic DNA adducts. To complement previous work geared toward understanding the mutagenicity of AA-derived adducts, which has almost exclusively studied (monoadducted) DNA containing a single lesion, the present work provides the first in-depth comparison of the structure of monoadducted and diadducted DNA duplexes. Specifically, molecular dynamics (MD) simulations were initially performed on DNA containing the nonmutagenic single-ringed N-(deoxyguanosin-8-yl)-aniline (ANdG) or the mutagenic four-ringed N-(deoxyguanosin-8-yl)-1-aminopyrene (APdG) lesion at G1, G2, or G3 in the AA deletion hotspot (5'-G1G2CG3CC) in the anti or syn glycosidic orientation (B/S duplex conformation). Subsequently, diadducted strands were assessed that span each combination of damaged sites (G1G2 (nearest neighbors), G2G3 (next-nearest neighbors), and G1G3 (two intervening nucleotides)) and anti/syn lesion glycosidic orientations. Despite other N-linked C8-dG adducts exhibiting sequence dependence conformational heterogeneity, a single ANdG or APdG lesion induces helical conformational homogeneity that is exclusively controlled by aryl moiety size. However, the preferred damaged DNA conformation can change upon the addition of a second adduct depending on lesion separation, with neighboring lesions stabilizing a nonmutagenic conformation and next-nearest damaged sites stabilizing a promutagenic conformation regardless of adduct size. As a result, diadducted DNA is found to adopt conformations that are unfavored for the corresponding monoadducted system, pointing to differential replication and repair outcomes for diadducted DNA compared to those for monoadducted DNA. Thus, although the toxicity of monoadducted DNA is most significantly dictated by lesion size, the toxicity can increase or decrease upon a second damaging event depending on lesion size and relative position. Overall, our work adds the number of lesions and their spatial separation to the growing list of factors that determine the structure and biological outcomes of adducted DNA.


Assuntos
Adutos de DNA , Simulação de Dinâmica Molecular , Aminas/toxicidade , DNA , Humanos , Conformação de Ácido Nucleico
19.
Chemosphere ; 274: 129991, 2021 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-33979922

RESUMO

Exposure to endogenous and exogenous factors can result in the formation of a wide variety of DNA adducts, and these may lead to gene mutations, thereby contributing to the development of cancer. DNA adductomics, a novel tool for exposomics, aims to detect the totality of DNA adducts. Liquid chromatography-high resolution mass spectrometry (LC-HRMS) is the state-of-the-art method for DNA adductomic analysis, although its high cost has limited widespread use. In this study, we compared the analytical performance between HRMS and the more popular/accessible triple-quadrupole MS (QqQ-MS). We initially developed and optimized a hybrid quadrupole-linear ion trap-orbitrap MS (Q-LIT-OT-MS) method, considering the detection of both purine and pyrimidine adducts. LC-Q-LIT-OT-MS and LC-QqQ-MS methods were compared by non-targeted screening of formaldehyde-induced DNA adducts. Using the results from Q-LIT-OT-MS as the gold standard, QqQ-MS successfully detected 12 out of 18 formaldehyde-induced DNA adducts/inter-strand crosslinks (ICLs). QqQ-MS however also produced nine false-positive results owing to the inherent instrumental mass resolution limits. To discriminate the false-positive results from the accurate ones, we firstly introduced a statistical analysis, partial least squares-discriminant analysis, which efficiently excluded the false signals. Six DNA adducts/ICLs were not detected by QqQ-MS, due to insufficient sensitivity. This could be overcome by employing a selected reaction monitoring scan mode with multiple injections. Overall, this study demonstrated that high resolution may not be a strict requirement for MS-based DNA adductomics. LC-QqQ-MS with statistical analysis, could also provide a comparable performance as HRMS for pre-screening purposes.


Assuntos
Adutos de DNA , Espectrometria de Massas em Tandem , Cromatografia Líquida
20.
Food Chem Toxicol ; 153: 112253, 2021 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-34015424

RESUMO

Estragole and anethole are secondary metabolites occurring in a variety of commonly used herbs like fennel, basil, and anise. Estragole is genotoxic and carcinogenic in rodents, which depends on the formation of 1'-sulfoxyestragole after hydroxylation and subsequent sulfoconjugation catalyzed by CYP and SULT, respectively. It was hypothesized recently that anethole may be bioactivated via the same metabolic pathways. Incubating estragole with hepatic S9-fractions from rats and humans, specific adducts with hemoglobin (N-(isoestragole-3-yl)-valine, IES-Val) and DNA (isoestragole-2'-deoxyguanosine and isoestragole-2'-deoxyadenosine) were formed. An isotope-dilution technique was developed for the quantification of IES-Val after cleavage with fluorescein isothiocyanate (FITC) according to a modified Edman degradation. The same adducts, albeit at lower levels, were also detected in reactions with anethole, indicating the formation of 3'-hydroxyanethole and the reactive 3'-sulfoxyanethole. Finally, we conducted a pilot investigation in which IES-Val levels in human blood were determined during and after the consumption of an estragole- and anethole-rich fennel tea for four weeks. A significant increase of IES-Val levels was observed during the consumption phase and followed by a continuous decrease during the washout period. IES-Val may be used to monitor the internal exposure to the common reactive genotoxic metabolites of estragole and anethole, 1'-sulfoxyestragole and 3'-sulfoxyanethole, respectively.


Assuntos
Derivados de Alilbenzenos/toxicidade , Anisóis/toxicidade , Adutos de DNA/química , Foeniculum/química , Hemoglobinas/química , Derivados de Alilbenzenos/metabolismo , Animais , Anisóis/metabolismo , Bebidas/análise , Biomarcadores/sangue , Humanos , Ratos
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