Duplex sequencing identifies genomic features that determine susceptibility to benzo(a)pyrene-induced in vivo mutations
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ABSTRACT: Supplementary Information
The online version contains supplementary material available at 10.1186/s12864-022-08752-w.
SUBMITTER: LeBlanc D
PROVIDER: S-EPMC9331077 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Understanding the effects of environmental exposures on germline mutation rates has been a decades-long pursuit in genetics. We used next-generation sequencing and comparative genomic hybridization arrays to investigate genome-wide mutations in the offspring of male mice exposed to benzo(a)pyrene (BaP), a common environmental pollutant. We demonstrate that offspring developing from sperm exposed during the mitotic or post-mitotic phases of spermatogenesis have significantly more de novo single nucleotide variants (1.8-fold; P?<?0.01) than controls. Both phases of spermatogenesis are susceptible to the induction of heritable mutations, although mutations arising from post-fertilization events are more common after post-mitotic exposure. In addition, the mutation spectra in sperm and offspring of BaP-exposed males are consistent. Finally, we report a significant increase in transmitted copy number duplications (P?=?0.001) in BaP-exposed sires. Our study demonstrates that germ cell mutagen exposures induce genome-wide mutations in the offspring that may be associated with adverse health outcomes.
| S-EPMC6586636 | biostudies-literature
Project description:De novo mutations are implicated in a variety of genetic diseases and arise primarily in the male germline. We investigated whether male germ cells have unique mechanisms for spontaneous or chemically-induced mutation relative to somatic cells using the MutaMouse model. We recovered lacZ transgenes from sperm 42 days after a 28-day exposure to benzo(a)pyrene (BaP, 100 mg/kg/day) to assess mutations arising in dividing spermatogonia. BaP caused a 3.4-fold increase in lacZ mutant frequency over controls which increased to 4.1-fold after clonal correction. We then used next generation sequencing to compare the spontaneous and BaP-induced mutation spectra in sperm and bone marrow. The spontaneous spectrum in sperm had significantly more G:C to A:T transitions and fewer mutations at A:T basepairs than bone marrow. BaP predominantly induced G:C to T:A transversions in both cell types, and both were enriched for mutations at CpG dinucleotides. However, BaP induced significantly more deletions in sperm, but more G:C to A:T transitions and G:C to C:G transversions in bone marrow. Differences in error-prone translesion DNA synthesis polymerases may underlie the observed spectrum differences between sperm and bone marrow. These findings suggest that mutations in sperm can arise via mechanisms that are unique to male germ cells.
| S-EPMC5103183 | biostudies-literature
Project description:Polycyclic aromatic hydrocarbons like benzo[a]pyrene (BaP) are ubiquitous environmental contaminants formed during incomplete combustion of organic materials. Our prior work showed that transplacental exposure to BaP depletes ovarian follicles and increases prevalence of epithelial ovarian tumors later in life. We used the MutaMouse transgenic rodent model to address the hypothesis that ovarian mutations play a role in tumorigenesis caused by prenatal exposure to BaP. Pregnant MutaMouse females were treated with 0, 10, 20, or 40 mg/(kg day) BaP orally on gestational days 7-16, covering critical windows of ovarian development. Female offspring were euthanized at 10 weeks of age; some ovaries with oviducts were processed for follicle counting; other ovaries/oviducts and bone marrow were processed for determination of lacZ mutant frequency (MF). Mutant plaques were pooled within dose groups and sequenced to determine the mutation spectrum. BaP exposure caused highly significant dose-related decreases in ovarian follicles and increases in ovarian/oviductal and bone marrow mutant frequencies at all doses. Absence of follicles, cell packets, and epithelial tubular structures were observed with 20 and 40 mg/(kg day) BaP. Depletion of ovarian germ cells was inversely associated with ovarian MF. BaP induced primarily G > T and G > C transversions and deletions in ovaries/oviducts and bone marrow cells and produced a mutation signature highly consistent with that of tobacco smoking in human cancers. Overall, our results show that prenatal BaP exposure significantly depletes ovarian germ cells, causes histopathological abnormalities, and increases the burden of ovarian/oviductal mutations, which may be involved in pathogenesis of epithelial ovarian tumors. Environ. Mol. Mutagen. 60:410-420, 2019. © 2018 Her Majesty the Queen in Right of Canada.
| S-EPMC6615722 | biostudies-literature
Project description:Persistent and ubiquitous organic pollutants, such as the polycyclic aromatic hydrocarbon benzo[⍺]pyrene (BaP), represent a major threat to aquatic organisms and human health. Beside some well-documented adverse effects on the development and reproduction of aquatic organisms, BaP was recently shown to affect fish bone formation and skeletal development through mechanisms that remain poorly understood. In this work, several bone-related in vivo assays were used in zebrafish to evaluate the osteotoxic effects of BaP during bone development and regeneration. Exposure to BaP for 3 days induced a dose-dependent reduction of the size of the opercular bone in 6 days post-fertilization (dpf) larvae, and affected osteoblast maturation as observed by the expression of the mature marker, osteocalcin. Exposure to BaP for 27 days affected the development of the axial skeleton and increased the incidence and severity of skeletal deformities. During bone regeneration, BaP affected the mineralization of newly formed fin rays and scales, while it impaired fin ray patterning and scale shape, through mechanisms that may involve an imbalance of bone remodeling. Transcriptomic and gene expression analyses indicate that BaP induced the activation of xenobiotic and metabolic pathways, while negatively impacting extracellular matrix formation and organization. Interestingly, BaP exposure positively regulated inflammation markers in 6 dpf larvae and increased neutrophil recruitment. A direct interaction between neutrophils and bone extracellular matrix or bone forming cells was observed in vivo, suggesting a role for neutrophils in the mechanisms underlying BaP osteotoxicity. Our work provides novel data on the cellular and molecular players involved in BaP osteotoxicity and brings new insights into a possible role for neutrophils in inflammatory bone reduction.
2024-04-29 | GSE174529 | GEO
Project description:Benzo[a]pyrene (B[a]P), a major human carcinogen in combustion products such as cigarette smoke and diesel exhaust, is metabolically activated into DNA-reactive metabolites via three different enzymatic pathways. The pathways are the anti-(+)-benzo[a]pyrene 7,8-diol 9,10-epoxide pathway (P450/epoxide hydrolase catalyzed) (B[a]PDE), the benzo[a]pyrene o-quinone pathway (aldo ketose reductase (AKR) catalyzed) and the B[a]P radical cation pathway (P450 peroxidase catalyzed). We used a yeast p53 mutagenesis system to assess mutagenesis by B[a]P radical cations. Because radical cations are short-lived, they were generated in situ by reacting B[a]P with cumene hydroperoxide (CuOOH) and horse radish peroxidase (HRP) and then monitoring the generation of the more stable downstream products, B[a]P-1,6-dione and B[a]P-3,6-dione. On the basis of B[a]P-1,6 and 3,6-dione formation, approximately 4 ?M of radical cation was generated. In the mutagenesis assays, the radical cations produced in situ showed a dose-dependent increase in mutagenicity from 0.25 ?M to 10 ?M B[a]P with no significant increase seen with further escalation to 50 ?M B[a]P. However, mutagenesis was 200-fold less than with the AKR pathway derived B[a]P, 7-8-dione. Mutant p53 plasmids, which yield red colonies, were recovered from the yeast to study the pattern and spectrum of mutations. The mutation pattern observed was G to T (31%) > G to C (29%) > G to A (14%). The frequency of codons mutated by the B[a]P radical cations was essentially random and not enriched at known cancer hotspots. The quinone products of radical cations, B[a]P-1,6-dione and B[a]P-3,6-dione were more mutagenic than the radical cation reactions, but still less mutagenic than AKR derived B[a]P-7,8-dione. We conclude that B[a]P radical cations and their quinone products are weakly mutagenic in this yeast-based system compared to redox cycling PAH o-quinones.
| S-EPMC3650728 | biostudies-literature
Project description:Exposure to environmental polycyclic aromatic hydrocarbons (PAHs), such as benzo(a)pyrene (B(a)P), has been linked to several health-threatening risks. PAHs were also shown to hinder adrenergic receptor (ADR) responses. As we previously demonstrated that B(a)P can directly interact with the ?2ADR, we investigated here whether B(a)P could decrease ?2ADR responsiveness by triggering receptor desensitization phenomena. We firstly showed that exposure to B(a)P reduced ?2ADR-mediated epinephrine-induced induction of NR4A gene mRNAs and of intracellular cAMP. Analysis of ?2ADR protein expression demonstrated that B(a)P rapidly decreased membrane expression of ?2ADR with a subsequent degradation of receptor protein. B(a)P exposure concomitantly rapidly increased the ?2ADR mRNA levels. The use of the ?-blockers, propranolol and ICI 118.551, demonstrated the involvement of ?2ADR itself in this increase. However, sustained exposure to B(a)P induced a diminution of ?2ADR mRNA steady-state as a result of the acceleration of its degradation. Together, these results show that, beside the well-known activation of the aryl hydrocarbon receptor, PAH deleterious effects may involve the dysfunction of adrenergic responses through, in part, the desensitization of ?2ADR. This may be taken in consideration when ?2-agonists/antagonists are administered in patients exposed to important concentrations of PAHs, e.g. in cigarette smokers.
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Project description:Melia azedarach-rhizosphere mediated degradation of benzo(a)pyrene (BaP), in the presence of cadmium (Cd) was studied, using efficient rhizobacterial isolate. Serratia marcescens S2I7, isolated from the petroleum-contaminated site, was able to tolerate up to 3.25 mM Cd. In the presence of Cd, the isolate S2I7 exhibited an increase in the activity of stress-responsive enzyme, glutathione-S-transferase. Gas Chromatography-Mass spectroscopy analysis revealed up to 59% in -vitro degradation of BaP after 21 days, while in the presence of Cd, the degradation was decreased by 14%. The bacterial isolate showed excellent plant growth-promoting attributes and could enhance the growth of host plant in Cd contaminated soil. The 52,41,555 bp genome of isolate S. marcescens S2I7 was sequenced, assembled and annotated into 4694 genes. Among these, 89 genes were identified for the metabolism of aromatic compounds and 172 genes for metal resistance, including the efflux pump system. A 2 MB segment of the genome was identified to contain operons for protocatechuate degradation, catechol degradation, benzoate degradation, and an IclR type regulatory protein pcaR, reported to be involved in the regulation of protocatechuate degradation. A pot trial was performed to validate the ability of S2I7 for rhizodegradation of BaP when applied through Melia azedarach rhizosphere. The rhizodegradation of BaP was significantly higher when augmented with S2I7 (85%) than degradation in bulk soil (68%), but decreased in the presence of Cd (71%).
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Project description:The microarray contains 9360 reporters, each present four times on the microarray. In our analysis 1326 human and viral reporters were used, of which 880 are unique human mature microRNAs. The other 8024 reporters represent microRNAs from other species. cDNA was generated using 1 μg of total RNA per sample. RNA was labeled only with Hy3 containing dye using the microRNACURY LNA™ microRNA Hy3 Power labeling kit (Exiqon, Denmark) according to the manufacturer’s protocol in a total volume of 25 μl. To adjust the volume to 100 μl, a hybridization buffer provided in miRCURY LNA™ microRNA Array, 5th generation kit (Exiqon, Denmark) was used.
2012-08-07 | GSE31252 | GEO
Project description:Benzo[a]pyrene (B[a]P), a polycyclic aromatic hydrocarbon, is a group 1 carcinogen that introduces mutagenic DNA adducts into the genome. In this study, we investigated the molecular mechanisms underlying the involvement of silymarin in the reduction of DNA adduct formation by B[a]P-7,8-dihydrodiol-9,10-epoxide (BPDE), induced by B[a]P. B[a]P exhibited toxicity in HepG2 cells, whereas co-treatment of the cells with B[a]P and silymarin reduced the formation of BPDE-DNA adducts, thereby increasing cell viability. Determination of the level of major B[a]P metabolites in the treated cells showed that BPDE levels were reduced by silymarin. Nuclear factor erythroid 2-related factor 2 (Nrf2) and pregnane X receptor (PXR) were found to be involved in the activation of detoxifying genes against B[a]P-mediated toxicity. Silymarin did not increase the expression of these major transcription factors, but greatly facilitated their nuclear translocation. In this manner, treatment of HepG2 cells with silymarin modulated detoxification enzymes through NRF2 and PXR to eliminate B[a]P metabolites. Knockdown of Nrf2 abolished the preventive effect of silymarin on BPDE-DNA adduct formation, indicating that activation of the Nrf2 pathway plays a key role in preventing B[a]P-induced genotoxicity. Our results suggest that silymarin has anti-genotoxic effects, as it prevents BPDE-DNA adduct formation by modulating the Nrf2 and PXR signaling pathways.
| S-EPMC7177818 | biostudies-literature
Project description:Benzo[a]pyrene (BaP) is a polycyclic aromatic hydrocarbon (PAH) that has been implicated in modulating aromatase enzyme function with the potential to interrupt normal reproductive function. The aim of this study was to use a fish model, Fundulus heteroclitus, to assess whether BaP exposure adversely impacts reproduction. Adult fish were exposed to waterborne BaP nominal concentrations of (0, 1, or 10 ?g/l) for 28 days. Males and females were combined for the second half of the exposure (days 14-28) in order to quantitate egg production and fertilization success. Egg fertilization and subsequent hatching success of F1 embryos was significantly decreased by the high dose of BaP. In males, both gonad weight and plasma testosterone concentrations were significantly reduced compared to controls by 10 ?g/l BaP. Histopathological examination of testes including spermatogonia, spermatocyte and spermatid cyst areas, percentage of cysts per phase, and area of spermatozoa per seminiferous tubule were not significantly affected. Other biomarkers, including male liver weight, liver vitellogenin (vtg) mRNA expression and sperm concentrations, were also not affected. In females, estradiol concentrations were significantly reduced after BaP exposure, but egg production, gonad weight, liver weight, vtg expression and oocyte maturation were not altered. Steroid concentrations in Fundulus larvae from exposed parents at 1 and 3 weeks posthatch were not significantly changed. BaP exposure at these environmentally relevant concentrations caused negative alterations particularly in male fish to both biochemical and phenotypic biomarkers associated with reproduction and multigenerational embryo survival.
| S-EPMC4133586 | biostudies-literature