Fomesafen (FSA) is widely used in soybean fields for weed control. However, the persisting characteristics of FSA in the agricultural soil or water may become a hidden danger causing environmental pollution and phytotoxicity to succession crops. In this study, the growth and physiological responses of rice to FSA were investigated. It was found that the growth of rice seedlings was obviously inhibited by FSA exposure especially at over 0.1 mg L⁻¹. To gain an insight into the molecular mechanisms for the potential ecotoxicology, four libraries of rice roots and shoots exposed to FSA were created and subjected to the global RNA-sequencing (RNA-Seq) combined with HRLC-Q-TOF-MS/MS analytical technologies to comprehensively characterize the biochemical processes and catalytic reactions involved in FSA metabolism in rice. Compared with those without FSA, 499 and 450 up-regulated genes in roots and shoots with FSA were detected. Many of them were closely correlated with the tolerance to environmental stress, detoxification of xenobiotics and molecular metabolism process including cytochrome P450, glutathione S-transferases and acetyltransferase. A total of eight metabolites and fourteen conjugates in the reactive pathways of hydrolysis, substitution, reduction, methylation, glycosylation, acetylation, and malonylation were characterized by HRLC-Q-TOF-MS/MS. The relationship between the metabolized derivatives of FSA and enhanced expression the corresponding enzymatic regulators was established. This study will help understand the mechanisms and pathways of FSA metabolism and inspire the further research on FSA degradation in the paddy crops and environmental or health risks.

Cadmium (Cd), which is considered a carcinogenic metal, promotes breast cancer (BC) progression, but the precise mechanism remains unclear. Herein, MCF-7 and T47-D cells were treated with 0.1, 1, and 10 μM cadmium chloride (CdCl₂) for 24, 48 and 72 h. In our study, Cd exposure significantly accelerated the proliferation, migration and invasion of MCF-7 and T47-D cells. Notably, Cd inhibited autophagic flux by suppressing ATG5-dependent autophagosome formation but had no significant effect on autophagosome-lysosome fusion and lysosomal function. The genetic enhancement of autophagy through ATG5 overexpression suppressed the Cd-mediated increases in proliferation, migration and invasion, which indicated a carcinogenic role of autophagy impairment in Cd-exposed BC cells. GSEA and GeneMANIA were utilized to demonstrate that the Cd-induced decrease in ACSS2 expression mechanistically inhibited ATG5-dependent autophagy in BC cells. Importantly, ACSS2 overexpression increased the level of H3K27 acetylation in the promoter region of ATG5, and this result maintained autophagic flux and abolished the Cd-induced increases in proliferation, migration and invasion. We also verified that the expression of ACSS2 in BC tissues was low and positively related to ATG5 expression. These findings indicated that the promoting effect of Cd on BC cell proliferation, migration and invasion through the impairment of ACSS2/ATG5-dependent autophagic flux suggests a new mechanism for BC cell proliferation and metastasis stimulated by Cd.

Heart development requires a precise temporal regulation of gene expression in cardiomyoblasts. Therefore, the transcriptional changes in differentiating cells can lead to congenital heart diseases. Although the genetic mutations underlie most of these alterations, exposure to environmental contaminants, such as bisphenol A (BPA), has been recently considered as a risk factor as well. In this study we investigated the genotoxic and epigenotoxic effects of BPA throughout cardiomyocyte differentiation. H9c2 cells (rat myoblasts) were exposed to 10 and 30 μM BPA before and during the last two days of cardiac-driven differentiation. Then, we have analysed the phenotypic and molecular modifications (at transcriptional, genetic and epigenetic level). The results showed that treated myoblasts developed a skeletal muscle cell-like phenotype. The transcriptional changes induced by BPA in genes codifying proteins involved in heart differentiation and function depend on the window of exposure to BPA. The exposure before differentiation repressed the expression of heart transcription factors (Hand2 and Gata4), whereas exposure during differentiation reduced the expression of cardiac-specific genes (Tnnt2, Myom2, Sln, and Atp2a1). Additionally, significant effects were observed regarding DNA damage and histone acetylation levels after the two periods of BPA exposure: in cells exposed to the toxicant the percentage of DNA repair foci (formed by the co-localization of γH2AX and 53BP1) increased in a dose-dependent manner, whereas the treatment with the toxicant triggered a decrease in the epigenetic marks H3K9ac and H3K27ac. Our in vitro results reveal that BPA seriously interferes with the process of cardiomyocyte differentiation, which could be related to the reported in vivo effects of this toxicant on cardiogenesis.

N⁶-methyladenosine (m6A) modification, the most prevalent form of RNA methylation, modulates gene expression post-transcriptionally. Di-(2-ethylhexyl) phthalate (DEHP) is a common environmental endocrine disrupting chemical that induces testicular injury due to the inhibition of the demethylase fat mass and obesity-associated protein (FTO) and increases the m6A modification. How FTO-mediated m6A modification in testicular Leydig cell injury induced by DEHP remains unclear. Here, the TM3 Leydig cell line was treated with mono-(2-ethylhexyl) phthalate (MEHP), the main metabolite of DEHP in the body, as well as FB23-2, an inhibitor of FTO. Decreased levels of testosterone in the culture supernatant, significantly increased apoptosis, and a remarkable upregulation of global m6A modification were found in both TM3 cells treated with MEHP and FB23-2. Transcriptome sequencing showed that both treatments significantly induced apoptosis-associated gene expression. Methylated RNA immunoprecipitation sequencing showed that the Leydig cell injury induced by upregulated m6A modification could be associated with multiple physiological disorders, including histone acetylation, reactive oxygen species biosynthesis, MAPK signaling pathway, hormone secretion regulation, autophagy regulation, and male gonadal development. Overall, the inhibition of FTO-mediated up-regulation of m6A could be involved in MEHP-induced Leydig cell apoptosis.

Lipid metabolism could be used as a biomarker for environmental monitoring of metal pollution, including Cu. Given the potential role of the Wnt/β-catenin signaling pathway and acetylation in lipid metabolism, the aim of this study was to investigate the mechanism of Wnt signaling and acetylation mediating Cu-induced lipogenesis. Grass carp Ctenopharyngodon idella, widely distributed freshwater teleost, were used as the model. We found that waterborne Cu exposure increased the accumulation of Cu and lipid, up-regulated lipogenesis, suppressed Wnt signaling, reduced β-catenin protein level and its nuclear location, reduced the sirt1 mRNA levels and up-regulated the β-catenin acetylation level. Further investigation found that Cu up-regulated lipogenesis through Wnt/β-catenin pathway; Cu regulated the β-catenin acetylation, and K311 was the key acetylated residue after Cu incubation. SIRT1 mediated Cu-induced changes of acetylated β-catenin and played an essential role in nuclear accumulation of β-catenin and Cu-induced lipogenesis. Cu facilitated lipid accumulation via the regulation of Wnt pathway by SIRT1. For the first time, our study uncovered the novel mechanism for Wnt/β-catenin pathway and β-catenin acetylation levels mediating Cu-induced lipid deposition, which provided insights into the association between Cu exposure and lipid metabolism in fish and had important environmental implications for monitoring metal pollution in the water by using new biomarkers involved in lipid metabolism.

Bisphenol A (BPA) is an endocrine disruptor whose ubiquitous presence in the environment has been related with impairment of male reproduction. BPA can cause both transcriptomic and epigenetic changes during spermatogenesis. To evaluate the potential effects of male exposure to BPA, adult zebrafish males were exposed during spermatogenesis to doses of 100 and 2000 μg/L, which were reported in contaminated water bodies and higher than those allowed for human consumption. Fertilization capacity and survival at hatching were analysed after mating with untreated females. Spermatogenic progress was analysed through a morphometrical study of testes and apoptosis was evaluated by TUNEL assay. Testicular gene expression was evaluated by RT-qPCR and epigenetics by using ELISA and immunocytochemistry. In vitro studies were performed to investigate the role of Gper. Chromatin fragmentation and the presence of transcripts were also evaluated in ejaculated sperm. Results on testes from males treated with the highest dose showed a significant decrease in spermatocytes, an increase in apoptosis, a downregulation of ccnb1 and sycp3, all of which point to an alteration of spermatogenesis and to meiotic arrest and an upregulation of gper1 and esrrga receptors. Additionally, BPA at 2000 μg/L caused missregulation of epigenetic remodelling enzymes transcripts in testes and promoted DNA hypermethylation and H3K27me3 demethylation. BPA also triggered an increase in histone acetyltransferase activity, which led to hyperacetylation of histones (H3K9ac, H3K14ac, H4K12ac). In vitro reversion of histone acetylation changes using a specific GPER antagonist, G-36, suggested this receptor as mediator of histone hyperacetylation. Males treated with the lower dose only showed an increase in some histone acetylation marks (H3K14ac, H4K12ac) but their progeny displayed very limited survival at hatching, revealing the deleterious effects of unbalanced paternal epigenetic information. Furthermore, the highest dose of BPA led to chromatin fragmentation, promoting direct reproductive effects, which are incompatible with embryo development.

Benzene is a common environmental carcinogen that induces leukemia. Studies suggest that metabolic disorder has a relationship with the toxicity of benzene. Pyruvate kinase M2 (PKM2) is a key rate-limiting enzyme in glycolysis. However, the upstream and downstream regulatory mechanisms of PKM2 in benzene-induced hematotoxicity and the therapeutic effects of targeting PKM2 in vivo are unclear. This study aims to provide insights into the new mechanism of benzene-induced hematotoxicity and reveal the therapeutic significance of targeting PKM2. Herein, we demonstrated that PKM2-dependent glycolysis contributes to benzene-induced hematotoxicity by regulating inflammation reaction. Mechanistically, acetylated proteomics revealed that 1,4-benzoquinone (1,4-BQ) induced acetylation of PKM2 at position K66, and this modification contributed to the increase of PKM2 expression and can be inhibited by inhibition of acetyltransferase GCN5. Meanwhile, the elevated PKM2 was shown to prompt the activation of nuclear phosphorylated Stat3 (p-Stat3) and IL17A. Clinically, pharmacological inhibition of PKM2 alleviated the blood toxicity induced by benzene, which was mainly characterized by an increase in routine blood parameters and improvement of hematopoietic imbalance. Besides, elevated PKM2 is a promising biomarker in people occupationally exposed to benzene. Overall, we identified PKM2/p-Stat3/IL-17A axis participates in the hematotoxicity of benzene, and targeting PKM2 has certain therapeutic implications in hematologic diseases.

Hydroquinone (HQ), one of the main metabolites of benzene, is a well-known human leukemogen. However, the specific mechanism of how benzene or HQ contributes to the development of leukemia is unknown. In a previous study, we demonstrated the upregulation of DNA methyltransferase (DNMT) expression in HQ-induced malignant transformed TK6 (HQ-TK6) cells. Here, we investigated whether a regulatory loop between the long noncoding RNA FAS-AS1 and DNMT3b exists in HQ-TK6 cells and benzene-exposed workers. We found that the expression of FAS-AS1 was downregulated in HQ-TK6 cells and workers exposed to benzene longer than 1.5 years via histone acetylation, and FAS-AS1 expression was negatively correlated with the time of benzene exposure. Restoration of FAS-AS1 in HQ-TK6 cells promoted apoptosis and inhibited tumorigenicity in female nude mice. Interestingly, treatment with a DNMT inhibitor (5-aza-2-deoxycytidine), histone deacetylase inhibitor (trichostatin A), or DNMT3b knockout led to increased FAS-AS1 through increased H3K27ac protein expression in HQ-TK6 cells, and DNMT3b knockout decreased H3K27ac and DNMT3b enrichment to the FAS-AS1 promoter region, which suggested that DNMT3b and/or histone acetylation involve FAS-AS1 expression. Importantly, restoration of FAS-AS1 resulted in reduced expression of DNMT3b and SIRT1 and increased expression of FAS in both HQ-TK6 cells and xenograft tissues. Moreover, the average DNMT3b expression in 17 paired workers exposed to benzene within 1.5 years was decreased, but that of the remaining 103 paired workers with longer exposure times was increased. Conversely, DNMT3b was negatively correlated with FAS-AS1 expression. Both FAS-AS1 and DNMT3b influenced the enrichment of H3K27ac in the FAS promoter region by regulating the expression of SIRT1, consequently upregulating FAS expression. Taken together, these observations demonstrate crosstalk between FAS-AS1 and DNMT3b via a mutual inhibition loop and indicate a new mechanism by which FAS-AS1 regulates the expression of FAS in benzene-related carcinogenesis.

Artificial light at night (ALAN) exposes us to prolonged illumination, that adversely affects female reproduction. However, it remains to be clarified how prolonged light exposure affects oocyte meiotic maturation and quality. To this end, we exposed female mice to a constant light (CL) of 250 lux for different durations. Our findings showed that CL exposure for 7 weeks reduced the oocyte maturation rate. Meanwhile, CL exposure caused greater abnormalities in spindle assembly and chromosome alignment and a higher rate of oocyte aneuploidy than the regular light dark cycle. CL exposure also induced oxidative stress and caused mitochondrial dysfunction, which resulted in oocyte apoptosis and autophagy. Notably, our results showed that CL exposure reduced the levels of α-tubulin acetylation, DNA methylation at 5 mC, RNA methylation at m⁶A and histone methylation at H3K4me2 but increased the levels of histone methylation at H3K27me2 in oocytes. In summary, our findings demonstrate that constant bright light exposure causes oocyte meiotic defects and reduces cytoplasmic quality. These results extend the current understanding of ALAN-mediated defects in female reproduction.

N⁶-methyladenosine (m⁶A), the most abundant and reversible RNA modification, plays critical a role in tumorigenesis. However, whether m⁶A can regulate p53, a leading antitumor protein remains poorly understood. In this study, we explored the regulatory role of m⁶A on p53 activation using an arsenite-transformed keratinocyte model, the HaCaT-T cell line. We created the cell line by exposing human keratinocyte HaCaT cells to 1 μM arsenite for 5 months. We found that the cells exhibited an increased m⁶A level along with an aberrant expression of the methyltransferases, demethylase, and readers of m⁶A. Moreover, the cells exhibited decreased p53 activity and reduced p53 phosphorylation, acetylation, and transactivation with a high nucleus export rate of p53. Knockdown of the m⁶A methyltransferase, METTL3 significantly decreased m⁶A level, restoring p53 activation and inhibiting cellular transformation phenotypes in the arsenite-transformed cells. Further, using both a bioinformatics analysis and experimental approaches, we demonstrated that m⁶A downregulated the expression of the positive p53 regulator, PRDM2, through the YTHDF2-promoted decay of PRDM2 mRNAs. We showed that m⁶A upregulated the expression of the negative p53 regulator, YY1 and MDM2 through YTHDF1-stimulated translation of YY1 and MDM2 mRNA. Taken together, our study revealed the novel role of m⁶A in mediating arsenite-induced human keratinocyte transformation by suppressing p53 activation. This study further sheds light on the mechanisms of arsenic carcinogenesis via RNA epigenetics.