DNA methylation changes induced by BDE-209 are related to DNA damage response and germ cell development in GC-2spd
Graphical abstract
Introduction
Persistent organic pollutants (POPs) are born with the rapid development of industrialization, which have been ubiquitous in the surrounding environment (Chen et al., 2019a). Due to the persistence and bioaccumulation, POPs have brought global problems of environmental pollution (Wong et al., 2005). In the past few decades, POPs have caused adverse effects on wild life and human health (Li et al., 2006). Polybrominated diphenyl ethers (PBDEs) are a group of environmental persistent chemicals, which have been widely used as brominated flame retardants (BFRs) on household consumer products since 1970s. Decabrominated diphenyl ethers (BDE-209), a common homologue of PBDEs, which has been listed in the Stockholm Convention on POPs (UNEP, 2017). Exposure to BDE-209 could have multiple harmful effects on the environment and organisms (Chen et al., 2018b; Hou et al., 2019; Wang et al., 2018). Both animal and epidemiologic studies have proved that PBDEs induces biological toxicity in male reproduction and birth outcomes (Chen et al., 2018a; Zhang et al., 2020a).
Epigenetics plays a vital role in normal embryonic development and human disease etiology by participating in the regulation of cellular gene expression (Illum et al., 2018). DNA methylation is one of the most common mechanisms of epigenetic regulation, which has been considered to be an essential process in normal development of mammal. The addition of a methyl group to the 5′ position of cytosine is the important modifier of DNA methylation for genome organization (Zemach and Zilberman, 2010). DNA methylation has extensive impacts on the transcriptional regulation (Bird, 2002), and plays a major role in fetal growth and sperm maturation (Bernstein et al., 2007; Lambrot et al., 2013).
The abnormal gene methylation in germ lines may lead to the failure of spermiogenesis or embryonic development. A meta-analytic study found that male infertility is associated with altered sperm methylation (Santi et al., 2017). PBDEs exposure could increase aberrant DNA methylation in epididymal sperms (Suvorov et al., 2018). Besides, we previously reported that BDE-209 caused decreased sperm quality and quantity with DNA double-strand breaks (DSBs) and DNA damage response (DDR) (Li et al., 2019). Tseng et al. (2013) has also suggested the involvement of sperm DNA damage in BDE-209-induced male reproductive effects. However, it remains unclear whether DDR is associated with BDE-209-induced DNA methylation in male reproduction. There are different cell types and dynamically expressed DNA methyltransferases during spermatogenesis due to the active cell differentiation and meiosis in mammals (Uysal et al., 2016). Considering the essential differentiation of spermatocyte into mature sperm, we chose the stable GC-2spd cell line of mouse spermatocytes to accurately investigate the DNA methylation status after BDE-209 exposure.
In this study, we analyzed the global DNA methylation changes in mouse GC-2spd (GC-2) cells after BDE-209 treatment for 24 hr by using microarray technology. Remarkably, p53-related DNA damage response was determined on the basis of Gene Ontology (GO) enrichment. Gene verification related to cell growth and spermatogenesis was also conducted by qRT-PCR assay. This study aimed to clarify the underlying impacts of BDE-209 on the epigenetic mechanism of spermatocyte, focused on the gene transcription changes in germ cell development. Our findings revealed a preliminary epigenetic perspective to address the biological mechanism of BDE-209-induced spermatogenesis disorder, which might provide evidence for elucidating the male reproductive toxicity of BDE-209.
Section snippets
Cell culture and BDE-209 exposure
In this study, GC-2 cells were derived from Kunming cell bank, Chinese Academy of Sciences with the origin of American Type Culture Collection (ATCC). GC-2 cell was established by the stable co-transfection of freshly isolated spermatocytes, which have lost their differentiation potential, and currently arrested at a premeiotic stage. Cells were cultured in Dulbecco's Modified Eagle's Medium (DMEM; Gibco, USA) supplemented with 100 U/mL penicillin and 100 μg/mL streptomycin, 10% fetal bovine
BDE-209 induced extensive DNA methylation in GC-2 cells
Differential methylated CpGs were located on all analyzed chromosomes. We identified 32,083 (1%) differentially methylated CpG sites with the criteria of p-value < 0.05 and the methylation difference > 20% between BDE-209 group and the control (Fig. 1a). Among them, 16,164 (50.38%) were hypermethylated and 15,919 (49.62%) were hypomethylated (Fig. 1b). We identified the differentially methylated CpG sites by their different locations of genetic distribution and CpG content. Both the hyper-
Discussion
This study found that exposure to BDE-209 resulted in genome wide DNA methylation changes in GC-2 cells. Hyper- and hypo-methylated genes were functionally enriched and analyzed. BDE-209-induced gene methylation were related to germ cell development and DNA damage response in mouse spermatocyte in vitro. This could explain the potential mechanism of male reproductive toxicity caused by BDE-209 exposure. Our previous study has demonstrated that exposure to BDE-209 for 24 h inhibited the
Conclusions
In summary, BDE-209 induced aberrant DNA methylation changes in GC-2 cells. BDE-209 exposure could interfere with spermatogenic cell growth and development by affecting the functional gene transcription activity. Remarkably, BDE-209 altered the gene transcription of p53-dependent DNA damage response, which was consistent with our previous results. Altogether, the negative impacts of BDE-209 on germ line development might be highly interrelated with the changes of genome wide DNA methylation.
Acknowledgments
This work was supported by the National Natural Science Foundation of China (Nos. 31770441, 31971415).
References (63)
- et al.
The mammalian epigenome
Cell
(2007) - et al.
A review of toxicity induced by persistent organic pollutants (POPs) and endocrine-disrupting chemicals (EDCs) in the nematode Caenorhabditis elegans
J. Environ. Manage.
(2019) - et al.
Organophosphate flame retardants (OPFRs) induce genotoxicity in vivo: a survey on apoptosis, DNA methylation, DNA oxidative damage, liver metabolites, and transcriptomics
Environ. Int.
(2019) - et al.
Disruption of thyroid hormone levels by decabrominated diphenyl ethers (BDE-209) in occupational workers from a deca-BDE manufacturing plant
Environ. Int.
(2018) - et al.
Effects of BDE-209 contaminated sediments on zebrafish development and potential implications to human health
Environ. Int.
(2014) - et al.
Germline development from human pluripotent stem cells toward disease modeling of infertility
Fertil. Steril.
(2012) - et al.
BDE-209 induces autophagy and apoptosis via IRE1alpha/Akt/mTOR signaling pathway in human umbilical vein endothelial cells
Environ. Pollut.
(2019) - et al.
DNA methylation homeostasis in human and mouse development
Curr. Opin. Genet. Dev.
(2017) - et al.
p53 and reproduction
Fertil. Steril.
(2018) - et al.
Maternal exposures to persistent organic pollutants are associated with DNA methylation of thyroid hormone-related genes in placenta differently by infant sex
Environ. Int.
(2019)
BDE-209 and DBDPE induce male reproductive toxicity through telomere-related cell senescence and apoptosis in SD rat
Environ. Int.
BDE-209 induces male reproductive toxicity via cell cycle arrest and apoptosis mediated by DNA damage response signaling pathways
Environ. Pollut.
Toxicogenomic analyses of the effects of BDE-47/209, TBBPA/S and TCBPA on early neural development with a human embryonic stem cell in vitro differentiation system
Toxicol. Appl. Pharmacol.
Parental exposure to natural mixtures of POPs reduced embryo production and altered gene transcription in zebrafish embryos
Aquat. Toxicol.
Combinatorial function of transcription factors and cofactors
Curr. Opie. Genet. Dev.
Perinatal exposure to low dose 2,2′,4,4′-tetrabromodiphenyl ether (BDE-47) alters sperm DNA methylation in adult rats
Reprod. Toxicol.
DNA methyltransferases exhibit dynamic expression during spermatogenesis
Reprod. Biomed. Online
Oncogenic roles of DNA hypomethylation through the activation of cancer-germline genes
Cancer Lett.
Studies on the interaction of BDE-47 and BDE-209 with acetylcholinesterase (AChE) based on the neurotoxicity through fluorescence, UV–Vis spectra, and molecular docking
Toxicol. Lett.
A review on the usage of POP pesticides in China, with emphasis on DDT loadings in human milk
Chemosphere
Associations between PBDEs exposure from house dust and human semen quality at an e-waste areas in South China – a pilot study
Chemosphere
Evolution of eukaryotic DNA methylation and the pursuit of safer sex
Curr. Biol.
Toxicity of polybrominated diphenyl ethers (PBDEs) on rodent male reproductive system: a systematic review and meta-analysis of randomized control studies
Sci. Total Environ.
Umbilical cord blood PBDEs concentrations are associated with placental DNA methylation
Environ. Int.
Associations between in utero exposure to polybrominated diphenyl ethers, pathophysiological state of fetal growth and placental DNA methylation changes
Environ. Int.
FOXM1: an emerging master regulator of DNA damage response and genotoxic agent resistance
Biochim. Biophys. Acta
DNA methylation in human sperm: a systematic review
Hum. Reprod. Update
Total DNA methylation as a biomarker of DNA damage and tumor malignancy in intracranial meningiomas
BMC Cancer
DNA methylation patterns and epigenetic memory
Genes Dev.
Polybrominated diphenyl ethers in cord blood and perinatal outcomes from Laizhou Wan Birth Cohort, China
Environ. Sci. Pollut. Res. Int.
Transcriptional trajectories of human kidney injury progression
JCI Insight
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These authors contributed equally to this work.