Understanding the role of ten-eleven translocation family proteins in kidney diseases.

Epigenetic mechanisms play a critical role in the pathogenesis of human diseases including kidney disorders. As the erasers of DNA methylation, Ten-eleven translocation (TET) family proteins can oxidize 5-methylcytosine (5mC) to 5-hydroxymethylcytosine (5hmC), 5-formylcytosine (5fC), and 5-carboxylcytosine (5caC), thus leading to passive or active DNA demethylation. Similarly, TET family proteins can also catalyze the same reaction on RNA. In addition, TET family proteins can also regulate chromatin structure and gene expression in a catalytic activity-independent manner through recruiting the SIN3A/HDAC co-repressor complex. In 2012, we reported for the first time that the genomic 5-hydroxymethylcytosine level and the mRNA levels of Tet1 and Tet2 were significantly downregulated in murine kidneys upon ischemia and reperfusion injury. Since then, accumulating evidences have eventually established an indispensable role of TET family proteins in not only acute kidney injury but also chronic kidney disease. In this review, we summarize the upstream regulatory mechanisms and the pathophysiological role of TET family proteins in major types of kidney diseases and discuss their potential values in clinical diagnosis and treatment.

Placenta-derived SOD3 deletion impairs maternal behavior via alterations in FGF/FGFR-prolactin signaling axis.

Offspring growth requires establishing maternal behavior associated with the maternal endocrine profile. Placentae support the adaptations of the mother, producing bioactive molecules that affect maternal organs. We recently reported that placentae produce superoxide dismutase 3 (SOD3) that exerts sustained effects on the offspring liver via epigenetic modifications. Here, we demonstrate that placenta-specific Sod3 knockout (Sod3-/-) dams exhibited impaired maternal behavior and decreased prolactin levels. Most fibroblast growth factor (FGF)-regulated pathways were downregulated in the pituitary tissues from Sod3-/- dams. FGF1-, FGF2-, and FGF4-induced prolactin expression and signaling via the phosphoinositide 3-kinase (PI3K)-phospholipase C-γ1 (PLCγ1)-protein kinase-Cδ (PKC)δ axis were reduced in primary pituitary cells from Sod3-/- dams. Mechanistically, FGF1/FGF receptor (FGFR)2 expressions were inhibited by the suppression of the ten-eleven translocation (TET)/isocitrate dehydrogenase (IDH)/α-ketoglutarate pathway and DNA demethylation levels at the zinc finger and BTB domain containing 18 (ZBTB18)-targeted promoters of Fgf1/Fgfr2. Importantly, offspring from Sod3-/- dams also showed impaired nurturing behavior to their grandoffspring. Collectively, placenta-derived SOD3 promotes maternal behavior via epigenetic programming of the FGF/FGFR-prolactin axis.

The Role of DNMT Methyltransferases and TET Dioxygenases in the Maintenance of the DNA Methylation Level.

This review deals with the functional characteristics and biological roles of enzymes participating in DNA methylation and demethylation as key factors in epigenetic regulation of gene expression. The set of enzymes that carry out such processes in human cells is limited to representatives of two families, namely DNMT (DNA methyltransferases) and TET (DNA dioxygenases). The review presents detailed information known today about each functionally important member of these families and describes the catalytic activity and roles in the mammalian body while also providing examples of dysregulation of the expression and/or activity of these enzymes in conjunction with the development of some human disorders, including cancers, neurodegenerative diseases, and developmental pathologies. By combining the up-to-date information on the dysfunction of various enzymes that control the DNA "methylome" in the human body, we hope not only to draw attention to the importance of the maintenance of a required DNA methylation level (ensuring epigenetic regulation of gene expression and normal functioning of the entire body) but also to help identify new targets for directed control over the activity of the enzymes that implement the balance between processes of DNA methylation and demethylation.

Genome-wide characterization of dynamic DNA 5-hydroxymethylcytosine and TET2-related DNA demethylation during breast tumorigenesis.

BACKGROUND: Breast tumorigenesis is a complex and multistep process accompanied by both genetic and epigenetic dysregulation. In contrast to the extensive studies on DNA epigenetic modifications 5-hydroxymethylcytosine (5hmC) and 5-methylcytosine (5mC) in malignant breast tumors, their roles in the early phases of breast tumorigenesis remain ambiguous. RESULTS: DNA 5hmC and 5mC exhibited a consistent and significant decrease from usual ductal hyperplasia to atypical ductal hyperplasia and subsequently to ductal carcinoma in situ (DCIS). However, 5hmC showed a modest increase in invasive ductal breast cancer compared to DCIS. Genomic analyses showed that the changes in 5hmC and 5mC levels occurred around the transcription start sites (TSSs), and the modification levels were strongly correlated with gene expression levels. Meanwhile, it was found that differentially hydroxymethylated regions (DhMRs) and differentially methylated regions (DMRs) were overlapped in the early phases and accompanied by the enrichment of active histone marks. In addition, TET2-related DNA demethylation was found to be involved in breast tumorigenesis, and four transcription factor binding sites (TFs: ESR1, FOXA1, GATA3, FOS) were enriched in TET2-related DhMRs/DMRs. Intriguingly, we also identified a certain number of common DhMRs between tumor samples and cell-free DNA (cfDNA). CONCLUSIONS: Our study reveals that dynamic changes in DNA 5hmC and 5mC play a vital role in propelling breast tumorigenesis. Both TFs and active histone marks are involved in TET2-related DNA demethylation. Concurrent changes in 5hmC signals in primary breast tumors and cfDNA may play a promising role in breast cancer screening.

Development of a rapid mass spectrometric method for the analysis of ten-eleven translocation enzymes.

In DNA, methylation at the fifth position of cytosine (5mC) by DNA methyltransferases is essential for eukaryotic gene regulation. Methylation patterns are dynamically controlled by epigenetic machinery. Erasure of 5mC by Fe2+ and 2-ketoglutarate (2KG) dependent dioxygenases in the ten-eleven translocation family (TET1-3), plays a key role in nuclear processes. Through the event of active demethylation, TET proteins iteratively oxidize 5mC to 5-hydroxymethyl cytosine (5hmC), 5-formylcytosine (5fC) and 5-carboxycytosine (5caC), each of which has been implicated in numerous diseases when aberrantly generated. A wide range of biochemical assays have been developed to characterize TET activity, many of which require multi-step processing to detect and quantify the 5mC oxidized products. Herein, we describe the development and optimization of a sensitive MALDI mass spectrometry-based technique that directly measures TET activity and eliminates tedious processing steps. Employing optimized assay conditions, we report the steady-state activity of wild type TET2 enzymes to furnish 5hmC, 5fC and 5caC. We next determine IC50 values of several small-molecule inhibitors of TETs. The utility of this assay is further demonstrated by analyzing the activity of V1395A which is an activating mutant of TET2 that primarily generates 5caC. Lastly, we describe the development of a secondary assay that utilizes bisulfite chemistry to further examine the activity of wildtype TET2 and V1395A in a base-resolution manner. The combined results demonstrate that the activity of TET proteins can be gauged, and their products accurately quantified using our methods.

Fucoidan prevents diabetic cognitive dysfunction via promoting TET2-mediated active DNA demethylation in high-fat diet induced diabetic mice.

Diabetic cognitive dysfunction (DCD) refers to cognitive impairment in individuals with diabetes, which is one of the most important comorbidities and complications. Preliminary evidence suggests that consuming sufficient dietary fiber could have benefits for both diabetes and cognitive function. However, the effect and underlying mechanism of dietary fiber on DCD remain unclear. We conducted a cross-sectional analysis using data from NHANES involving 2072 diabetics and indicated a significant positive dose-response relationship between the dietary fiber intake and cognitive performance in diabetics. Furthermore, we observed disrupted cognitive function and neuronal morphology in high-fat diet induced DCD mice, both of which were effectively restored by fucoidan supplementation through alleviating DNA epigenetic metabolic disorders. Moreover, fucoidan supplementation enhanced the levels of short-chain fatty acids (SCFAs) in the cecum of diabetic mice. These SCFAs enhanced TET2 protein stability by activating phosphorylated AMPK and improved TETs activity by reducing the ratio of (succinic acid + fumaric acid)/ α-ketoglutaric acid, subsequently enhancing TET2 function. The positive correlation between dietary fiber intake and cognitive function in diabetics was supported by human and animal studies alike. Importantly, fucoidan can prevent the occurrence of DCD by promoting TET2-mediated active DNA demethylation in the cerebral cortex of diabetic mice.

Experimental Varicocele Impairs DNA Methylation and Demethylation in Germ Cells, TESE, and Epididymal Spermatozoa, Impacting Active DNA Demethylation in Zygotes.

Varicocele causes infertility. The current study has investigated the impact of experimental varicocele on DNA methylation, demethylation, and damage in the germ cells, TESE-derived and epididymal spermatozoa. Moreover, the results were compared between epidydimal and TESE-derived spermatozoa. Finally, the varicocele-induced effect on active DNA demethylation (ADD) of male pronucleus and pre-implantation embryo development was assessed. The mature male rats were divided into control, control-sham (undergone simple laparotomy), and experimental varicocele-induced groups (n = 6/each group). The left renal vein semi-ligation was considered to induce varicocele. The expression levels of DNA methyltransferase 1 (DNMT1) and ten-eleven-translocation proteins (TET1, 2, 3), and global DNA methylation in testicular tissue, TESE, and epididymis-derived spermatozoa, and the ADD in zygotes male pronucleus as well as pre-implantation embryo development were assessed. The expression levels of DNMT1 and TET1, 2, 3 in testicles, TESE, and epididymis-derived spermatozoa were decreased in the varicocele group compared to the control and control-sham groups. The TESE-derived spermatozoa exhibited higher DNMT1, higher DNMT1, and TET 1, 2, and no change in TET3 expression compared to epididymis-derived spermatozoa. The varicocele group represented lower DNA methylation in the testicles, TESE-derived and epididymal spermatozoa, higher 5mC+ signal in male pronucleus, and a lower pre-implantation embryo development compared to control and control-sham rats. The TESE-derived spermatozoa exhibited higher 5mC protein expression compared to epididymal spermatozoa. In conclusion, varicocele can negatively impact the DNA methylation/demethylation processes impairing spermatogenesis and leading to fertilization failure, which may ultimately result in a decrease in embryo development by increasing susceptibility to DNA damage.

TET enzyme driven epigenetic reprogramming in early embryos and its implication on long-term health.

Mammalian embryo development is initiated by the union of paternal and maternal gametes. Upon fertilization, their epigenome landscape is transformed through a series of finely orchestrated mechanisms that are crucial for survival and successful embryogenesis. Specifically, maternal or oocyte-specific reprogramming factors modulate germ cell specific epigenetic marks into their embryonic states. Rapid and dynamic changes in epigenetic marks such as DNA methylation and histone modifications are observed during early embryo development. These changes govern the structure of embryonic genome prior to zygotic genome activation. Differential changes in epigenetic marks are observed between paternal and maternal genomes because the structure of the parental genomes allows interaction with specific oocyte reprogramming factors. For instance, the paternal genome is targeted by the TET family of enzymes which oxidize the 5-methylcytosine (5mC) epigenetic mark into 5-hydroxymethylcytosine (5hmC) to lower the level of DNA methylation. The maternal genome is mainly protected from TET3-mediated oxidation by the maternal factor, STELLA. The TET3-mediated DNA demethylation occurs at the global level and is clearly observed in many mammalian species. Other epigenetic modulating enzymes, such as DNA methyltransferases, provide fine tuning of the DNA methylation level by initiating de novo methylation. The mechanisms which initiate the epigenetic reprogramming of gametes are critical for proper activation of embryonic genome and subsequent establishment of pluripotency and normal development. Clinical cases or diseases linked to mutations in reprogramming modulators exist, emphasizing the need to understand mechanistic actions of these modulators. In addition, embryos generated via in vitro embryo production system often present epigenetic abnormalities. Understanding mechanistic actions of the epigenetic modulators will potentially improve the well-being of individuals suffering from these epigenetic disorders and correct epigenetic abnormalities in embryos produced in vitro. This review will summarize the current understanding of epigenetic reprogramming by TET enzymes during early embryogenesis and highlight its clinical relevance and potential implication for assisted reproductive technologies.

Selective Estrogen Receptor Modulators' (SERMs) Influence on TET3 Expression in Breast Cancer Cell Lines with Distinct Biological Subtypes.

Tamoxifen, a selective estrogen receptor modulator (SERM), exhibits dual agonist or antagonist effects contingent upon its binding to either G-protein-coupled estrogen receptor (GPER) or estrogen nuclear receptor (ESR). Estrogen signaling plays a pivotal role in initiating epigenetic alterations and regulating estrogen-responsive genes in breast cancer. Employing three distinct breast cancer cell lines-MCF-7 (ESR+; GPER+), MDA-MB-231 (ESR-; GPER-), and SkBr3 (ESR-; GPER+)-this study subjected them to treatment with two tamoxifen derivatives: 4-hydroxytamoxifen (4-HT) and endoxifen (Endox). Through 2D high-performance liquid chromatography with tandem mass spectrometry detection (HPLC-MS/MS), varying levels of 5-methylcytosine (5-mC) were found, with MCF-7 displaying the highest levels. Furthermore, TET3 mRNA expression levels varied among the cell lines, with MCF-7 exhibiting the lowest expression. Notably, treatment with 4-HT induced significant changes in TET3 expression across all cell lines, with the most pronounced increase seen in MCF-7 and the least in MDA-MB-231. These findings underscore the influence of tamoxifen derivatives on DNA methylation patterns, particularly through modulating TET3 expression, which appears to be contingent on the presence of estrogen receptors. This study highlights the potential of targeting epigenetic modifications for personalized anti-cancer therapy, offering a novel avenue to improve treatment outcomes.

Effect of Vitis vinifera zygotic embryo cryopreservation and post-cryopreservation on the gene expression of DNA demethylases.

Grapevine (Vitis vinifera L.) crops are continuously exposed to biotic and abiotic stresses, which can cause genetic and epigenetic alterations. To determine the possible effects of grapevine cryopreservation on the regulation of DNA demethylase genes, this work studied the expression of DNA demethylase genes in cryopreserved and post-cryopreserved grapevine tissues. V. vinifera DNA demethylases were characterized by in silico analysis, and gene expression quantification was conducted by RT‒qPCR. Three DNA demethylase sequences were found: VIT_13s0074g00450 (VvDMT), VIT_08s0007g03920 (VvROS1), and VIT_06s0061g01270 (VvDML3). Phylogenetic analysis revealed that the sequences from V. vinifera and A. thaliana had a common ancestry. In the promoters of responsive elements to transcription factors such as AP-2, Myb, bZIP, TBP, and GATA, the conserved domains RRM DME and Perm CXXC were detected. These responsive elements play roles in the response to abiotic stress and the regulation of cell growth. These data helped us characterize the V. vinifera DNA demethylase genes. Gene expression analysis indicated that plant vitrification solution 2 (PVS2) treatment does not alter the expression of DNA demethylase genes. The expression levels of VvDMT and VvROS1 increased in response to cryopreservation by vitrification. Furthermore, in post-cryopreservation, VvROS1 was highly induced, and VvDML3 was repressed in all the treatment groups. Gene expression differences between different treatments and tissues may play roles in controlling methylation patterns during gene regulation in tissues stressed by cryopreservation procedures and in the post-cryopreservation period during plant growth and development.

TET1 displays catalytic and non-catalytic functions in the adult mouse cortex.

TET1/2/3 dioxygenases iteratively demethylate 5-methylcytosine, beginning with the formation of 5-hydroxymethylcytosine (5hmC). The post-mitotic brain maintains higher levels of 5hmC than most peripheral tissues, and TET1 ablation studies have underscored the critical role of TET1 in brain physiology. However, deletion of Tet1 precludes the disentangling of the catalytic and non-catalytic functions of TET1. Here, we dissect these functions of TET1 by comparing adult cortex of Tet1 wildtype (Tet1 WT), a novel Tet1 catalytically dead mutant (Tet1 HxD), and Tet1 knockout (Tet1 KO) mice. Using DNA methylation array, we uncover that Tet1 HxD and KO mutations perturb the methylation status of distinct subsets of CpG sites. Gene ontology (GO) analysis on specific differential 5hmC regions indicates that TET1's catalytic activity is linked to neuronal-specific functions. RNA-Seq further shows that Tet1 mutations predominantly impact the genes that are associated with alternative splicing. Lastly, we performed High-performance Liquid Chromatography Mass-Spectrometry lipidomics on WT and mutant cortices and uncover accumulation of lysophospholipids lysophosphatidylethanolamine and lysophosphatidylcholine in Tet1 HxD cortex. In summary, we show that Tet1 HxD does not completely phenocopy Tet1 KO, providing evidence that TET1 modulates distinct cortical functions through its catalytic and non-catalytic roles.

The biological function of demethylase ALKBH1 and its role in human diseases.

AlkB homolog 1 (ALKBH1) is a member of the AlkB family of dioxygenases that are dependent on Fe(II) and α-ketoglutarate. Mounting evidence demonstrates that ALKBH1 exhibits enzymatic activity against various substrates, including N6-methyladenosine (m6A), N1-methyladenosine (m1A), N3-methylcytidine (m3C), 5-methylcytosine (m5C), N6-methyladenine (N6-mA, 6mA), and H2A, indicating its dual roles in different biological processes and involvement in human diseases. Up to the present, there is ongoing debate regarding ALKBH1's enzymatic activity. In this review, we present a comprehensive summary of recent research on ALKBH1, including its substrate diversity and pathological roles in a wide range of human disorders, the underlying mechanisms of its functions, and its dysregulation. We also explored the potential of ALKBH1 as a prognostic target.

Loss of Tet hydroxymethylase activity causes mouse embryonic stem cell differentiation bias and developmental defects.

The TET family is well known for active DNA demethylation and plays important roles in regulating transcription, the epigenome and development. Nevertheless, previous studies using knockdown (KD) or knockout (KO) models to investigate the function of TET have faced challenges in distinguishing its enzymatic and nonenzymatic roles, as well as compensatory effects among TET family members, which has made the understanding of the enzymatic role of TET not accurate enough. To solve this problem, we successfully generated mice catalytically inactive for specific Tet members (Tetm/m). We observed that, compared with the reported KO mice, mutant mice exhibited distinct developmental defects, including growth retardation, sex imbalance, infertility, and perinatal lethality. Notably, Tetm/m mouse embryonic stem cells (mESCs) were successfully established but entered an impaired developmental program, demonstrating extended pluripotency and defects in ectodermal differentiation caused by abnormal DNA methylation. Intriguingly, Tet3, traditionally considered less critical for mESCs due to its lower expression level, had a significant impact on the global hydroxymethylation, gene expression, and differentiation potential of mESCs. Notably, there were common regulatory regions between Tet1 and Tet3 in pluripotency regulation. In summary, our study provides a more accurate reference for the functional mechanism of Tet hydroxymethylase activity in mouse development and ESC pluripotency regulation.

Active DNA Demethylase, TET1, Increases Oxidative Phosphorylation and Sensitizes Ovarian Cancer Stem Cells to Mitochondrial Complex I Inhibitor.

Ten-eleven translocation 1 (TET1) is a methylcytosine dioxygenase involved in active DNA demethylation. In our previous study, we demonstrated that TET1 reprogrammed the ovarian cancer epigenome, increased stem properties, and activated various regulatory networks, including metabolic networks. However, the role of TET1 in cancer metabolism remains poorly understood. Herein, we uncovered a demethylated metabolic gene network, especially oxidative phosphorylation (OXPHOS). Contrary to the concept of the Warburg effect in cancer cells, TET1 increased energy production mainly using OXPHOS rather than using glycolysis. Notably, TET1 increased the mitochondrial mass and DNA copy number. TET1 also activated mitochondrial biogenesis genes and adenosine triphosphate production. However, the reactive oxygen species levels were surprisingly decreased. In addition, TET1 increased the basal and maximal respiratory capacities. In an analysis of tricarboxylic acid cycle metabolites, TET1 increased the levels of α-ketoglutarate, which is a coenzyme of TET1 dioxygenase and may provide a positive feedback loop to modify the epigenomic landscape. TET1 also increased the mitochondrial complex I activity. Moreover, the mitochondrial complex I inhibitor, which had synergistic effects with the casein kinase 2 inhibitor, affected ovarian cancer growth. Altogether, TET1-reprogrammed ovarian cancer stem cells shifted the energy source to OXPHOS, which suggested that metabolic intervention might be a novel strategy for ovarian cancer treatment.

AGEs induce MMP-9 promoter demethylation through the GADD45α-mediated BER pathway to promote breast cancer metastasis in patients with diabetes.

Scientific evidence has linked diabetes to a higher incidence and increased aggressiveness of breast cancer; however, mechanistic studies of the numerous regulators involved in this process are insufficiently thorough. Advanced glycation end products (AGEs) play an important role in the chronic complications of diabetes, but the mechanisms of AGEs in breast cancer are largely unexplored. In this study, we first demonstrate that high AGE levels in breast cancer tissues are associated with the diabetic state and poor patient outcomes. Furthermore, AGEs interact with the receptor for AGEs (RAGE) to promote breast cancer cell migration and invasion. Mechanistically, based on RNA sequencing (RNA-seq) analysis, we reveal that growth arrest and DNA damage gene 45α (GADD45α) is a vital protein upregulated by AGEs through a P53-dependent pathway. Next, GADD45α recruits thymine DNA glycosylase for base excision repair to form the demethylation complex at the promoter region of MMP-9 and enhance MMP-9 transactivation through DNA demethylation. Overall, our results indicate a critical regulatory role of AGEs in patients with breast cancer and diabetes and reveal a novel mechanism of epigenetic modification in promoting breast cancer metastasis.

FLI1 is associated with regulation of DNA methylation and megakaryocytic differentiation in FPDMM caused by a RUNX1 transactivation domain mutation.

Familial platelet disorder with associated myeloid malignancies (FPDMM) is an autosomal dominant disease caused by heterozygous germline mutations in RUNX1. It is characterized by thrombocytopenia, platelet dysfunction, and a predisposition to hematological malignancies. Although FPDMM is a precursor for diseases involving abnormal DNA methylation, the DNA methylation status in FPDMM remains unknown, largely due to a lack of animal models and challenges in obtaining patient-derived samples. Here, using genome editing techniques, we established two lines of human induced pluripotent stem cells (iPSCs) with different FPDMM-mimicking heterozygous RUNX1 mutations. These iPSCs showed defective differentiation of hematopoietic progenitor cells (HPCs) and megakaryocytes (Mks), consistent with FPDMM. The FPDMM-mimicking HPCs showed DNA methylation patterns distinct from those of wild-type HPCs, with hypermethylated regions showing the enrichment of ETS transcription factor (TF) motifs. We found that the expression of FLI1, an ETS family member, was significantly downregulated in FPDMM-mimicking HPCs with a RUNX1 transactivation domain (TAD) mutation. We demonstrated that FLI1 promoted binding-site-directed DNA demethylation, and that overexpression of FLI1 restored their megakaryocytic differentiation efficiency and hypermethylation status. These findings suggest that FLI1 plays a crucial role in regulating DNA methylation and correcting defective megakaryocytic differentiation in FPDMM-mimicking HPCs with a RUNX1 TAD mutation.

Identification of DEMETER-like DNA demethylase gene family in citrus and their role in drought stress-adaptive responses.

DEMETER-Like DNA demethylases (DMLs) are epigenetic regulators of many developmental and biological processes in plants. No comprehensive information about the DML gene family in citrus is available to date. Here, a total of three DML genes in the genomes of Citrus sinensis (named CsDML1-3) and C. clementina (named CcDML1-3) were identified and analyzed. They encode hydrophilic and relatively large proteins, with prediction of nuclear localization, containing the conserved domains and motifs typical of plant DMLs. Protein interaction network analysis suggested that they interact primarily with proteins related to the maintenance of DNA methylation and remodeling of chromatin. Analysis of their promoter regions led to the identification of several cis-acting regulatory elements involved in stress response, including drought, heat and cold stresses. The presence of several miRNA targets and potential phosphorylation sites suggest that their expression is also regulated at post-transcriptional and post-translational levels. RNA-Seq data and quantitative real-time PCR analysis showed a low and drought-regulated gene expression of the citrus DMLs in different plant tissues. CsDML1 and CsDML3 were also differentially regulated by deficit irrigation in fruits at different developmental stages, with a positive and significant correlation found between CsDML1 and PHYTOENE SYNTHASE (PSY) and between CsDML3 and ATP CITRATE LYASEs (ACLs) and ZETA-CAROTENE DESATURASE (ZDS) gene expression. These results indicate that the citrus DMLs are potentially functional enzymes involved in developmental processes and drought stress-adaptive responses, providing a useful reference for further investigation of their functions and applications on the citrus improvement.

Prenatal exposure to low-dose bisphenol A disrupts hippocampal DNA methylation and demethylation in male rat offspring.

Earlier research has demonstrated that developmental exposure to bisphenol A (BPA) has persistent impacts on both adult brain growth and actions. It has been suggested that BPA might obstruct the methylation coding of the genes in the brain. In this study, the methylation changes in the hippocampus tissue of male rat pups were examined following prenatal BPA exposure. Pregnant Sprague-Dawley rats were treated with either vehicle (tocopherol-stripped corn oil) or BPA (4, 40, or 400 µg/kg·body weight/day) throughout the entire duration of gestation and lactation. At 3 weeks of age, the male rat offspring were euthanized, and the hippocampus were dissected out for analysis. The expression levels of DNA methyltransferases (DNMT1, DNMT3A, and DNMT3B) and DNA demethylases (TET1, Gadd45a, Gadd45b, and Apobec1) were analyzed in the hippocampus by means of quantitative real-time polymerase chain reaction and Western blotting, respectively. The results showed that prenatal exposure to BPA upregulated the expression of enzymes associated with DNA methylation and demethylation processes in the hippocampus of male rat offspring. These findings suggest that prenatal exposure to a low dose of BPA could potentially disrupt the balance of methylation and demethylation in the hippocampus, thereby perturbing epigenetic modifications. This may represent a neurotoxicity mechanism of BPA.

DPPA3 facilitates genome-wide DNA demethylation in mouse primordial germ cells.

BACKGROUND: Genome-wide DNA demethylation occurs in mammalian primordial germ cells (PGCs) as part of the epigenetic reprogramming important for gametogenesis and resetting the epigenetic information for totipotency. Dppa3 (also known as Stella or Pgc7) is highly expressed in mouse PGCs and oocytes and encodes a factor essential for female fertility. It prevents excessive DNA methylation in oocytes and ensures proper gene expression in preimplantation embryos: however, its role in PGCs is largely unexplored. In the present study, we investigated whether or not DPPA3 has an impact on CG methylation/demethylation in mouse PGCs. RESULTS: We show that DPPA3 plays a role in genome-wide demethylation in PGCs even before sex differentiation. Dppa3 knockout female PGCs show aberrant hypermethylation, most predominantly at H3K9me3-marked retrotransposons, which persists up to the fully-grown oocyte stage. DPPA3 works downstream of PRDM14, a master regulator of epigenetic reprogramming in embryonic stem cells and PGCs, and independently of TET1, an enzyme that hydroxylates 5-methylcytosine. CONCLUSIONS: The results suggest that DPPA3 facilitates DNA demethylation through a replication-coupled passive mechanism in PGCs. Our study identifies DPPA3 as a novel epigenetic reprogramming factor in mouse PGCs.

Epstein-Barr virus-driven metabolic alterations contribute to the viral lytic reactivation and tumor progression in nasopharyngeal carcinoma.

Metabolic reprogramming induced by Epstein-Barr virus (EBV) often mirrors metabolic changes observed in cancer cells. Accumulating evidence suggests that lytic reactivation is crucial in EBV-associated oncogenesis. The aim of this study was to explore the role of metabolite changes in EBV-associated malignancies and viral life cycle control. We first revealed that EBV (LMP1) accelerates the secretion of the oncometabolite D-2HG, and serum D-2HG level is a potential diagnostic biomarker for NPC. EBV (LMP1)-driven metabolite changes disrupts the homeostasis of global DNA methylation and demethylation, which have a significantly inhibitory effect on active DNA demethylation and 5hmC content. We found that loss of 5hmC indicates a poor prognosis for NPC patients, and that 5hmC modification is a restriction factor of EBV reactivation. We confirmed a novel EBV reactivation inhibitor, α-KG, which inhibits the expression of EBV lytic genes with CpG-containing ZREs and the latent-lytic switch by enhancing 5hmC modification. Our results demonstrate a novel mechanism of which metabolite abnormality driven by EBV controls the viral lytic reactivation through epigenetic modification. This study presents a potential strategy for blocking EBV reactivation, and provides potential targets for the diagnosis and therapy of NPC.