The structure of DNA methyltransferase DNMT3C reveals an activity-tuning mechanism for DNA methylation.

DNA methylation is one of the major epigenetic mechanisms crucial for gene regulation and genome stability. De novo DNA methyltransferase DNMT3C is required for silencing evolutionarily young transposons during mice spermatogenesis. Mutation of DNMT3C led to a sterility phenotype that cannot be rescued by its homologs DNMT3A and DNMT3B. However, the structural basis of DNMT3C-mediated DNA methylation remains unknown. Here, we report the structure and mechanism of DNMT3C-mediated DNA methylation. The DNMT3C methyltransferase domain recognizes CpG-containing DNA in a manner similar to that of DNMT3A and DNMT3B, in line with their high sequence similarity. However, two evolutionary covariation sites, C543 and E590, diversify the substrate interaction among DNMT3C, DNMT3A, and DNMT3B, resulting in distinct DNA methylation activity and specificity between DNMT3C, DNMT3A, and DNMT3B in vitro. In addition, our combined structural and biochemical analysis reveals that the disease-causing rahu mutation of DNMT3C compromises its oligomerization and DNA-binding activities, explaining the loss of DNA methylation activity caused by this mutation. This study provides a mechanistic insight into DNMT3C-mediated DNA methylation that complements DNMT3A- and DNMT3B-mediated DNA methylation in mice, unraveling a regulatory mechanism by which evolutionary conservation and diversification fine-tune the activity of de novo DNA methyltransferases.

Genome-wide comparative methylation analysis reveals the fate of germ stem cells after surrogate production in teleost.

BACKGROUND: Surrogate production by germline stem cell transplantation is a powerful method to produce donor-derived gametes via a host, a practice known as surrogacy. The gametes produced by surrogates are often analysed on the basis of their morphology and species-specific genotyping, which enables conclusion to be drawn about the donor's characteristics. However, in-depth information, such as data on epigenetic changes, is rarely acquired. Germ cells develop in close contact with supporting somatic cells during gametogenesis in vertebrates, and we hypothesize that the recipient's gonadal environment may cause epigenetic changes in produced gametes and progeny. Here, we extensively characterize the DNA methylome of donor-derived sperm and their intergenerational effects in both inter- and intraspecific surrogates. RESULTS: We found more than 3000 differentially methylated regions in both the sperm and progeny derived from inter- and intraspecific surrogates. Hypermethylation in the promoter regions of the protocadherin gamma gene in the intraspecific surrogates was found to be associated with germline transmission. On the contrary, gene expression level and the embryonic development of the offspring remained unaffected. We also discovered MAPK/p53 pathway disruption in interspecific surrogates due to promoter hypermethylation and identified that the inefficient removal of meiotic-arrested endogenous germ cells in hybrid gonads led to the production of infertile spermatozoa. CONCLUSIONS: Donor-derived sperm and progeny from inter- and intraspecific surrogates were more globally hypermethylated than those of the donors. The observed changes in DNA methylation marks in the surrogates had no significant phenotypic effects in the offspring.

Differential CpG methylation at Nnat in the early establishment of beta cell heterogeneity.

AIMS/HYPOTHESIS: Beta cells within the pancreatic islet represent a heterogenous population wherein individual sub-groups of cells make distinct contributions to the overall control of insulin secretion. These include a subpopulation of highly connected 'hub' cells, important for the propagation of intercellular Ca2+ waves. Functional subpopulations have also been demonstrated in human beta cells, with an altered subtype distribution apparent in type 2 diabetes. At present, the molecular mechanisms through which beta cell hierarchy is established are poorly understood. Changes at the level of the epigenome provide one such possibility, which we explore here by focusing on the imprinted gene Nnat (encoding neuronatin [NNAT]), which is required for normal insulin synthesis and secretion. METHODS: Single-cell RNA-seq datasets were examined using Seurat 4.0 and ClusterProfiler running under R. Transgenic mice expressing enhanced GFP under the control of the Nnat enhancer/promoter regions were generated for FACS of beta cells and downstream analysis of CpG methylation by bisulphite sequencing and RNA-seq, respectively. Animals deleted for the de novo methyltransferase DNA methyltransferase 3 alpha (DNMT3A) from the pancreatic progenitor stage were used to explore control of promoter methylation. Proteomics was performed using affinity purification mass spectrometry and Ca2+ dynamics explored by rapid confocal imaging of Cal-520 AM and Cal-590 AM. Insulin secretion was measured using homogeneous time-resolved fluorescence imaging. RESULTS: Nnat mRNA was differentially expressed in a discrete beta cell population in a developmental stage- and DNA methylation (DNMT3A)-dependent manner. Thus, pseudo-time analysis of embryonic datasets demonstrated the early establishment of Nnat-positive and -negative subpopulations during embryogenesis. NNAT expression is also restricted to a subset of beta cells across the human islet that is maintained throughout adult life. NNAT+ beta cells also displayed a discrete transcriptome at adult stages, representing a subpopulation specialised for insulin production, and were diminished in db/db mice. 'Hub' cells were less abundant in the NNAT+ population, consistent with epigenetic control of this functional specialisation. CONCLUSIONS/INTERPRETATION: These findings demonstrate that differential DNA methylation at Nnat represents a novel means through which beta cell heterogeneity is established during development. We therefore hypothesise that changes in methylation at this locus may contribute to a loss of beta cell hierarchy and connectivity, potentially contributing to defective insulin secretion in some forms of diabetes. DATA AVAILABILITY: The mass spectrometry proteomics data have been deposited to the ProteomeXchange Consortium via the PRIDE partner repository with the dataset identifier PXD048465.

5-mC methylation study of sORFs in 3'UTR of transcription factor JUNGBRUNNEN 1-like during leaf rust pathogenesis in wheat.

BACKGROUND: JUB1, a NAC domain containing hydrogen peroxide-induced transcription factor, plays a critical role in plant immunity. Little is known about how JUB1 responds to leaf rust disease in wheat. Recent discoveries in genomics have also unveiled a multitude of sORFs often assumed to be non-functional, to argue for the necessity of including them as potential regulatory players of translation. However, whether methylation on sORFs spanning the 3'UTR of regulatory genes like JUB1 modulate gene expression, remains unclear. METHODS AND RESULTS: In this study, we identified the methylation states of two sORFs in 3'UTR of a homologous gene of JUB1 in wheat, TaJUB1-L, at cytosine residues in CpG, CHH and CHG sites at different time points of disease progression in two near-isogenic lines of wheat (HD2329), with and without Lr24 gene during leaf rust pathogenesis. Here, we report a significant demethylation of the CpG dinucleotides occurring in the sORFs of the 3'UTR in the resistant isolines after 24 h post-infection. Also, the up-regulated gene expression observed through RT-qPCR was directly proportional to the demethylation of the CpG sites in the sORFs. CONCLUSIONS: Our findings indicate that TaJUB1-L might be a positive regulator in providing tolerance during leaf rust pathogenesis and cytosine methylation at 3'UTR might act as a switch for its expression control. These results enrich the potential benefit of conventional methylation assay techniques for unraveling the unexplored enigma in epigenetics during plant-pathogen interaction in a cost-effective and confidentially conclusive manner.

Glioblastoma epigenomics discloses a complex biology and potential therapeutic targets.

Background and purpose:

Glioblastoma (GBM), a highly aggressive form of brain tumors, has been extensively studied using OMICS methods, and the most characteristic molecular determinants have been incorporated into the histopathological diagnosis. Research data, nevertheless, only partially have been adopted in clinical practice. Here we aimed to present results of our epige­no­mic GBM profiling to better understand early and late determinants of these tumors, and to share main elements of our findings with practicing professionals.

GBM specimens were surgically obtained after first diagnosis (GBM1) and at recurrence (GBM2). DNA was extracted from 24 sequential pairs of formalin-fixed, paraffin-embedded tumor tissues. The Reduced Representation Bisulfite Sequencing kit was used for library preparation. Pooled libraries were sequenced on an Illumina NextSeq 550 instrument. Methylation controls (MC) were obtained from a publicly available database. Bioinformatic analyses were performed to identify differentially methylated pathways and their elements in cohorts of MC, GBM1 and GBM2.

Several differentially methylated pathways involved in basic intracellular and brain tissue developmental processes were identified in the GBM1 vs. MC and GBM2 vs. MC comparisons. Among differentially me­thylated pathways, those involved in immune regulation, neurotransmitter (particularly dopaminergic, noradrenergic and glutaminergic) responses and regulation of stem cell differentiation and proliferation stood out in the GBM2 vs. GBM1 comparisons.

Our study revealed biological complexity of early and late gliomagenesis encompassing mechanisms from basic intracellular through distorted neurodevelopmental processes to more specific immune and highjacked neurotransmitter pathways in the tumor microenvironment. These findings may offer considerations for therapeutic approaches.

SARS-CoV-2 hot-spot mutations are significantly enriched within inverted repeats and CpG island loci.

SARS-CoV-2 is an intensively investigated virus from the order Nidovirales (Coronaviridae family) that causes COVID-19 disease in humans. Through enormous scientific effort, thousands of viral strains have been sequenced to date, thereby creating a strong background for deep bioinformatics studies of the SARS-CoV-2 genome. In this study, we inspected high-frequency mutations of SARS-CoV-2 and carried out systematic analyses of their overlay with inverted repeat (IR) loci and CpG islands. The main conclusion of our study is that SARS-CoV-2 hot-spot mutations are significantly enriched within both IRs and CpG island loci. This points to their role in genomic instability and may predict further mutational drive of the SARS-CoV-2 genome. Moreover, CpG islands are strongly enriched upstream from viral ORFs and thus could play important roles in transcription and the viral life cycle. We hypothesize that hypermethylation of these loci will decrease the transcription of viral ORFs and could therefore limit the progression of the disease.

Loss of TSC complex enhances gluconeogenesis via upregulation of Dlk1-Dio3 locus miRNAs.

Loss of the tumor suppressor tuberous sclerosis complex 1 (Tsc1) in the liver promotes gluconeogenesis and glucose intolerance. We asked whether this could be attributed to aberrant expression of small RNAs. We performed small-RNA sequencing on liver of Tsc1-knockout mice, and found that miRNAs of the delta-like homolog 1 (Dlk1)-deiodinase iodothyronine type III (Dio3) locus are up-regulated in an mTORC1-dependent manner. Sustained mTORC1 signaling during development prevented CpG methylation and silencing of the Dlk1-Dio3 locus, thereby increasing miRNA transcription. Deletion of miRNAs encoded by the Dlk1-Dio3 locus reduced gluconeogenesis, glucose intolerance, and fasting blood glucose levels. Thus, miRNAs contribute to the metabolic effects observed upon loss of TSC1 and hyperactivation of mTORC1 in the liver. Furthermore, we show that miRNA is a downstream effector of hyperactive mTORC1 signaling.

A methyl-TROSY approach for NMR studies of high-molecular-weight DNA with application to the nucleosome core particle.

The development of methyl-transverse relaxation-optimized spectroscopy (methyl-TROSY)-based NMR methods, in concert with robust strategies for incorporation of methyl-group probes of structure and dynamics into the protein of interest, has facilitated quantitative studies of high-molecular-weight protein complexes. Here we develop a one-pot in vitro reaction for producing NMR quantities of methyl-labeled DNA at the C5 and N6 positions of cytosine (5mC) and adenine (6mA) nucleobases, respectively, enabling the study of high-molecular-weight DNA molecules using TROSY approaches originally developed for protein applications. Our biosynthetic strategy exploits the large number of naturally available methyltransferases to specifically methylate DNA at a desired number of sites that serve as probes of structure and dynamics. We illustrate the methodology with studies of the 153-base pair Widom DNA molecule that is simultaneously methyl-labeled at five sites, showing that high-quality 13C-1H spectra can be recorded on 100 µM samples in a few minutes. NMR spin relaxation studies of labeled methyl groups in both DNA and the H2B histone protein component of the 200-kDa nucleosome core particle (NCP) establish that methyl groups at 5mC and 6mA positions are, in general, more rigid than Ile, Leu, and Val methyl probes in protein side chains. Studies focusing on histone H2B of NCPs wrapped with either wild-type DNA or DNA methylated at all 26 CpG sites highlight the utility of NMR in investigating the structural dynamics of the NCP and how its histone core is affected through DNA methylation, an important regulator of transcription.

Cooperative In Situ Assembly of G-Quadruplex DNAzyme Nanowires for One-Step Sensing of CpG Methylation in Human Genomes.

CpG methylation is one the most predominant epigenetic modification that has been recognized as a molecular-level biomarker for various human diseases. Taking advantage of methylation-dependent cleavage and encoding flexibility in nucleic acid functions and structures, we demonstrate the cooperative in situ assembly of G-quadruplex DNAzyme nanowires for one-step sensing of CpG methylation in human genomes. This nanodevice displays good specificity and high sensitivity with a limit of detection (LOD) of 0.565 aM in vitro and 1 cell in vivo. It can distinguish 0.001% CpG methylation level from excess unmethylated DNA, quantify different CpG methylation targets from diverse human cancer cells, and even discriminate CpG methylation expressions between lung tumor and precancerous tissues. Importantly, this nanodevice can be performed isothermally in one step within 2 h in a label-free manner without any bisulfite conversion, fluorescence tagging, and PCR amplification process, providing a new platform for genomic methylation-related clinical diagnosis and biomedical research.

Perinatal S-Adenosylmethionine Supplementation Represses PSEN1 Expression by the Cellular Epigenetic Memory of CpG and Non-CpG Methylation in Adult TgCRD8 Mice.

DNA methylation, the main epigenetic modification regulating gene expression, plays a role in the pathophysiology of neurodegeneration. Previous evidence indicates that 5'-flanking hypomethylation of PSEN1, a gene involved in the amyloidogenic pathway in Alzheimer's disease (AD), boosts the AD-like phenotype in transgenic TgCRND8 mice. Supplementation with S-adenosylmethionine (SAM), the methyl donor in the DNA methylation reactions, reverts the pathological phenotype. Several studies indicate that epigenetic signatures, driving the shift between normal and diseased aging, can be acquired during the first stages of life, even in utero, and manifest phenotypically later on in life. Therefore, we decided to test whether SAM supplementation during the perinatal period (i.e., supplementing the mothers from mating to weaning) could exert a protective role towards AD-like symptom manifestation. We therefore compared the effect of post-weaning vs. perinatal SAM treatment in TgCRND8 mice by assessing PSEN1 methylation and expression and the development of amyloid plaques. We found that short-term perinatal supplementation was as effective as the longer post-weaning supplementation in repressing PSEN1 expression and amyloid deposition in adult mice. These results highlight the importance of epigenetic memory and methyl donor availability during early life to promote healthy aging and stress the functional role of non-CpG methylation.

Epigenetic Modification of Cytosines in Hematopoietic Differentiation and Malignant Transformation.

The mammalian DNA methylation landscape is established and maintained by the combined activities of the two key epigenetic modifiers, DNA methyltransferases (DNMT) and Ten-eleven-translocation (TET) enzymes. Once DNMTs produce 5-methylcytosine (5mC), TET proteins fine-tune the DNA methylation status by consecutively oxidizing 5mC to 5-hydroxymethylcytosine (5hmC) and further oxidized derivatives. The 5mC and oxidized methylcytosines are essential for the maintenance of cellular identity and function during differentiation. Cytosine modifications with DNMT and TET enzymes exert pleiotropic effects on various aspects of hematopoiesis, including self-renewal of hematopoietic stem/progenitor cells (HSPCs), lineage determination, differentiation, and function. Under pathological conditions, these enzymes are frequently dysregulated, leading to loss of function. In particular, the loss of DNMT3A and TET2 function is conspicuous in diverse hematological disorders, including myeloid and lymphoid malignancies, and causally related to clonal hematopoiesis and malignant transformation. Here, we update recent advances in understanding how the maintenance of DNA methylation homeostasis by DNMT and TET proteins influences normal hematopoiesis and malignant transformation, highlighting the potential impact of DNMT3A and TET2 dysregulation on clonal dominance and evolution of pre-leukemic stem cells to full-blown malignancies. Clarification of the normal and pathological functions of DNA-modifying epigenetic regulators will be crucial to future innovations in epigenetic therapies for treating hematological disorders.

FZD7, Regulated by Non-CpG Methylation, Plays an Important Role in Immature Porcine Sertoli Cell Proliferation.

The regulatory role of non-CpG methylation in mammals has been important in whole-genome bisulfite sequencing. It has also been suggested that non-CpG methylation regulates gene expression to affect the development and health of mammals. However, the dynamic regulatory mechanisms of genome-wide, non-CpG methylation during testicular development still require intensive study. In this study, we analyzed the dataset from the whole-genome bisulfite sequencing (WGBS) and the RNA-seq of precocious porcine testicular tissues across two developmental stages (1 and 75 days old) in order to explore the regulatory roles of non-CpG methylation. Our results showed that genes regulated by non-CpG methylation affect the development of testes in multiple pathways. Furthermore, several hub genes that are regulated by non-CpG methylation during testicular development-such as VEGFA, PECAM1, and FZD7-were also identified. We also found that the relative expression of FZD7 was downregulated by the zebularine-induced demethylation of the first exon of FZD7. This regulatory relationship was consistent with the results of the WGBS and RNA-seq analysis. The immature porcine Sertoli cells were transfected with RNAi to mimic the expression patterns of FZD7 during testicular development. The results of the simulation test showed that cell proliferation was significantly impeded and that cell cycle arrest at the G2 phase was caused by the siRNA-induced FZD7 inhibition. We also found that the percentage of early apoptotic Sertoli cells was decreased by transfecting them with the RNAi for FZD7. This indicates that FZD7 is an important factor in linking the proliferation and apoptosis of Sertoli cells. We further demonstrated that Sertoli cells that were treated with the medium collected from apoptotic cells could stimulate proliferation. These findings will contribute to the exploration of the regulatory mechanisms of non-CpG methylation in testicular development and of the relationship between the proliferation and apoptosis of normal somatic cells.

nc886, an RNA Polymerase III-Transcribed Noncoding RNA Whose Expression Is Dynamic and Regulated by Intriguing Mechanisms.

nc886 is a medium-sized non-coding RNA that is transcribed by RNA polymerase III (Pol III) and plays diverse roles in tumorigenesis, innate immunity, and other cellular processes. Although Pol III-transcribed ncRNAs were previously thought to be expressed constitutively, this concept is evolving, and nc886 is the most notable example. The transcription of nc886 in a cell, as well as in human individuals, is controlled by multiple mechanisms, including its promoter CpG DNA methylation and transcription factor activity. Additionally, the RNA instability of nc886 contributes to its highly variable steady-state expression levels in a given situation. This comprehensive review discusses nc886's variable expression in physiological and pathological conditions and critically examines the regulatory factors that determine its expression levels.

The Methylation Analysis of the Glucose-Dependent Insulinotropic Polypeptide Receptor (GIPR) Locus in GH-Secreting Pituitary Adenomas.

The glucose-dependent insulinotropic polypeptide receptor (GIPR) is aberrantly expressed in about one-third of GH-secreting pituitary adenomas (GH-PAs) and has been associated with a paradoxical increase of GH after a glucose load. The reason for such an overexpression has not yet been clarified. In this work, we aimed to evaluate whether locus-specific changes in DNA methylation patterns could contribute to this phenomenon. By cloning bisulfite-sequencing PCR, we compared the methylation pattern of the GIPR locus in GIPR-positive (GIPR+) and GIPR-negative (GIPR-) GH-PAs. Then, to assess the correlation between Gipr expression and locus methylation, we induced global DNA methylation changes by treating the lactosomatotroph GH3 cells with 5-aza-2'-deoxycytidine. Differences in methylation levels were observed between GIPR+ and GIPR- GH-PAs, both within the promoter (31.9% vs. 68.2%, p < 0.05) and at two gene body regions (GB_1 20.7% vs. 9.1%; GB_2 51.2% vs. 65.8%, p < 0.05). GH3 cells treated with 5-aza-2'-deoxycytidine showed a ~75% reduction in Gipr steady-state level, possibly associated with the observed decrease in CpGs methylation. These results indicate that epigenetic regulation affects GIPR expression in GH-PAs, even though this possibly represents only a part of a much more complex regulatory mechanism.

Stabilization of VEGF i-motif structure by CpG methylation.

The intercalated motif (i-motif) is a non-canonical nucleic acid structure formed by intercalated hemi-protonated cytosine base pairs (C-C+) under acidic conditions. The i-motif structure formation is involved in biological processes such as transcription regulation. Therefore, the identification of factors controlling i-motif formation is important in elucidating the cellular functions it controls. We previously reported that the VEGF G-quadruplex structure is stabilized by CpG methylation. In this study, the effect of CpG methylation on the stability of the VEGF i-motif structure was investigated. The VEGF i-motif-forming oligonucleotide contains four cytosines on CpG sites, and three of the four cytosines (C4, C15, and C20) are involved in C-C+ formation in the i-motif structure. Circular dichroism (CD) spectra analysis demonstrated that full CpG methylation increased the pH of mid transition (pHT) of the i-motif structure by 0.1, and the melting temperature (Tm) by 5.1 °C in 25 mM sodium cacodylate buffer at pH 5.0. Moreover, single methylation at C4, C15, and C20 increased Tm by 0.5, 1.7, and 2.0 °C in the buffer, respectively. These results demonstrated that CpG methylation stabilized the VEGF i-motif structure.

TET3 dioxygenase modulates gene conversion at the avian immunoglobulin variable region via demethylation of non-CpG sites in pseudogene templates.

Diversification of the avian primary immunoglobulin (Ig) repertoire is achieved in developing B cells by somatic hypermutation (SHM) and gene conversion (GCV). GCV is a type of homologous recombination that unidirectionally transfers segments of Ig pseudogenes to Ig variable domains. It is regulated by epigenetic mechanisms like histone modifications, but the role of DNA methylation remains unclear. Here, we demonstrate that the chicken B-cell line DT40 lacking TET3, a member of the TET (Ten-eleven translocation) family dioxygenases that facilitate DNA demethylation, exhibited a marked reduction in GCV activity in Ig variable regions. This was accompanied by a drop in the bulk levels of 5-hydroxymethylcytosine, an oxidized derivative of 5-methylcytosine, whereas TET1-deficient or TET2-deficient DT40 strains did not exhibit such effects. Deletion of TET3 caused little effects on the expression of proteins required for SHM and GCV, but induced hypermethylation in some Ig pseudogene templates. Notably, the enhanced methylation occurred preferably on non-CpG cytosines. Disruption of both TET1 and TET3 significantly inhibited the expression of activation-induced cytidine deaminase (AID), an essential player in Ig diversification. These results uncover unique roles of TET proteins in avian Ig diversification, highlighting the potential importance of TET3 in maintaining hypomethylation In Ig pseudogenes.

Simultaneous measurement of the size and methylation of chromosome 4qA-D4Z4 repeats in facioscapulohumeral muscular dystrophy by long-read sequencing.

BACKGROUND: Facioscapulohumeral muscular dystrophy (FSHD) is an autosomal dominant muscular disorder characterized by asymmetric muscle wasting and weakness. FSHD can be subdivided into two types: FSHD1, caused by contraction of the D4Z4 repeat on chromosome 4q35, and FSHD2, caused by mild contraction of the D4Z4 repeat plus aberrant hypomethylation mediated by genetic variants in SMCHD1, DNMT3B, or LRIF1. Genetic diagnosis of FSHD is challenging because of the complex procedures required. METHODS: We applied Nanopore CRISPR/Cas9-targeted resequencing for the diagnosis of FSHD by simultaneous detection of D4Z4 repeat length and methylation status at nucleotide level in genetically-confirmed and suspected patients. RESULTS: We found significant hypomethylation of contracted 4q-D4Z4 repeats in FSHD1, and both 4q- and 10q-D4Z4 repeats in FSHD2. We also found that the hypomethylation in the contracted D4Z4 in FSHD1 is moderately correlated with patient phenotypes. CONCLUSIONS: Our method contributes to the development for the diagnosis of FSHD using Nanopore long-read sequencing. This finding might give insight into the mechanisms by which repeat contraction causes disease pathogenesis.

Association between prenatal cadmium exposure and cord blood DNA methylation.

Prenatal cadmium exposure is known to affect infant growth and organ development. Nonetheless, the role of DNA methylation in cadmium-related health effects has yet to be determined. To this end, we investigated the relationship between prenatal cadmium exposure and cord blood DNA methylation in Korean infants through an epigenome-wide association study. Cadmium concentrations in maternal blood during early and late pregnancy and in cord blood collected from newborns were measured using atomic adsorption spectrometry and DNA methylation analysis was conducted using HumanMethylationEPIC BeadChip kits. After adjusting for infant sex, maternal pregnancy body mass index, smoking status, and estimated leukocyte composition, we analyzed the association between CpG methylation and cadmium concentration in 364 samples. Among 835,252 CpG sites, maternal blood cadmium concentration in early pregnancy was significantly associated with two differentially methylated CpG sites, cg05537752 and cg24904393, which were annotated ATP9A and no gene, respectively. The study findings indicate that prenatal cadmium exposure is significantly associated with methylation statuses of several CpG sites and regions in Korean infants, especially during early pregnancy.

Gene-Targeted DNA Methylation: Towards Long-Lasting Reprogramming of Gene Expression?

DNA methylation is an essential epigenetic mark, strongly associated with gene expression regulation. Aberrant DNA methylation patterns underlie various diseases and efforts to intervene with DNA methylation signatures are of great clinical interest. Technological developments to target writers or erasers of DNA methylation to specific genomic loci by epigenetic editing resulted in successful gene expression modulation, also in in vivo models. Application of epigenetic editing in human health could have a huge impact, but clinical translation is still challenging. Despite successes for a wide variety of genes, not all genes mitotically maintain their (de)methylation signatures after editing, and reprogramming requires further understanding of chromatin context-dependency. In addition, difficulties of current delivery systems and off-target effects are hurdles to be tackled. The present review describes findings towards effective and sustained DNA (de)methylation by epigenetic editing and discusses the need for multi-effector approaches to achieve highly efficient long-lasting reprogramming.

Identification of novel genetic and epigenetic regulators of different tissue types of cervical cancer.

OBJECTIVES: The study aimed to find differential gene mutations, DNA methylation, and expression profiles among different categories of cervical cancer samples. METHODS: The study was based on freely available gene mutations, promoter methylation, and gene expression status of The Cancer Genome Atlas (TCGA) cervical cancer samples and adjacent normal tissues in the Genomic Data Commons (GDC) portal. The association of CpG island methylation with gene expression was determined through negative correlation analysis. RESULTS: We identified that the ErbB signaling pathway and proteoglycans pathway was significantly associated with adenocarcinoma cervical cancers patients. In these pathways, missense mutation especially S310F in the ERBB2 gene as well as G12D and A146T in the KRAS gene were significantly associated with adenocarcinoma cases. Furthermore, a comparison of SCC cases with adjacent control tissues revealed differential hypermethylation of two CpG positions of the KAAG1 gene and differential downregulation of NPY1R and NPY5R genes in cervical squamous cell carcinoma compared to cervical adenocarcinoma cases and adjacent normal tissues. Specifically, the hypermethylation of the promoter region of the KAAG1 gene might be responsible for the carcinogenesis of cervical squamous cells exclusively and methylation marks can be reversible by the widely used drug, azacytidine. In contrast, adenocarcinoma cervical cancer cases may be treated with floxuridine which is successfully utilized for other tissue-specific adenocarcinoma cases. CONCLUSIONS: These results provide valuable insight into the differential molecular markers among the categories of cervical cancer, which helps our ability to classify these cancers and for targeted therapy.