Maintenance Of DNA Methylation Research Articles

DNA hypomethylation promotes UHRF1-and SUV39H1/H2-dependent crosstalk between H3K18ub and H3K9me3 to reinforce heterochromatin states.

Mono-ubiquitination of lysine 18 on histone H3 (H3K18ub), catalyzed by UHRF1, is a DNMT1 docking site that facilitates replication-coupled DNA methylation maintenance. Its functions beyond this are unknown. Here, we genomically map simultaneous increases in UHRF1-dependent H3K18ub and SUV39H1/H2-dependent H3K9me3 following DNMT1 inhibition. Mechanistically, transient accumulation of hemi-methylated DNA at CpG islands facilitates UHRF1 recruitment and E3 ligase activity toward H3K18. Notably, H3K18ub enhances SUV39H1/H2 methyltransferase activity and, in colon cancer cells, nucleates new H3K9me3 domains at CpG island promoters of DNA methylation-silenced tumor suppressor genes (TSGs). Disrupting UHRF1 enzyme activity prevents H3K9me3 accumulation while promoting PRC2-dependent H3K27me3 as a tertiary layer of gene repression in these regions. By contrast, disrupting H3K18ub-dependent SUV39H1/H2 activity enhances the transcriptional activating and antiproliferative effects of DNMT1 inhibition. Collectively, these findings reveal roles for UHRF1 and H3K18ub in regulating a hierarchy of repressive histone methylation signaling and rationalize a combination strategy for epigenetic cancer therapy.

UHRF1 ubiquitin ligase activity supports the maintenance of low-density CpG methylation.

The RING E3 ubiquitin ligase UHRF1 is an established cofactor for DNA methylation inheritance. The model posits that nucleosomal engagement through histone and DNA interactions directs UHRF1 ubiquitin ligase activity toward lysines on histone H3 tails, creating binding sites for DNMT1 through ubiquitin interacting motifs (UIM1 and UIM2). However, the extent to which DNMT1 relies on ubiquitin signaling through UHRF1 in support of DNA methylation maintenance remains unclear. Here, with integrative epigenomic and biochemical analyses, we reveal that DNA methylation maintenance at low-density cytosine-guanine dinucleotides (CpGs) is particularly vulnerable to disruption of UHRF1 ubiquitin ligase activity and DNMT1 ubiquitin reading activity through UIM1. Hypomethylation of low-density CpGs in this manner induces formation of partially methylated domains (PMDs), a methylation signature observed across human cancers. In contrast, UIM2 disruption completely abolishes the DNA methylation maintenance function of DNMT1 in a CpG density-independent manner. In the context of DNA methylation recovery following acute DNMT1 depletion, we further reveal a 'bookmarking' function for UHRF1 ubiquitin ligase activity in support of DNA re-methylation. Collectively, these studies show that DNMT1-dependent DNA methylation inheritance is a ubiquitin-regulated process that is partially reliant on UHRF1 and suggest a disrupted UHRF1-DNMT1 ubiquitin signaling axis contributes to PMD formation in cancers.

Aging drives a program of DNA methylation decay in plant organs.

How organisms age is a question with broad implications for human health. In mammals, DNA methylation is a biomarker for biological age, which may predict age more accurately than date of birth. However, limitations in mammalian models make it difficult to identify mechanisms underpinning age-related DNA methylation changes. Here, we show that the short-lived model plant Arabidopsis thaliana exhibits a loss of epigenetic integrity during aging, causing heterochromatin DNA methylation decay and the expression of transposable elements. We show that the rate of epigenetic aging can be manipulated by extending or curtailing lifespan, and that shoot apical meristems are protected from this aging process. We demonstrate that a program of transcriptional repression suppresses DNA methylation maintenance pathways during aging, and that mutants of this mechanism display a complete absence of epigenetic decay. This presents a new paradigm in which a gene regulatory program sets the rate of epigenomic information loss during aging.

Regulation of de novo and maintenance DNA methylation by DNA methyltransferases in post-implantation embryos.

DNA methylation is mainly catalyzed by three DNA methyltransferase (DNMT) proteins in mammals. Usually DNMT1 is considered the primary DNMT for maintenance DNA methylation, whereas DNMT3A and DNMT3B function in de novo DNA methylation. Interestingly, we found DNMT3A and DNMT3B exerted maintenance and de novo DNA methylation in post-implantation mouse embryos. Together with DNMT1, they maintained DNA methylation at some pluripotent genes and lineage marker genes. Germline-derived DNA methylation at the imprinting control regions (ICRs) is stably maintained in embryos. DNMT1 maintained DNA methylation at most ICRs in post-implantation embryos. Surprisingly, DNA methylation was increased at five ICRs after implantation, and two DNMT3 proteins maintained the newly acquired DNA methylation at two of these five ICRs. Intriguingly, DNMT3A and DNMT3B maintained pre-existing DNA methylation at four other ICRs, similar to what we found in embryonic stem (ES) cells before. These results suggest that DNA methylation is more dynamic than originally thought during embryogenesis including the ICRs of the imprinted regions. DNMT3A and DNMT3B exert both de novo and maintenance DNA methylation functions after implantation. They maintain large portions of newly acquired DNA methylation at variable degrees across the genome in mouse embryos, together with DNMT1. Furthermore, they contribute to maintenance of pre-existing DNA methylation at a subset of ICRs as well as in the CpG islands (CGIs) and certain lineage marker gene. These findings may have some implications for the important roles of DNMT proteins in development and human diseases.

UHRF1-mediated ubiquitination of nonhomologous end joining factor XLF promotes DNA repair in human tumor cells

UHRF1 (Ubiquitin-like with PHD and Ring Finger domains 1) is a crucial E3 ubiquitin ligase and epigenetic regulator with pivotal roles in various biological processes, including the maintenance of DNA methylation, regulation of gene expression, and facilitation of DNA damage repair. In this study, we unveil that UHRF1 interacts with the non-homologous end joining (NHEJ) factor XLF (also known as Cernunnos) following DNA double strand breaks (DSBs) in HeLa cells. Furthermore, we demonstrate that UHRF1 catalyzes lysine 63-linked polyubiquitination of XLF, rather than lysine 48-linked polyubiquitination. Notably, this polyubiquitination of XLF by UHRF1 does not affect its protein stability; instead, it enhances the recruitment of XLF to the sites of DNA damage. These findings shed light on the role of UHRF1 as a novel regulator of DNA repair through XLF in tumor cells.

CDCA7 is an evolutionarily conserved hemimethylated DNA sensor in eukaryotes.

Mutations of the SNF2 family ATPase HELLS and its activator CDCA7 cause immunodeficiency, centromeric instability, and facial anomalies syndrome, characterized by DNA hypomethylation at heterochromatin. It remains unclear why CDCA7-HELLS is the sole nucleosome remodeling complex whose deficiency abrogates the maintenance of DNA methylation. We here identify the unique zinc-finger domain of CDCA7 as an evolutionarily conserved hemimethylation-sensing zinc finger (HMZF) domain. Cryo-electron microscopy structural analysis of the CDCA7-nucleosome complex reveals that the HMZF domain can recognize hemimethylated CpG in the outward-facing DNA major groove within the nucleosome core particle, whereas UHRF1, the critical activator of the maintenance methyltransferase DNMT1, cannot. CDCA7 recruits HELLS to hemimethylated chromatin and facilitates UHRF1-mediated H3 ubiquitylation associated with replication-uncoupled maintenance DNA methylation. We propose that the CDCA7-HELLS nucleosome remodeling complex assists the maintenance of DNA methylation on chromatin by sensing hemimethylated CpG that is otherwise inaccessible to UHRF1 and DNMT1.

Aberrant DNA methylation associated with the development of metabolic dysfunction-associated fatty liver disease

The literature review deals with DNA methylation, a key epigenetic mechanism that controls the activity of gene transcription, plays a decisive role in the formation of genomic imprinting, gene silencing, X-chromosome inactivation, RNA splicing, DNA repair, cell differentiation and cell reprogramming, and also determines the occurrence and development of liver steatotic lesions and metabolic disorders. Methylation of DNA cytosine dinucleotide (CpG) can be represented in two types: de novo CpG methylation, which is carried out by 5mC DNA writers — DNA-(cytosine-5)-methyltransferase (DNMT) 3a and 3b, and suppor­ting DNA methylation, which is performed by DNMT1 during DNA replication. It has been found that the maintenance DNA methylation allows the preservation of the methylation pattern characteristic of progenitor cells in the cells of the new generation, and the DNA methylation of the gene body is associated with its increased expression. Active demethylation of 5mC is carried out by TET dioxygenases, including three enzymatic representatives: TET1, TET2 and TET3. It has been demonstrated that aberrant methylation of DNA nucleotides is directly related to the activity of lipid synthesis, the degree of oxidative stress, the development of liver steatosis, low-grade inflammation, insulin resistance, and the progression of liver fibrosis. The authors presented in detail the functions and features of DNA methyltransferases, erasers, and readers of 5mC sites; possible violations of the balance of activity of writers and erasers of 5mC DNA; DNA methylation landscape and patterns; clinical significance of DNA methylation signatures in metabolic dysfunction-associated fatty liver disease. Global hypomethylation of genome, at least 55 genes, is observed in patients with metabolic dysfunction-associated fatty liver disease. The authors emphasize that the use of DNA methylation signatures is a promising direction for early diagnosis and prognosis of the course of metabolic dysfunction-associated fatty liver disease, while the study of molecular components of DNA methylation mechanisms involved in the regulation of gene expression, the dependence of their activity on exposure to the exposome will allow to persona­lize and improve recommendations for lifestyle and diet modification in patients with metabolic dysfunction-associated fatty liver disease.

DNMT1 can induce primary germ layer differentiation through de novo DNA methylation.

DNA methyltransferases and Ten-Eleven Translocation (TET) proteins regulate the DNA methylation and demethylation cycles during mouse embryonic development. Although DNMT1 mainly plays a role in the maintenance of DNA methylation after DNA replication, it is also reported to possess de novo methyltransferase capacity. However, its physiological significance remains unclear. Here, we demonstrate that full-length DNMT1 (FL) and a mutant lacking the N-terminus necessary for its maintenance activity (602) confer the differentiation potential of mouse Dnmt1, Dnmt3a, and Dnmt3b (Dnmts-TKO) embryonic stem cells (ESCs). Both FL and 602 inhibit the spontaneous differentiation of Dnmts-TKO ESCs in the undifferentiated state. Dnmts-TKO ESCs showed loss of DNA methylation and de-repression of primitive endoderm-related genes, but these defects were partially restored in Dnmts-TKO + FL and Dnmts-TKO + 602 ESCs. Upon differentiation, Dnmts-TKO + FL ESCs show increased 5mC and 5hmC levels across chromosomes, including pericentromeric regions. In contrast, Dnmts-TKO + 602 ESCs didn't accumulate 5mC, and sister chromatids showed 5hmC asynchronously. Furthermore, in comparison with DNMT1_602, DNMT1_FL effectively promoted commitment to the epiblast-like cells and beyond, driving cell-autonomous mesendodermal and germline differentiation through embryoid body-based methods. With precise target selectivity achieved by its N-terminal region, DNMT1 may play a role in gene regulation leading to germline development.

Dynamics of DNA Methylation During Gametogenesis: Insights and Detection Methods

For many years, scientific research has dedicated significant attention to unraveling the complexities of inheritance, the process by which traits are passed from one generation to the next. At the heart of this exploration lies DNA methylation, a pivotal mechanism that exerts profound influence over gene expression, cellular identity, and the overarching development of organisms. This in-depth review meticulously probes the intricate dynamics of DNA methylation, meticulously dissecting its multifaceted impact on inheritance patterns and the spectrum of phenotypic variations observed. The review scrutinizes the underlying mechanisms governing the establishment and maintenance of DNA methylation, offering a nuanced understanding of its regulatory roles. Through a synthesis of insights gleaned from the fields of molecular biology, epigenetics, and genomics, it illuminates the far-reaching implications of DNA methylation for both health and disease. By peering into the molecular machinery that orchestrates these epigenetic modifications, the review provides a comprehensive framework for comprehending the interplay between DNA methylation and the transmission of hereditary traits. Moreover, by contextualizing these findings within the broader landscape of genetics and inheritance, the review offers valuable perspectives for guiding future research endeavors. These insights not only deepen our understanding of the intricate processes underlying heredity but also hold potential implications for the development of novel therapeutic interventions aimed at modulating DNA methylation patterns. In essence, this review serves as a cornerstone in the ongoing quest to unravel the mysteries of inheritance, paving the way for innovative approaches to address complex biological phenomena.

Structure of human DPPA3 bound to the UHRF1 PHD finger reveals its functional and structural differences from mouse DPPA3

DNA methylation maintenance is essential for cell fate inheritance. In differentiated cells, this involves orchestrated actions of DNMT1 and UHRF1. In mice, the high-affinity binding of DPPA3 to the UHRF1 PHD finger regulates UHRF1 chromatin dissociation and cytosolic localization, which is required for oocyte maturation and early embryo development. However, the human DPPA3 ortholog functions during these stages remain unclear. Here, we report the structural basis for human DPPA3 binding to the UHRF1 PHD finger. The conserved human DPPA3 85VRT87 motif binds to the acidic surface of UHRF1 PHD finger, whereas mouse DPPA3 binding additionally utilizes two unique α-helices. The binding affinity of human DPPA3 for the UHRF1 PHD finger was weaker than that of mouse DPPA3. Consequently, human DPPA3, unlike mouse DPPA3, failed to inhibit UHRF1 chromatin binding and DNA remethylation in Xenopus egg extracts effectively. Our data provide novel insights into the distinct function and structure of human DPPA3.

Semimethylation is a feature of diffuse large B-cell lymphoma, and subgroups with poor prognosis are characterized by global hypomethylation and short telomere length

BackgroundLarge B-cell lymphoma (LBCL) is the most common lymphoma and is known to be a biologically heterogeneous disease regarding genetic, phenotypic, and clinical features. Although the prognosis is good, one-third has a primary refractory or relapsing disease which underscores the importance of developing predictive biological markers capable of identifying high- and low-risk patients. DNA methylation (DNAm) and telomere maintenance alterations are hallmarks of cancer and aging. Both these alterations may contribute to the heterogeneity of the disease, and potentially influence the prognosis of LBCL.ResultsWe studied the DNAm profiles (Infinium MethylationEPIC BeadChip) and relative telomere lengths (RTL) with qPCR of 93 LBCL cases: Diffuse large B-cell lymphoma not otherwise specified (DLBCL, n = 66), High-grade B-cell lymphoma (n = 7), Primary CNS lymphoma (n = 8), and transformation of indolent B-cell lymphoma (n = 12). There was a substantial methylation heterogeneity in DLBCL and other LBCL entities compared to normal cells and other B-cell neoplasms. LBCL cases had a particularly aberrant semimethylated pattern (0.15 ≤ β ≤ 0.8) with large intertumor variation and overall low hypermethylation (β > 0.8). DNAm patterns could not be used to distinguish between germinal center B-cell-like (GC) and non-GC DLBCL cases. In cases treated with R-CHOP-like regimens, a high percentage of global hypomethylation (β < 0.15) was in multivariable analysis associated with worse disease-specific survival (DSS) (HR 6.920, 95% CI 1.499–31.943) and progression-free survival (PFS) (HR 4.923, 95% CI 1.286–18.849) in DLBCL and with worse DSS (HR 5.147, 95% CI 1.239–21.388) in LBCL. These cases with a high percentage of global hypomethylation also had a higher degree of CpG island methylation, including islands in promoter-associated regions, than the cases with less hypomethylation. Additionally, telomere length was heterogenous in LBCL, with a subset of the DLBCL-GC cases accounting for the longest RTL. Short RTL was independently associated with worse DSS (HR 6.011, 95% CI 1.319–27.397) and PFS (HR 4.689, 95% CI 1.102–19.963) in LBCL treated with R-CHOP-like regimens.ConclusionWe hypothesize that subclones with high global hypomethylation and hypermethylated CpG islands could have advantages in tumor progression, e.g. by inactivating tumor suppressor genes or promoting treatment resistance. Our findings suggest that cases with high global hypomethylation and thus poor prognosis could be candidates for alternative treatment regimens including hypomethylating drugs.

Recent Advances in Studies of Genomic DNA Methylation and Its Involvement in Regulating Drought Stress Response in Crops.

As global arid conditions worsen and groundwater resources diminish, drought stress has emerged as a critical impediment to plant growth and development globally, notably causing declines in crop yields and even the extinction of certain cultivated species. Numerous studies on drought resistance have demonstrated that DNA methylation dynamically interacts with plant responses to drought stress by modulating gene expression and developmental processes. However, the precise mechanisms underlying these interactions remain elusive. This article consolidates the latest research on the role of DNA methylation in plant responses to drought stress across various species, focusing on methods of methylation detection, mechanisms of methylation pattern alteration (including DNA de novo methylation, DNA maintenance methylation, and DNA demethylation), and overall responses to drought conditions. While many studies have observed significant shifts in genome-wide or gene promoter methylation levels in drought-stressed plants, the identification of specific genes and pathways involved remains limited. This review aims to furnish a reference for detailed research into plant responses to drought stress through epigenetic approaches, striving to identify drought resistance genes regulated by DNA methylation, specific signaling pathways, and their molecular mechanisms of action.

Combinatorial quantification of 5mC and 5hmC at individual CpG dyads and the transcriptome in single cells reveals modulators of DNA methylation maintenance fidelity.

Inheritance of 5-methylcytosine from one cell generation to the next by DNA methyltransferase 1 (DNMT1) plays a key role in regulating cellular identity. While recent work has shown that the activity of DNMT1 is imprecise, it remains unclear how the fidelity of DNMT1 is tuned in different genomic and cell state contexts. Here we describe Dyad-seq, a method to quantify the genome-wide methylation status of cytosines at the resolution of individual CpG dinucleotides to find that the fidelity of DNMT1-mediated maintenance methylation is related to the local density of DNA methylation and the landscape of histone modifications. To gain deeper insights into methylation/demethylation turnover dynamics, we first extended Dyad-seq to quantify all combinations of 5-methylcytosine and 5-hydroxymethylcytosine at individual CpG dyads. Next, to understand how cell state transitions impact maintenance methylation, we scaled the method down to jointly profile genome-wide methylation levels, maintenance methylation fidelity and the transcriptome from single cells (scDyad&T-seq). Using scDyad&T-seq, we demonstrate that, while distinct cell states can substantially impact the activity of the maintenance methylation machinery, locally there exists an intrinsic relationship between DNA methylation density, histone modifications and DNMT1-mediated maintenance methylation fidelity that is independent of cell state.

Non-canonical functions of UHRF1 maintain DNA methylation homeostasis in cancer cells

DNA methylation is an essential epigenetic chromatin modification, and its maintenance in mammals requires the protein UHRF1. It is yet unclear if UHRF1 functions solely by stimulating DNA methylation maintenance by DNMT1, or if it has important additional functions. Using degron alleles, we show that UHRF1 depletion causes a much greater loss of DNA methylation than DNMT1 depletion. This is not caused by passive demethylation as UHRF1-depleted cells proliferate more slowly than DNMT1-depleted cells. Instead, bioinformatics, proteomics and genetics experiments establish that UHRF1, besides activating DNMT1, interacts with DNMT3A and DNMT3B and promotes their activity. In addition, we show that UHRF1 antagonizes active DNA demethylation by TET2. Therefore, UHRF1 has non-canonical roles that contribute importantly to DNA methylation homeostasis; these findings have practical implications for epigenetics in health and disease.

ICBP90, an epigenetic regulator, induces DKK3 promoter methylation, promotes glioma progression, and reduces sensitivity to cis-platinum

Glioma is the most common brain malignancy, characterized by high morbidity, high mortality, and treatment-resistance. Inverted CCAAT box Binding Protein of 90 kDa (ICBP90) has been reported to be involved in tumor progression and the maintenance of DNA methylation. Herein, we constructed ICBP90 over-expression and knockdown glioma cell lines, and found that ICBP90 knockdown inhibited glioma cell proliferation, migration, and invasion. ICBP90 silencing potentially enhanced cellular sensitivity to cis-platinum (DDP) and exacerbated DDP-induced pyroptosis, manifested by the elevated levels of gasdermin D-N-terminal and cleaved caspase 1; whereas, ICBP90 over-expression exhibited the opposite effects. Consistently, ICBP90 knockdown inhibited tumor growth in an in vivo mouse xenograft study using U251 cells stably expressing sh-ICBP90 and oe-ICBP90. Further experiments found that ICBP90 reduced the expression of Dickkopf 3 homolog (DKK3), a negative regulator of β-catenin, by binding its promoter and inducing DNA methylation. ICBP90 knockdown prevented the nuclear translocation of β‐catenin and suppressed the expression of c-Myc and cyclin D1. Besides, DKK3 over-expression restored the effects of ICBP90 over-expression on cell proliferation, migration, invasion, and DDP sensitivity. Our findings suggest that ICBP90 inhibits the expression of DKK3 in glioma by maintaining DKK3 promoter methylation, thereby conducing to ICBP90-mediated carcinogenesis and drug insensitivity.

Population epigenetics: DNA methylation in the plant omics era.

DNA methylation plays an important role in many biological processes. The mechanisms underlying the establishment and maintenance of DNA methylation are well understood thanks to decades of research using DNA methylation mutants, primarily in Arabidopsis (Arabidopsis thaliana) accession Col-0. Recent genome-wide association studies (GWASs) using the methylomes of natural accessions have uncovered a complex and distinct genetic basis of variation in DNA methylation at the population level. Sequencing following bisulfite treatment has served as an excellent method for quantifying DNA methylation. Unlike studies focusing on specific accessions with reference genomes, population-scale methylome research often requires an additional round of sequencing beyond obtaining genome assemblies or genetic variations from whole-genome sequencing data, which can be cost prohibitive. Here, we provide an overview of recently developed bisulfite-free methods for quantifying methylation and cost-effective approaches for the simultaneous detection of genetic and epigenetic information. We also discuss the plasticity of DNA methylation in a specific Arabidopsis accession, the contribution of DNA methylation to plant adaptation, and the genetic determinants of variation in DNA methylation in natural populations. The recently developed technology and knowledge will greatly benefit future studies in population epigenomes.

DNA hypomethylation activates Cdk4/6 and Atr to induce DNA replication and cell cycle arrest to constrain liver outgrowth in zebrafish.

Coordinating epigenomic inheritance and cell cycle progression is essential for organogenesis. UHRF1 connects these functions during development by facilitating maintenance of DNA methylation and cell cycle progression. Here, we provide evidence resolving the paradoxical phenotype of uhrf1 mutant zebrafish embryos which have activation of pro-proliferative genes and increased number of hepatocytes in S-phase, but the liver fails to grow. We uncover decreased Cdkn2a/b and persistent Cdk4/6 activation as the mechanism driving uhrf1 mutant hepatocytes into S-phase. This induces replication stress, DNA damage and Atr activation. Palbociclib treatment of uhrf1 mutants prevented aberrant S-phase entry, reduced DNA damage, and rescued most cellular and developmental phenotypes, but it did not rescue DNA hypomethylation, transposon expression or the interferon response. Inhibiting Atr reduced DNA replication and increased liver size in uhrf1 mutants, suggesting that Atr activation leads to dormant origin firing and prevents hepatocyte proliferation. Cdkn2a/b was downregulated pro-proliferative genes were also induced in a Cdk4/6 dependent fashion in the liver of dnmt1 mutants, suggesting DNA hypomethylation as a mechanism of Cdk4/6 activation during development. This shows that the developmental defects caused by DNA hypomethylation are attributed to persistent Cdk4/6 activation, DNA replication stress, dormant origin firing and cell cycle inhibition.

Antagonistic interactions safeguard mitotic propagation of genetic and epigenetic information in zebrafish

The stability of cellular phenotypes in developing organisms depends on error-free transmission of epigenetic and genetic information during mitosis. Methylation of cytosine residues in genomic DNA is a key epigenetic mark that modulates gene expression and prevents genome instability. Here, we report on a genetic test of the relationship between DNA replication and methylation in the context of the developing vertebrate organism instead of cell lines. Our analysis is based on the identification of hypomorphic alleles of dnmt1, encoding the DNA maintenance methylase Dnmt1, and pole1, encoding the catalytic subunit of leading-strand DNA polymerase epsilon holoenzyme (Pole). Homozygous dnmt1 mutants exhibit genome-wide DNA hypomethylation, whereas the pole1 mutation is associated with increased DNA methylation levels. In dnmt1/pole1 double-mutant zebrafish larvae, DNA methylation levels are restored to near normal values, associated with partial rescue of mutant-associated transcriptional changes and phenotypes. Hence, a balancing antagonism between DNA replication and maintenance methylation buffers against replicative errors contributing to the robustness of vertebrate development.

Epigenetics comprehensive experimental course based on the integration of science and education to cultivate students' ability of cutting-edge innovation.

The integration of science and education is an effective way for universities to cultivate students in cutting-edge innovative interests. Epigenetics is the expansion of classical genetics, the corresponding experimental courses of which have not been integrated into the current teaching system. In this paper, by taking advantage of our laboratory's research on the DNA methylation maintenance gene, OsMET1-2 in rice, we have integrated our innovative findings in the education curriculum, and built a comprehensive teaching system on experimentation research, which greatly stimulates the curiosity of the students. Taking the OsMET1-2 mutants and its isogenic wild-type rice plants as experimental materials, this course has successfully demonstrated a causal link between genetic mutation and epigenetic variation, a topic widely interested by the students in learning genetics and epigenetics. Through the practice of this course, students have a deeper understanding of the important role of epigenetic modifications, their scientific research capabilities have been greatly improved, thereby strongly supporting the cultivation of top innovative talents among the students.

MBD4 loss results in global reactivation of promoters and retroelements with low methylated CpG density

BackgroundInherited defects in the base-excision repair gene MBD4 predispose individuals to adenomatous polyposis and colorectal cancer, which is characterized by an accumulation of C > T transitions resulting from spontaneous deamination of 5’-methylcytosine.MethodsHere, we have investigated the potential role of MBD4 in regulating DNA methylation levels using genome-wide transcriptome and methylome analyses. Additionally, we have elucidated its function through a series of in vitro experiments.ResultsHere we show that the protein MBD4 is required for DNA methylation maintenance and G/T mismatch repair. Transcriptome and methylome analyses reveal a genome-wide hypomethylation of promoters, gene bodies and repetitive elements in the absence of MBD4 in vivo. Methylation mark loss is accompanied by a broad transcriptional derepression phenotype affecting promoters and retroelements with low methylated CpG density. MBD4 in vivo forms a complex with the mismatch repair proteins (MMR), which exhibits high bi-functional glycosylase/AP-lyase endonuclease specific activity towards methylated DNA substrates containing a G/T mismatch. Experiments using recombinant proteins reveal that the association of MBD4 with the MMR protein MLH1 is required for this activity.ConclusionsOur data identify MBD4 as an enzyme specifically designed to repair deaminated 5-methylcytosines and underscores its critical role in safeguarding against methylation damage. Furthermore, it illustrates how MBD4 functions in normal and pathological conditions.