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Nature Genetics Dec 2022Cytosine methylation efficiently silences CpG-rich regulatory regions of genes and repeats in mammalian genomes. To what extent this entails direct inhibition of...
Cytosine methylation efficiently silences CpG-rich regulatory regions of genes and repeats in mammalian genomes. To what extent this entails direct inhibition of transcription factor (TF) binding versus indirect inhibition via recruitment of methyl-CpG-binding domain (MBD) proteins is unclear. Here we show that combinatorial genetic deletions of all four proteins with functional MBDs in mouse embryonic stem cells, derived neurons or a human cell line do not reactivate genes or repeats with methylated promoters. These do, however, become activated by methylation-restricted TFs if DNA methylation is removed. We identify several causal TFs in neurons, including ONECUT1, which is methylation sensitive only at a motif variant. Rampantly upregulated retrotransposons in methylation-free neurons feature a CRE motif, which activates them in the absence of DNA methylation via methylation-sensitive binding of CREB1. Our study reveals methylation-sensitive TFs in vivo and argues that direct inhibition, rather than indirect repression by the tested MBD proteins, is the prevailing mechanism of methylation-mediated repression at regulatory regions and repeats.
Topics: Animals; Humans; Mice; DNA Methylation; Hepatocyte Nuclear Factor 6; Mammals; Transcription Factors
PubMed: 36471082
DOI: 10.1038/s41588-022-01241-6 -
Acta Neuropathologica Communications Jul 2023Chordomas are rare tumors of notochord remnants, occurring mainly in the sacrum and skull base. Despite of their unusually slow growth, chordomas are highly invasive and...
Chordomas are rare tumors of notochord remnants, occurring mainly in the sacrum and skull base. Despite of their unusually slow growth, chordomas are highly invasive and the involvement of adjacent critical structures causes treatment challenges. Due to the low incidence, the molecular pathogenesis of this entity remains largely unknown. This study aimed to investigate DNA methylation abnormalities and their impact on gene expression profiles in skull base chordomas. 32 tumor and 4 normal nucleus pulposus samples were subjected to DNA methylation and gene expression profiling with methylation microarrays and RNA sequencing. Genome-wide DNA methylation analysis revealed two distinct clusters for chordoma (termed subtypes C and I) with different patterns of aberrant DNA methylation. C Chordomas were characterized by general hypomethylation with hypermethylation of CpG islands, while I chordomas were generally hypermethylated. These differences were reflected by distinct distribution of differentially methylated probes (DMPs). Differentially methylated regions (DMRs) were identified, indicating aberrant methylation in known tumor-related genes in booth chordoma subtypes and regions encoding small RNAs in subtype C chordomas. Correlation between methylation and expression was observed in a minority of genes. Upregulation of TBXT in chordomas appeared to be related to lower methylation of tumor-specific DMR in gene promoter. Gene expression-based clusters of tumor samples did not overlap with DNA methylation-based subtypes. Nevertheless, they differ in transcriptomic profile that shows immune infiltration in I chordomas and up-regulation of cell cycle in C chordomas. Immune enrichment in chordomas I was confirmed with 3 independent deconvolution methods and immunohistochemistry. Copy number analysis showed higher chromosomal instability in C chordomas. Nine out of eight had deletion of CDKN2A/B loci and downregulation of genes encoded in related chromosomal band. No significant difference in patients' survival was observed between tumor subtypes, however, shorter survival was observed in patients with higher number of copy number alterations.
Topics: Humans; DNA Methylation; Chordoma; CpG Islands; Gene Expression Profiling; Sequence Analysis, RNA; Tumor Microenvironment
PubMed: 37434245
DOI: 10.1186/s40478-023-01610-0 -
Nature Reviews. Genetics Aug 2016Recent technological advances have made it possible to decode DNA methylomes at single-base-pair resolution under various physiological conditions. Many aberrant or... (Review)
Review
Recent technological advances have made it possible to decode DNA methylomes at single-base-pair resolution under various physiological conditions. Many aberrant or differentially methylated sites have been discovered, but the mechanisms by which changes in DNA methylation lead to observed phenotypes, such as cancer, remain elusive. The classical view of methylation-mediated protein-DNA interactions is that only proteins with a methyl-CpG binding domain (MBD) can interact with methylated DNA. However, evidence is emerging to suggest that transcription factors lacking a MBD can also interact with methylated DNA. The identification of these proteins and the elucidation of their characteristics and the biological consequences of methylation-dependent transcription factor-DNA interactions are important stepping stones towards a mechanistic understanding of methylation-mediated biological processes, which have crucial implications for human development and disease.
Topics: DNA; DNA Methylation; DNA-Binding Proteins; Humans; Transcription Factors; Transcription, Genetic
PubMed: 27479905
DOI: 10.1038/nrg.2016.83 -
Nature Communications Jun 2023O-Methylated stilbenes are prominent nutraceuticals but rarely produced by crops. Here, the inherent ability of two Saccharinae grasses to produce regioselectively...
O-Methylated stilbenes are prominent nutraceuticals but rarely produced by crops. Here, the inherent ability of two Saccharinae grasses to produce regioselectively O-methylated stilbenes is reported. A stilbene O-methyltransferase, SbSOMT, is first shown to be indispensable for pathogen-inducible pterostilbene (3,5-bis-O-methylated) biosynthesis in sorghum (Sorghum bicolor). Phylogenetic analysis indicates the recruitment of genus-specific SOMTs from canonical caffeic acid O-methyltransferases (COMTs) after the divergence of Sorghum spp. from Saccharum spp. In recombinant enzyme assays, SbSOMT and COMTs regioselectively catalyze O-methylation of stilbene A-ring and B-ring respectively. Subsequently, SOMT-stilbene crystal structures are presented. Whilst SbSOMT shows global structural resemblance to SbCOMT, molecular characterizations illustrate two hydrophobic residues (Ile144/Phe337) crucial for substrate binding orientation leading to 3,5-bis-O-methylations in the A-ring. In contrast, the equivalent residues (Asn128/Asn323) in SbCOMT facilitate an opposite orientation that favors 3'-O-methylation in the B-ring. Consistently, a highly-conserved COMT is likely involved in isorhapontigenin (3'-O-methylated) formation in wounded wild sugarcane (Saccharum spontaneum). Altogether, our work reveals the potential of Saccharinae grasses as a source of O-methylated stilbenes, and rationalize the regioselectivity of SOMT activities for bioengineering of O-methylated stilbenes.
Topics: Poaceae; Methylation; Phylogeny; Sorghum; Saccharum
PubMed: 37308495
DOI: 10.1038/s41467-023-38908-5 -
International Journal of Molecular... Dec 2020Methylation is a universal biochemical process which covalently adds methyl groups to a variety of molecular targets. It plays a critical role in two major global... (Review)
Review
Methylation is a universal biochemical process which covalently adds methyl groups to a variety of molecular targets. It plays a critical role in two major global regulatory mechanisms, epigenetic modifications and imprinting, via methyl tagging on histones and DNA. During reproduction, the two genomes that unite to create a new individual are complementary but not equivalent. Methylation determines the complementary regulatory characteristics of male and female genomes. DNA methylation is executed by methyltransferases that transfer a methyl group from S-adenosylmethionine, the universal methyl donor, to cytosine residues of CG (also designated CpG). Histones are methylated mainly on lysine and arginine residues. The methylation processes regulate the main steps in reproductive physiology: gametogenesis, and early and late embryo development. A focus will be made on the impact of assisted reproductive technology and on the impact of endocrine disruptors (EDCs) via generation of oxidative stress.
Topics: Animals; DNA Methylation; Embryonic Development; Epigenesis, Genetic; Gametogenesis; Histone Code; Humans; Reproductive Techniques, Assisted
PubMed: 33297303
DOI: 10.3390/ijms21239311 -
Molecular Cancer Jun 2024RNA methylation, a prevalent post-transcriptional modification, has garnered considerable attention in research circles. It exerts regulatory control over diverse... (Review)
Review
RNA methylation, a prevalent post-transcriptional modification, has garnered considerable attention in research circles. It exerts regulatory control over diverse biological functions by modulating RNA splicing, translation, transport, and stability. Notably, studies have illuminated the substantial impact of RNA methylation on tumor immunity. The primary types of RNA methylation encompass N6-methyladenosine (m6A), 5-methylcytosine (m5C), N1-methyladenosine (m1A), and N7-methylguanosine (m7G), and 3-methylcytidine (m3C). Compelling evidence underscores the involvement of RNA methylation in regulating the tumor microenvironment (TME). By affecting RNA translation and stability through the "writers", "erasers" and "readers", RNA methylation exerts influence over the dysregulation of immune cells and immune factors. Consequently, RNA methylation plays a pivotal role in modulating tumor immunity and mediating various biological behaviors, encompassing proliferation, invasion, metastasis, etc. In this review, we discussed the mechanisms and functions of several RNA methylations, providing a comprehensive overview of their biological roles and underlying mechanisms within the tumor microenvironment and among immunocytes. By exploring how these RNA modifications mediate tumor immune evasion, we also examine their potential applications in immunotherapy. This review aims to provide novel insights and strategies for identifying novel targets in RNA methylation and advancing cancer immunotherapy efficacy.
Topics: Humans; Neoplasms; Immunotherapy; Methylation; Tumor Microenvironment; Animals; RNA Processing, Post-Transcriptional; RNA; Gene Expression Regulation, Neoplastic; RNA Methylation
PubMed: 38902779
DOI: 10.1186/s12943-024-02041-8 -
Journal of Translational Medicine Feb 2021Neurotrophic tropomyosin receptor kinases (NTRKs) are a gene family function as oncogene or tumor suppressor gene in distinct cancers. We aimed to investigate the...
BACKGROUND
Neurotrophic tropomyosin receptor kinases (NTRKs) are a gene family function as oncogene or tumor suppressor gene in distinct cancers. We aimed to investigate the methylation and expression profiles and prognostic value of NTRKs gene in colorectal cancer (CRC).
METHODS
An analysis of DNA methylation and expression profiles in CRC patients was performed to explore the critical methylations within NTRKs genes. The methylation marker was validated in a retrospectively collected cohort of 229 CRC patients and tested in other tumor types from TCGA. DNA methylation status was determined by quantitative methylation-specific PCR (QMSP).
RESULTS
The profiles in six CRC cohorts showed that NTRKs gene promoter was more frequently methylated in CRC compared to normal mucosa, which was associated with suppressed gene expression. We identified a specific methylated region within NTRK3 promoter targeted by cg27034819 and cg11525479 that best predicted survival outcome in CRC. NTRK3 promoter methylation showed independently predictive value for survival outcome in the validation cohort (P = 0.004, HR 2.688, 95% CI [1.355, 5.333]). Based on this, a nomogram predicting survival outcome was developed with a C-index of 0.705. Furthermore, the addition of NTRK3 promoter methylation improved the performance of currently-used prognostic model (AIC: 516.49 vs 513.91; LR: 39.06 vs 43.64, P = 0.032). Finally, NTRK3 promoter methylation also predicted survival in other tumors, including pancreatic cancer, glioblastoma and stomach adenocarcinoma.
CONCLUSIONS
This study highlights the essential value of NTRK3 methylation in prognostic evaluation and the potential to improve current prognostic models in CRC and other tumors.
Topics: Biomarkers, Tumor; Colorectal Neoplasms; DNA Methylation; Humans; Prognosis; Receptor, trkC; Retrospective Studies; Tropomyosin
PubMed: 33593392
DOI: 10.1186/s12967-021-02740-6 -
BMC Genomics Dec 2023Atherosclerosis (AS) is a critical pathological event during the progression of cardiovascular diseases. It exhibits fibrofatty lesions on the arterial wall and lacks...
BACKGROUND
Atherosclerosis (AS) is a critical pathological event during the progression of cardiovascular diseases. It exhibits fibrofatty lesions on the arterial wall and lacks effective treatment. N-methyladenosine (mA) is the most common modification of eukaryotic RNA and plays an important role in regulating the development and progression of cardiovascular diseases. However, the role of mA modification in AS remains largely unknown. Therefore, in this study, we explored the transcriptome distribution of mA modification in AS and its potential mechanism.
METHODS
Methylation Quantification Kit was used to detect the global mA levels in the aorta of AS mice. Western blot was used to analyze the protein level of methyltransferases. Methylated RNA immunoprecipitation with next-generation sequencing (MeRIP-seq) and RNA sequencing (RNA-seq) were used to obtain the first transcriptome range analysis of the mA methylene map in the aorta of AS mice, followed by bioinformatics analysis. qRT-PCR and MeRIP-qRT-PCR were used to measure the mRNA and mA levels in target genes.
RESULTS
The global mA and protein levels of methyltransferase METTL3 were significantly increased in the aorta of AS mice. However, the protein level of demethylase ALKBH5 was significantly decreased. Through MeRIP-seq, we obtained mA methylation maps in AS and control mice. In total, 26,918 mA peaks associated with 13,744 genes were detected in AS group, whereas 26,157 mA peaks associated with 13,283 genes were detected in the control group. Peaks mainly appeared in the coding sequence (CDS) regions close to the stop codon with the RRACH motif. Moreover, functional enrichment analysis demonstrated that mA-containing genes were significantly enriched in AS-relevant pathways. Interestingly, a negative correlation between mA methylation abundance and gene expression level was found through the integrated analysis of MeRIP-seq and RNA-seq data. Among the mA-modified genes, a hypo-methylated but up-regulated (hypo-up) gene Fabp5 may be a potential biomarker of AS.
CONCLUSIONS
Our study provides transcriptome-wide mA methylation for the first time to determine the association between mA modification and AS progression. Our study lays a foundation for further exploring the pathogenesis of AS and provides a new direction for the treatment of AS.
Topics: Mice; Animals; Transcriptome; Cardiovascular Diseases; Methylation; RNA
PubMed: 38097926
DOI: 10.1186/s12864-023-09878-1 -
Bioinformatics (Oxford, England) Sep 2019The identification of differentially methylated regions (DMRs) among phenotypes is one of the main goals of epigenetic analysis. Although there are several methods...
MOTIVATION
The identification of differentially methylated regions (DMRs) among phenotypes is one of the main goals of epigenetic analysis. Although there are several methods developed to detect DMRs, most of them are focused on detecting relatively large differences in methylation levels and fail to detect moderate, but consistent, methylation changes that might be associated to complex disorders.
RESULTS
We present mCSEA, an R package that implements a Gene Set Enrichment Analysis method to identify DMRs from Illumina450K and EPIC array data. It is especially useful for detecting subtle, but consistent, methylation differences in complex phenotypes. mCSEA also implements functions to integrate gene expression data and to detect genes with significant correlations among methylation and gene expression patterns. Using simulated datasets we show that mCSEA outperforms other tools in detecting DMRs. In addition, we applied mCSEA to a previously published dataset of sibling pairs discordant for intrauterine hyperglycemia exposure. We found several differentially methylated promoters in genes related to metabolic disorders like obesity and diabetes, demonstrating the potential of mCSEA to identify DMRs not detected by other methods.
AVAILABILITY AND IMPLEMENTATION
mCSEA is freely available from the Bioconductor repository.
SUPPLEMENTARY INFORMATION
Supplementary data are available at Bioinformatics online.
Topics: CpG Islands; DNA Methylation; Promoter Regions, Genetic; Software
PubMed: 30753302
DOI: 10.1093/bioinformatics/btz096 -
International Journal of Molecular... Jun 2019Epigenetic modifications on individual bases in DNA and RNA can encode inheritable genetic information beyond the canonical bases. Among the nucleic acid modifications,... (Review)
Review
Epigenetic modifications on individual bases in DNA and RNA can encode inheritable genetic information beyond the canonical bases. Among the nucleic acid modifications, DNA N6-methadenine (6mA) and RNA N6-methyladenosine (m6A) have recently been well-studied due to the technological development of detection strategies and the functional identification of modification enzymes. The current findings demonstrate a wide spectrum of 6mA and m6A distributions from prokaryotes to eukaryotes and critical roles in multiple cellular processes. It is interesting that the processes of modification in which the methyl group is added to adenine and adenosine are the same, but the outcomes of these modifications in terms of their physiological impacts in organisms are quite different. In this review, we summarize the latest progress in the study of enzymes involved in the 6mA and m6A methylation machinery, including methyltransferases and demethylases, and their functions in various biological pathways. In particular, we focus on the mechanisms by which 6mA and m6A regulate the expression of target genes, and we highlight the future challenges in epigenetic regulation.
Topics: Adenosine; Animals; DNA Methylation; Epigenesis, Genetic; Humans; Methyltransferases; RNA
PubMed: 31208067
DOI: 10.3390/ijms20122931