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1.
Immunol Rev ; 305(1): 137-151, 2022 01.
Article in English | MEDLINE | ID: mdl-34935162

ABSTRACT

Epigenetic regulation of gene transcription in the immune system is important for proper control of protective and pathogenic inflammation. Aberrant epigenetic modifications are often associated with dysregulation of the immune cells, including lymphocytes and macrophages, leading to pathogenic inflammation and autoimmune diseases. Two classical epigenetic markers-histone modifications and DNA cytosine methylation, the latter is the 5 position of the cytosine base in the context of CpG dinucleotides-play multiple roles in the immune system. CxxC domain-containing proteins, which basically bind to the non-methylated CpG (i.e., epigenetic "readers"), often function as "writers" of the epigenetic markers via their catalytic domain within the proteins or by interacting with other epigenetic modifiers. We herein report the most recent advances in our understanding of the functions of CxxC domain-containing proteins in the immune system and inflammation, mainly focusing on T cells and macrophages.


Subject(s)
DNA Methylation , Epigenesis, Genetic , CpG Islands , DNA , Humans , Inflammation/genetics
2.
Sci Rep ; 12(1): 7494, 2022 May 06.
Article in English | MEDLINE | ID: mdl-35523936

ABSTRACT

Ovarian cancer is one of the lethal gynecologic cancers. Chemoresistance is an essential reason for treatment failure and high mortality. Emerging evidence connects epithelial-mesenchymal transition (EMT) like changes and acquisition of chemoresistance in cancers. Including EMT, DNA methylation influences cellular processes. Here, EMT-like changes were investigated in cisplatin-resistant A2780 ovarian cancer cells (A2780cis), wherein role of DNA methylation in some EMT genes regulations was studied. Cell viability assay was carried out to test the sensitivity of A2780, and A2780cis human cancer cell lines to cisplatin. Differential mRNA expression of EMT markers using qPCR was conducted to investigate EMT like changes. CpG methylation role in gene expression regulation was investigated by 5-azacytidine (5-aza) treatment. DNA methylation changes in EMT genes were identified using Methylscreen assay between A2780 and A2780cis cells. In order to evaluate if DNA methylation changes are causally underlying EMT, treatment with 5-aza followed by Cisplatin was done on A2780cis cells. Accordingly, morphological changes were studied under the microscope, whereas EMT marker's gene expression changes were investigated using qPCR. In this respect, A2780cis cell line has maintained its cisplatin tolerance ability and exhibits phenotypic changes congruent with EMT. Methylscreen assay and qPCR study have revealed DNA hypermethylation in promoters of epithelial adhesion molecules CDH1 and EPCAM in A2780cis compared to the cisplatin-sensitive parental cells. These changes were concomitant with gene expression down-regulation. DNA hypomethylation associated with transcription up-regulation of the mesenchymal marker TWIST2 was observed in the resistant cells. Azacytidine treatment confirmed DNA methylation role in regulating gene expression of CDH1, EPCAM and TWIST2 genes. A2780cis cell line undergoes EMT like changes, and EMT genes are regulated by DNA methylation. To that end, a better understanding of the molecular alterations that correlate with chemoresistance may lead to therapeutic benefits such as chemosensitivity restoration.


Subject(s)
CpG Islands , DNA Methylation , Epithelial-Mesenchymal Transition , Ovarian Neoplasms , Azacitidine/metabolism , Cell Line, Tumor , Cisplatin/therapeutic use , Drug Resistance, Neoplasm/genetics , Epithelial Cell Adhesion Molecule/metabolism , Epithelial-Mesenchymal Transition/genetics , Female , Gene Expression Regulation, Neoplastic , Humans , Ovarian Neoplasms/drug therapy , Ovarian Neoplasms/genetics , Ovarian Neoplasms/metabolism
3.
Acta Neuropathol Commun ; 10(1): 71, 2022 May 08.
Article in English | MEDLINE | ID: mdl-35527288

ABSTRACT

DNA methylation of cytosines in CpG sites throughout the genome is an epigenetic mark contributing to gene expression regulation. DNA methylation patterns are specific to tissue type, conserved throughout life and reflect changes during tumorigenesis. DNA methylation recently emerged as a diagnostic tool to classify tumors based on a combination of preserved developmental and mutation induced signatures. In addition to the tumor classification, DNA methylation data can also be used to evaluate copy number variation, assess promoter methylation status of specific genes, such as MGMT or MLH1, and deconvolute the tumor microenvironment, assessing the tumor immune infiltrate as a potential biomarker for immunotherapy. Here we review the role for DNA methylation in tumor diagnosis.


Subject(s)
DNA Methylation , Neoplasms , CpG Islands , DNA Copy Number Variations , DNA Modification Methylases/genetics , DNA Repair Enzymes/genetics , Humans , Mutation , Neoplasms/diagnosis , Neoplasms/genetics , Promoter Regions, Genetic , Tumor Microenvironment
4.
PLoS Genet ; 18(5): e1010181, 2022 May.
Article in English | MEDLINE | ID: mdl-35522715

ABSTRACT

Gene body methylation (GBM) is an ancestral mode of DNA methylation whose role in development has been obscured by the more prominent roles of promoter and CpG island methylation. The wasp Nasonia vitripennis has little promoter and CpG island methylation, yet retains strong GBM, making it an excellent model for elucidating the roles of GBM. Here we show that N. vitripennis DNA methyltransferase 1a (Nv-Dnmt1a) knockdown leads to failures in cellularization and gastrulation of the embryo. Both of these disrupted events are hallmarks of the maternal-zygotic transition (MZT) in insects. Analysis of the embryonic transcriptome and methylome revealed strong reduction of GBM and widespread disruption of gene expression during embryogenesis after Nv-Dnmt1a knockdown. Strikingly, there was a strong correlation between loss of GBM and reduced gene expression in thousands of methylated loci, consistent with the hypothesis that GBM directly facilitates high levels of transcription. We propose that lower expression levels of methylated genes due to reduced GBM is the crucial direct effect of Nv-Dnmt1 knockdown. Subsequently, the disruption of methylated genes leads to downstream dysregulation of the MZT, culminating in developmental failure at gastrulation.


Subject(s)
Wasps , Animals , CpG Islands/genetics , DNA Methylation/genetics , Genome , Wasps/genetics , Zygote/metabolism
5.
Nat Commun ; 13(1): 2408, 2022 May 03.
Article in English | MEDLINE | ID: mdl-35504910

ABSTRACT

We performed a multi-ethnic Epigenome Wide Association study on 22,774 individuals to describe the DNA methylation signature of chronic low-grade inflammation as measured by C-Reactive protein (CRP). We find 1,511 independent differentially methylated loci associated with CRP. These CpG sites show correlation structures across chromosomes, and are primarily situated in euchromatin, depleted in CpG islands. These genomic loci are predominantly situated in transcription factor binding sites and genomic enhancer regions. Mendelian randomization analysis suggests altered CpG methylation is a consequence of increased blood CRP levels. Mediation analysis reveals obesity and smoking as important underlying driving factors for changed CpG methylation. Finally, we find that an activated CpG signature significantly increases the risk for cardiometabolic diseases and COPD.


Subject(s)
DNA Methylation , Inflammation , C-Reactive Protein/genetics , CpG Islands/genetics , DNA Methylation/genetics , Humans , Inflammation/genetics , Nucleotide Motifs
6.
Methods Mol Biol ; 2432: 15-24, 2022.
Article in English | MEDLINE | ID: mdl-35505204

ABSTRACT

DNA methylation is a widely studied epigenetic phenomenon. Alterations in methylation patterns influence human phenotypes and risk of disease. The Illumina Infinium HumanMethylation450 (HM450) and MethylationEPIC (EPIC) BeadChip are widely used microarray-based platforms for epigenome-wide association studies (EWASs). In this chapter, we will discuss the use of intraclass correlation coefficient (ICC) for assessing technical variations induced by methylation arrays at single-CpG level. ICC compares variation of methylation levels within- and between-replicate measurements, ranging between 0 and 1. We further characterize the distribution of ICCs using a mixture of truncated normal and normal distributions, and cluster CpG sites on the arrays into low- and high-reliability groups. In practice, we recommend that extra caution needs to be taken for associations at the CpG sites with low ICC values.


Subject(s)
DNA Methylation , Epigenomics , CpG Islands , Oligonucleotide Array Sequence Analysis , Reproducibility of Results
7.
Int J Mol Sci ; 23(7)2022 Apr 05.
Article in English | MEDLINE | ID: mdl-35409379

ABSTRACT

Gene expression is controlled by epigenetic deregulation, a hallmark of cancer. The DNA methylome of canine diffuse large B-cell lymphoma (cDLBCL), the most frequent malignancy of B-lymphocytes in dog, has recently been investigated, suggesting that aberrant hypermethylation of CpG loci is associated with gene silencing. Here, we used a multi-omics approach (DNA methylome, transcriptome and copy number variations) combined with functional in vitro assays, to identify putative tumour suppressor genes subjected to DNA methylation in cDLBCL. Using four cDLBCL primary cell cultures and CLBL-1 cells, we found that CiDEA, MAL and PCDH17, which were significantly suppressed in DLBCL samples, were hypermethylated and also responsive (at the DNA, mRNA and protein level) to pharmacological unmasking with hypomethylating drugs and histone deacetylase inhibitors. The regulatory mechanism underneath the methylation-dependent inhibition of those target genes expression was then investigated through luciferase and in vitro methylation assays. In the most responsive CpG-rich regions, an in silico analysis allowed the prediction of putative transcription factor binding sites influenced by DNA methylation. Interestingly, regulatory elements for AP2, MZF1, NF-kB, PAX5 and SP1 were commonly identified in all three genes. This study provides a foundation for characterisation and experimental validation of novel epigenetically-dysregulated pathways in cDLBCL.


Subject(s)
DNA Copy Number Variations , DNA Methylation , Animals , Cell Line, Tumor , CpG Islands , Dogs , Gene Expression Regulation, Neoplastic , Gene Silencing , Genes, Tumor Suppressor
8.
Nucleic Acids Res ; 50(9): 4813-4839, 2022 May 20.
Article in English | MEDLINE | ID: mdl-35489059

ABSTRACT

Polycomb group proteins predominantly exist in polycomb repressive complexes (PRCs) that cooperate to maintain the repressed state of thousands of cell-type-specific genes. Targeting PRCs to the correct sites in chromatin is essential for their function. However, the mechanisms by which PRCs are recruited to their target genes in mammals are multifactorial and complex. Here we review DNA binding by polycomb group proteins. There is strong evidence that the DNA-binding subunits of PRCs and their DNA-binding activities are required for chromatin binding and CpG targeting in cells. In vitro, CpG-specific binding was observed for truncated proteins externally to the context of their PRCs. Yet, the mere DNA sequence cannot fully explain the subset of CpG islands that are targeted by PRCs in any given cell type. At this time we find very little structural and biophysical evidence to support a model where sequence-specific DNA-binding activity is required or sufficient for the targeting of CpG-dinucleotide sequences by polycomb group proteins while they are within the context of their respective PRCs, either PRC1 or PRC2. We discuss the current knowledge and open questions on how the DNA-binding activities of polycomb group proteins facilitate the targeting of PRCs to chromatin.


Subject(s)
Chromatin , DNA , Animals , Chromatin/genetics , CpG Islands/genetics , DNA/genetics , Mammals/genetics , Polycomb Repressive Complex 1/metabolism , Polycomb-Group Proteins/genetics , Polycomb-Group Proteins/metabolism
9.
Sci Rep ; 12(1): 5606, 2022 04 04.
Article in English | MEDLINE | ID: mdl-35379837

ABSTRACT

Handedness has low heritability and epigenetic mechanisms have been proposed as an etiological mechanism. To examine this hypothesis, we performed an epigenome-wide association study of left-handedness. In a meta-analysis of 3914 adults of whole-blood DNA methylation, we observed that CpG sites located in proximity of handedness-associated genetic variants were more strongly associated with left-handedness than other CpG sites (P = 0.04), but did not identify any differentially methylated positions. In longitudinal analyses of DNA methylation in peripheral blood and buccal cells from children (N = 1737), we observed moderately stable associations across age (correlation range [0.355-0.578]), but inconsistent across tissues (correlation range [- 0.384 to 0.318]). We conclude that DNA methylation in peripheral tissues captures little of the variance in handedness. Future investigations should consider other more targeted sources of tissue, such as the brain.


Subject(s)
DNA Methylation , Mouth Mucosa , Adult , Child , CpG Islands , Functional Laterality/genetics , Genome-Wide Association Study , Humans
10.
Clin Epigenetics ; 14(1): 53, 2022 Apr 19.
Article in English | MEDLINE | ID: mdl-35440009

ABSTRACT

BACKGROUND: DNA methylation is an epigenetic mechanism involved in human development. Numerous epigenome-wide association studies (EWAS) have investigated the associations of DNA methylation at single CpG sites with childhood outcomes. However, the overall contribution of DNA methylation across the genome (R2Methylation) towards childhood phenotypes is unknown. An estimate of R2Methylation would provide context regarding the importance of DNA methylation explaining variance in health outcomes. We therefore estimated the variance explained by epigenome-wide cord blood methylation (R2Methylation) for five childhood phenotypes: gestational age, birth weight, and body mass index (BMI), IQ and ADHD symptoms at school age. We adapted a genome-based restricted maximum likelihood (GREML) approach with cross-validation (CV) to DNA methylation data and applied it in two population-based birth cohorts: ALSPAC (n = 775) and Generation R (n = 1382). RESULTS: Using information from > 470,000 autosomal probes we estimated that DNA methylation at birth explains 32% (SDCV = 0.06) of gestational age variance and 5% (SDCV = 0.02) of birth weight variance. The R2Methylation estimates for BMI, IQ and ADHD symptoms at school age estimates were near 0% across almost all cross-validation iterations. CONCLUSIONS: The results suggest that cord blood methylation explains a moderate degree of variance in gestational age and birth weight, in line with the success of previous EWAS in identifying numerous CpG sites associated with these phenotypes. In contrast, we could not obtain a reliable estimate for school-age BMI, IQ and ADHD symptoms. This may reflect a null bias due to insufficient sample size to detect variance explained in more weakly associated phenotypes, although the true R2Methylation for these phenotypes is likely below that of gestational age and birth weight when using DNA methylation at birth.


Subject(s)
Epigenome , Individuality , Birth Weight/genetics , CpG Islands , DNA Methylation , Epigenesis, Genetic , Genome-Wide Association Study , Humans , Phenotype
11.
Oncogene ; 41(19): 2778-2785, 2022 May.
Article in English | MEDLINE | ID: mdl-35361883

ABSTRACT

In prostate cancers, elongation initiation factor 4A1 (eIF4A1) supports an oncogenic translation program and is highly expressed, but its role remains elusive. By the use of human specimens and cell models, we addressed the role of eIF4A1 in prostate cancer in vitro and in vivo. EIF4A1 expression, as determined by mRNA and protein levels, was higher in primary prostate cancers relative to normal prostate tissue. Also, for primary prostate cancers, elevated mRNA levels of EIF4A1 correlated with DNA hypomethylation levels in the CpG-rich island of EIF4A1. Using a DNMT3a CRISPR-Cas9-based tool for specific targeting of DNA methylation, we characterized, in human prostate cancer cells, the epigenetic regulation of EIF4A1 transcripts through DNA methylation in the CpG-rich island of EIF4A1. Next, we investigated the oncogenic effect of EIF4A1 on cancer cell proliferation in vitro and tumor growth in vivo. For prostate cancer cells, EIF4A1 heterozygous knockout or knockdown inhibited protein translation and tumor growth. In addition, using RNA immunoprecipitation with RNA sequencing, we discovered the eIF4A1-mediated translational regulation of the oncogene BRD2, which contains the most enriched eIF4A1-binding motifs in its 5' untranslated region, establishing an eIF4A1-BRD2 axis for oncogenic translation. Finally, we found a positive correlation between expression levels of eIF4A1 and BRD2 in primary prostate cancers. Our results demonstrate, for prostate cancer cells, epigenetic regulation of EIF4A1 transcripts through DNA methylation and an oncogenic role of eIF4A1 through BRD2 signaling.


Subject(s)
DNA Methylation , Eukaryotic Initiation Factor-4A/genetics , Prostatic Neoplasms , 5' Untranslated Regions , Carcinogenesis/genetics , CpG Islands , Epigenesis, Genetic , Gene Expression Regulation, Neoplastic , Humans , Male , Oncogenes , Peptide Initiation Factors/genetics , Prostatic Neoplasms/genetics , RNA, Messenger/genetics , Transcription Factors/genetics
12.
Arch Iran Med ; 25(3): 171-177, 2022 Mar 01.
Article in English | MEDLINE | ID: mdl-35429959

ABSTRACT

BACKGROUND: Medullary thyroid cancer (MTC) is a rare type of neuroendocrine tumor. This study aimed to investigate the gene and protein expression of RAP1GAP and DNA methylation patterns of its CpG74a , CpG74b , and CpG24 in an Iranian population with MTC. METHODS: In this case-control study, we selected 55 individuals who underwent thyroidectomy in Erfan hospital, Tehran, between 2018 and 2020. Samples were divided into normal thyroid tissues (control; n=20), benign nodule (n=20), and MTC (n=15). DNA methylation patterns were investigated using MSP (methylation-specific PCR). The protein level and mRNA expression of RAP1GAP were also evaluated using western blotting and real-time PCR, respectively. RESULTS: The hyper-methylation rates of CpG24 and CpG74a in the MTC samples were considerably higher than the controls (83% versus 15% and 74% versus 17%, respectively; P<0.001). The methylation/unmethylation ratio of CpG74a , and CpG24 was considerably higher than the controls (P<0.001). The methylation/unmethylation ratio of CpG24 in the benign nodules was also considerably greater than the controls (P<0.001). The mRNA expression and the protein level of RAP1GAP in the MTC group were considerably lower than the controls (P=0.005 and P=0.035, respectively). In the MTC group, aberrant methylation of CpG74a and CpG24 was significantly correlated with decreasing expression of the Rap1Gap gene (R2 : 0.23; P=0.032 and R2 : 0.56; P=0.001, respectively). CONCLUSION: Hyper-methylation in CpG24 and CpG74a and decreasing expression of RAP1GAP can be considered as diagnostic biomarkers for MTC.


Subject(s)
Carcinoma, Neuroendocrine , CpG Islands , GTPase-Activating Proteins , Thyroid Neoplasms , Carcinoma, Neuroendocrine/genetics , Carcinoma, Neuroendocrine/metabolism , Case-Control Studies , CpG Islands/genetics , DNA Methylation , GTPase-Activating Proteins/genetics , GTPase-Activating Proteins/metabolism , Humans , Iran , RNA, Messenger/genetics , RNA, Messenger/metabolism , Thyroid Neoplasms/genetics , Thyroid Neoplasms/metabolism
13.
Clin Epigenetics ; 14(1): 49, 2022 Apr 11.
Article in English | MEDLINE | ID: mdl-35410447

ABSTRACT

OBJECTIVE: Necrotizing enterocolitis (NEC) is the most common and lethal gastrointestinal disease affecting preterm infants. NEC develops suddenly and is characterized by gut barrier destruction, an inflammatory response, intestinal necrosis and multi-system organ failure. There is currently no method for early NEC detection, and the pathogenesis of NEC remains unclear. DESIGN: To further understand the molecular mechanisms that support NEC, we used solution phase hybridization and next-generation DNA sequencing of bisulfite converted DNA to perform targeted genome-wide analysis of DNA methylation at high read depth. RESULTS: We found that ileal samples from surgical NEC infants (n = 5) exist in a broadly hypermethylated state relative to their non-NEC counterparts (n = 9). These trends were not uniform, with hypermethylation being most consistently observed outside CpG islands and promoters. We further identified several biologically interesting gene promoters that displayed differential methylation in NEC and a number of biological pathways that appear dysregulated in NEC. We also found that DNA methylation patterns identified in ileal NEC tissue were correlated with those found and published previously in stool samples from NEC-affected infants. CONCLUSION: We confirmed that surgical NEC is associated with broad DNA hypermethylation in the ileum, and this may be detectable in stool samples of affected individuals. Thus, an epigenomic liquid biopsy of stool may have significant potential as a biomarker with respect to the diagnostic/predictive detection of NEC. Our findings, along with recent similar observations in colon, suggest that epigenomic dysregulation is a significant feature of surgical NEC. These findings motivate future studies which will involve the longitudinal screening of samples obtained prior to the onset of NEC. Our long-term goal is the development of novel screening, diagnostic and phenotyping methods for NEC.


Subject(s)
Enterocolitis, Necrotizing , Infant, Newborn, Diseases , CpG Islands , DNA Methylation , Enterocolitis, Necrotizing/diagnosis , Enterocolitis, Necrotizing/genetics , Humans , Infant , Infant, Newborn , Infant, Premature , Intestine, Small/pathology
14.
Mech Ageing Dev ; 204: 111676, 2022 Jun.
Article in English | MEDLINE | ID: mdl-35489615

ABSTRACT

The abundance of the biological data and the rapid evolution of the newer machine learning technologies have increased the epigenetics research in the last decade. This has enhanced the ability to measure the biological age of humans and different organisms via their omics data. DNA methylation array data are commonly used in the prediction of methylation age. Horvath clock has been adopted in various aging studies as a DNA methylation age predicting clock due to its higher accuracy and multi tissue prediction potential. In the current study, we have developed a pan tissue methylation-aging clock by using the publicly available illumina 450k and EPIC array methylation datasets. In doing that, we developed a highly accurate epigenetic clock, which predicts the age of multiple tissues with higher accuracy. We have also analyzed the selected probes for their biological relevance. Upon analyzing the selected features further, we found out evidences, which support the Antagonistic pleiotropy theory of aging.


Subject(s)
DNA Methylation , Epigenomics , Aging/genetics , CpG Islands , Epigenesis, Genetic , Epigenomics/methods , Humans , Machine Learning
15.
Transl Psychiatry ; 12(1): 132, 2022 Mar 31.
Article in English | MEDLINE | ID: mdl-35354798

ABSTRACT

Antenatal corticosteroids (ACS) are used to treat women at risk of preterm birth to improve neonatal survival. Though affected children may be at long-term risk of neurobehavioural disorders, the driving mechanisms remain unknown. Animal studies have shown that ACS exposure can lead to overlapping changes in DNA methylation between the blood and the brain, identifying gene pathways for neurodevelopment, which highlights the potential to examine peripheral blood as a surrogate for inaccessible human brain tissue. We hypothesized that differential methylation will be identified in blood of term-born neonates following ACS. Mother-infant dyads that received ACS were retrospectively identified through the Ontario Birth Study at Sinai Health Complex and matched to untreated controls for maternal age, BMI, parity and foetal sex (n = 14/group). Genome-wide methylation differences were examined at single-nucleotide resolution in DNA extracted from dried bloodspot cards using reduced representative bisulfite sequencing approaches. 505 differentially methylated CpG sites (DMCs) were identified, wherein 231 were hypermethylated and 274 were hypomethylated. These sites were annotated to 219 genes, of which USP48, SH3PXD2A, NTM, CAMK2N2, MAP6D1 were five of the top ten genes with known neurological function. Collectively, the set of hypermethylated genes were enriched for pathways of transcription regulation, while pathways of proteasome activity were enriched among the set of hypomethylated genes. This study is the first to identify DNA methylation changes in human neonatal blood following ACS. Understanding the epigenetic changes that occur in response to ACS will support future investigations to delineate the effects of prenatal glucocorticoid exposure on human development.


Subject(s)
DNA Methylation , Premature Birth , Adrenal Cortex Hormones/therapeutic use , CpG Islands , Female , Genome-Wide Association Study , Humans , Infant, Newborn , Mothers , Pregnancy , Premature Birth/genetics , Retrospective Studies , Sex Factors
16.
Clin Epigenetics ; 14(1): 37, 2022 03 10.
Article in English | MEDLINE | ID: mdl-35272673

ABSTRACT

BACKGROUND: DNA methylation alterations have emerged as hallmarks of cancer and have been proposed as screening, prognostic, and predictive biomarkers. Traditional approaches for methylation analysis have relied on bisulfite conversion of DNA, which can damage DNA and is not suitable for targeted gene analysis in low-input samples. Here, we have adapted methyl-CpG-binding domain protein 2 (MBD2)-based DNA enrichment for use on a semi-automated exclusion-based sample preparation (ESP) platform for robust and scalable enrichment of methylated DNA from low-input samples, called SEEMLIS. RESULTS: We show that combining methylation-sensitive enzyme digestion with ESP-based MBD2 enrichment allows for single gene analysis with high sensitivity for GSTP1 in highly impure, heterogenous samples. We also show that ESP-based MBD2 enrichment coupled with targeted pre-amplification allows for analysis of multiple genes with sensitivities approaching the single cell level in pure samples for GSTP1 and RASSF1 and sensitivity down to 14 cells for these genes in highly impure samples. Finally, we demonstrate the potential clinical utility of SEEMLIS by successful detection of methylated gene signatures in circulating tumor cells (CTCs) from patients with prostate cancer with varying CTC number and sample purity. CONCLUSIONS: SEEMLIS is a robust assay for targeted DNA methylation analysis in low-input samples, with flexibility at multiple steps. We demonstrate the feasibility of this assay to analyze DNA methylation in prostate cancer cells using CTCs from patients with prostate cancer as a real-world example of a low-input analyte of clinical importance. In summary, this novel assay provides a platform for determining methylation signatures in rare cell populations with broad implications for research as well as clinical applications.


Subject(s)
DNA Methylation , Prostatic Neoplasms , CpG Islands , DNA/metabolism , DNA-Binding Proteins/genetics , DNA-Binding Proteins/metabolism , Glutathione S-Transferase pi/genetics , Humans , Male , Prognosis , Prostatic Neoplasms/pathology
17.
Nat Methods ; 19(3): 296-306, 2022 03.
Article in English | MEDLINE | ID: mdl-35277705

ABSTRACT

Bulk-tissue DNA methylomes represent an average over many different cell types, hampering our understanding of cell-type-specific contributions to disease development. As single-cell methylomics is not scalable to large cohorts of individuals, cost-effective computational solutions are needed, yet current methods are limited to tissues such as blood. Here we leverage the high-resolution nature of tissue-specific single-cell RNA-sequencing datasets to construct a DNA methylation atlas defined for 13 solid tissue types and 40 cell types. We comprehensively validate this atlas in independent bulk and single-nucleus DNA methylation datasets. We demonstrate that it correctly predicts the cell of origin of diverse cancer types and discovers new prognostic associations in olfactory neuroblastoma and stage 2 melanoma. In brain, the atlas predicts a neuronal origin for schizophrenia, with neuron-specific differential DNA methylation enriched for corresponding genome-wide association study risk loci. In summary, the DNA methylation atlas enables the decomposition of 13 different human tissue types at a high cellular resolution, paving the way for an improved interpretation of epigenetic data.


Subject(s)
DNA Methylation , Epigenome , CpG Islands , Epigenesis, Genetic , Epigenomics , Genome-Wide Association Study , Humans , Neurons/metabolism
18.
BMC Biol ; 20(1): 70, 2022 03 23.
Article in English | MEDLINE | ID: mdl-35317801

ABSTRACT

BACKGROUND: Cytosine DNA methylation is a heritable epigenetic mark present in most eukaryotic groups. While the patterns and functions of DNA methylation have been extensively studied in mouse and human, their conservation in other vertebrates remains poorly explored. In this study, we interrogated the distribution and function of DNA methylation in primary fibroblasts of seven vertebrate species including bio-medical models and livestock species (human, mouse, rabbit, dog, cow, pig, and chicken). RESULTS: Our data highlight both divergence and conservation of DNA methylation patterns and functions. We show that the chicken genome is hypomethylated compared to other vertebrates. Furthermore, compared to mouse, other species show a higher frequency of methylation of CpG-rich DNA. We reveal the conservation of large unmethylated valleys and patterns of DNA methylation associated with X-chromosome inactivation through vertebrate evolution and make predictions of conserved sets of imprinted genes across mammals. Finally, using chemical inhibition of DNA methylation, we show that the silencing of germline genes and endogenous retroviruses (ERVs) are conserved functions of DNA methylation in vertebrates. CONCLUSIONS: Our data highlight conserved properties of DNA methylation in vertebrate genomes but at the same time point to differences between mouse and other vertebrate species.


Subject(s)
DNA Methylation , Epigenome , Animals , Cattle , CpG Islands , Dogs , Female , Genome , Germ Cells , Mammals/genetics , Mice , Rabbits , Swine/genetics , Vertebrates/genetics
19.
Gene ; 824: 146404, 2022 May 25.
Article in English | MEDLINE | ID: mdl-35278634

ABSTRACT

DNA methylation is an epigenetic mechanism that acts on cytosine residues. The methyl-CpG-binding domain proteins (MBD) are involved in the recognition of methyl-cytosines by activating a signaling cascade that induces the formation of heterochromatin or euchromatin, thereby regulating gene expression. In this study, we analyzed the evolution and conservation of MBD proteins in plants. First, we performed a genome-wide identification and analysis of the MBD family in common bean and soybean, since they have experienced one and two whole-genome duplication events, respectively. We found one pair of MBD paralogs in soybean (GmMBD2) has subfunctionalized after their recent divergence, which was corroborated with their expression profile. Phylogenetic analysis revealed that classes of MBD proteins clustered with human MBD. Interestingly, the MBD9 may have emerged after the hexaploidization event in eudicots. We found that plants and humans share a great similarity in MBDs' binding affinity in the mCpG context. MBD2 and MBD4 from different plant species have the conserved four amino acid residues -Arg (R), Asp (D), Tyr (Y) and Arg (R)- reported to be responsible for MBD-binding in the mCpG. However, MBD8, MBD9, MBD10, and MBD11 underwent substitutions in these residues, suggesting the non-interaction in the mCpG context, but a heterochromatin association as MBD5 and MBD6 from human. This study represents the first genome-wide analysis of the MBD gene family in eurosids I - soybean and common bean. The data presented here contribute towards understanding the evolution of MBDs proteins in plants and their specific binding affinity on mCpG site.


Subject(s)
DNA-Binding Proteins , Heterochromatin , CpG Islands/genetics , Cytosine , DNA Methylation , DNA-Binding Proteins/metabolism , Humans , Phylogeny , Plants/genetics , Plants/metabolism
20.
Sci Rep ; 12(1): 3574, 2022 03 04.
Article in English | MEDLINE | ID: mdl-35246549

ABSTRACT

Genetic and epigenetic modifications of genes involved in the key regulatory pathways play a significant role in the pathophysiology and progression of multifactorial diseases. The present study is an attempt to identify single nucleotide variations (SNVs) at CpG sites of promoters of ACAT1, APOB, APOE, CYBA, FAS, FLT1, KSR2, LDLR, MMP9, PCSK9, PHOX2A, REST, SH2B3, SORT1 and TIMP1 genes influencing CpG island (CGI) existence and size associated with the pathophysiology of Diabetes mellitus, Coronary artery disease and Cancers. Promoter sequences located between -2000 to + 2000 bp were retrieved from the EPDnew database and predicted the CpG island using MethPrimer. Further, SNVs at CpG sites were accessed from NCBI, Ensembl while transcription factor (TF) binding sites were accessed using AliBaba2.1. CGI existence and size were determined for each SNV at CpG site with respect to wild type and variant allele by MethPrimer. A total of 200 SNVs at CpG sites were analyzed from the promoters of ACAT1, APOB, APOE, CYBA, FAS, FLT1, KSR2, LDLR, MMP9, PCSK9, PHOX2A, REST, SH2B3, SORT1 and TIMP1 genes. Of these, only 17 (8.5%) SNVs were found to influence the loss of CGI while 70 (35%) SNVs were found to reduce the size of CGI. It has also been found that 59% (10) of CGI abolishing SNVs are showing differences in binding of TFs. The findings of the study suggest that the candidate SNVs at CpG sites regulating CGI existence and size might influence the DNA methylation status and expression of genes involved in molecular pathways associated with several diseases. The insights of the present study may pave the way for new experimental studies to undertake challenges in DNA methylation, gene expression and protein assays.


Subject(s)
Matrix Metalloproteinase 9 , Proprotein Convertase 9 , Apolipoproteins B/genetics , Apolipoproteins E/genetics , CpG Islands/genetics , DNA Methylation , Matrix Metalloproteinase 9/genetics , Nucleotides , Proprotein Convertase 9/genetics
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