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1.
Nat Commun ; 12(1): 5251, 2021 09 02.
Artigo em Inglês | MEDLINE | ID: mdl-34475392

RESUMO

DNA methylation (DNAm) is an epigenetic regulator of gene expression and a hallmark of gene-environment interaction. Using whole-genome bisulfite sequencing, we have surveyed DNAm in 344 samples of human postmortem brain tissue from neurotypical subjects and individuals with schizophrenia. We identify genetic influence on local methylation levels throughout the genome, both at CpG sites and CpH sites, with 86% of SNPs and 55% of CpGs being part of methylation quantitative trait loci (meQTLs). These associations can further be clustered into regions that are differentially methylated by a given SNP, highlighting the genes and regions with which these loci are epigenetically associated. These findings can be used to better characterize schizophrenia GWAS-identified variants as epigenetic risk variants. Regions differentially methylated by schizophrenia risk-SNPs explain much of the heritability associated with risk loci, despite covering only a fraction of the genomic space. We provide a comprehensive, single base resolution view of association between genetic variation and genomic methylation, and implicate schizophrenia GWAS-associated variants as influencing the epigenetic plasticity of the brain.


Assuntos
Metilação de DNA , Genoma Humano , Locos de Características Quantitativas/genética , Esquizofrenia/genética , Fatores Etários , Encéfalo/metabolismo , Encéfalo/patologia , Ilhas de CpG/genética , Epigênese Genética , Predisposição Genética para Doença/genética , Variação Genética , Estudo de Associação Genômica Ampla , Genótipo , Humanos , Polimorfismo de Nucleotídeo Único
2.
Transl Psychiatry ; 11(1): 416, 2021 08 02.
Artigo em Inglês | MEDLINE | ID: mdl-34341332

RESUMO

Depression is currently the leading cause of disability around the world. We conducted an epigenome-wide association study (EWAS) in a sample of 58 depression score-discordant monozygotic twin pairs, aiming to detect specific epigenetic variants potentially related to depression and further integrate with gene expression profile data. Association between the methylation level of each CpG site and depression score was tested by applying a linear mixed effect model. Weighted gene co-expression network analysis (WGCNA) was performed for gene expression data. The association of DNA methylation levels of 66 CpG sites with depression score reached the level of P < 1 × 10-4. These top CpG sites were located at 34 genes, especially PTPRN2, HES5, GATA2, PRDM7, and KCNIP1. Many ontology enrichments were highlighted, including Notch signaling pathway, Huntington disease, p53 pathway by glucose deprivation, hedgehog signaling pathway, DNA binding, and nucleic acid metabolic process. We detected 19 differentially methylated regions (DMRs), some of which were located at GRIK2, DGKA, and NIPA2. While integrating with gene expression data, HELZ2, PTPRN2, GATA2, and ZNF624 were differentially expressed. In WGCNA, one specific module was positively correlated with depression score (r = 0.62, P = 0.002). Some common genes (including BMP2, PRDM7, KCNIP1, and GRIK2) and enrichment terms (including complement and coagulation cascades pathway, DNA binding, neuron fate specification, glial cell differentiation, and thyroid gland development) were both identified in methylation analysis and WGCNA. Our study identifies specific epigenetic variations which are significantly involved in regions, functional genes, biological function, and pathways that mediate depression disorder.


Assuntos
Metilação de DNA , Gêmeos Monozigóticos , Biomarcadores , Ilhas de CpG/genética , DNA , Depressão , Epigênese Genética , Proteínas Hedgehog , Humanos , Transcriptoma , Gêmeos Monozigóticos/genética
3.
Nature ; 596(7870): 133-137, 2021 08.
Artigo em Inglês | MEDLINE | ID: mdl-34234345

RESUMO

The majority of gene transcripts generated by RNA polymerase II in mammalian genomes initiate at CpG island (CGI) promoters1,2, yet our understanding of their regulation remains limited. This is in part due to the incomplete information that we have on transcription factors, their DNA-binding motifs and which genomic binding sites are functional in any given cell type3-5. In addition, there are orphan motifs without known binders, such as the CGCG element, which is associated with highly expressed genes across human tissues and enriched near the transcription start site of a subset of CGI promoters6-8. Here we combine single-molecule footprinting with interaction proteomics to identify BTG3-associated nuclear protein (BANP) as the transcription factor that binds this element in the mouse and human genome. We show that BANP is a strong CGI activator that controls essential metabolic genes in pluripotent stem and terminally differentiated neuronal cells. BANP binding is repelled by DNA methylation of its motif in vitro and in vivo, which epigenetically restricts most binding to CGIs and accounts for differential binding at aberrantly methylated CGI promoters in cancer cells. Upon binding to an unmethylated motif, BANP opens chromatin and phases nucleosomes. These findings establish BANP as a critical activator of a set of essential genes and suggest a model in which the activity of CGI promoters relies on methylation-sensitive transcription factors that are capable of chromatin opening.


Assuntos
Proteínas de Ciclo Celular/metabolismo , Montagem e Desmontagem da Cromatina , Cromatina/genética , Cromatina/metabolismo , Ilhas de CpG/genética , Proteínas de Ligação a DNA/metabolismo , Proteínas Nucleares/metabolismo , Animais , Sequência de Bases , Linhagem Celular Tumoral , Cromatina/química , Montagem e Desmontagem da Cromatina/genética , Metilação de DNA , Regulação da Expressão Gênica , Genes Essenciais , Humanos , Camundongos , Imagem Individual de Molécula
4.
Nat Commun ; 12(1): 4549, 2021 07 27.
Artigo em Inglês | MEDLINE | ID: mdl-34315901

RESUMO

Germline pathogenic variants in DNMT3A were recently described in patients with overgrowth, obesity, behavioral, and learning difficulties (DNMT3A Overgrowth Syndrome/DOS). Somatic mutations in the DNMT3A gene are also the most common cause of clonal hematopoiesis, and can initiate acute myeloid leukemia (AML). Using whole genome bisulfite sequencing, we studied DNA methylation in peripheral blood cells of 11 DOS patients and found a focal, canonical hypomethylation phenotype, which is most severe with the dominant negative DNMT3AR882H mutation. A germline mouse model expressing the homologous Dnmt3aR878H mutation phenocopies most aspects of the human DOS syndrome, including the methylation phenotype and an increased incidence of spontaneous hematopoietic malignancies, suggesting that all aspects of this syndrome are caused by this mutation.


Assuntos
Anormalidades Múltiplas/genética , DNA (Citosina-5-)-Metiltransferases/genética , Epigênese Genética , Anormalidades Múltiplas/sangue , Adolescente , Adulto , Animais , Comportamento Animal , Peso Corporal/genética , Células da Medula Óssea/metabolismo , Criança , Pré-Escolar , Ilhas de CpG/genética , Metilação de DNA/genética , Feminino , Perfilação da Expressão Gênica , Mutação em Linhagem Germinativa/genética , Hematopoese/genética , Células-Tronco Hematopoéticas/metabolismo , Humanos , Lactente , Leucemia/genética , Leucemia/patologia , Masculino , Camundongos Endogâmicos C57BL , Obesidade/genética , Fenótipo , Síndrome , Transcrição Genética
5.
Int J Mol Sci ; 22(12)2021 Jun 15.
Artigo em Inglês | MEDLINE | ID: mdl-34203623

RESUMO

To increase the efficiency of assisted reproductive techniques (ART), molecular studies have been performed to identify the best predictive biomarkers for selecting the most suitable germ cells for fertilization and the best embryo for intra-uterine transfer. However, across different studies, no universal markers have been found. In this study, we addressed this issue by generating gene expression and CpG methylation profiles of outer cumulus cells obtained during intra-cytoplasmic sperm injection (ICSI). We also studied the association of the generated genomic data with the clinical parameters (spindle presence, zona pellucida birefringence, pronuclear pattern, estrogen level, endometrium size and lead follicle size) and the pregnancy result. Our data highlighted the presence of several parameters that affect analysis, such as inter-individual differences, inter-treatment differences, and, above all, specific treatment protocol differences. When comparing the pregnancy outcome following the long protocol (GnRH agonist) of ovarian stimulation, we identified the single gene markers (NME6 and ASAP1, FDR < 5%) which were also correlated with endometrium size, upstream regulators (e.g., EIF2AK3, FSH, ATF4, MKNK1, and TP53) and several bio-functions related to cell death (apoptosis) and cellular growth and proliferation. In conclusion, our study highlighted the need to stratify samples that are very heterogeneous and to use pathway analysis as a more reliable and universal method for identifying markers that can predict oocyte development potential.


Assuntos
Biomarcadores/metabolismo , Células do Cúmulo/metabolismo , Desenvolvimento Embrionário , Oócitos/metabolismo , Adulto , Ilhas de CpG/genética , Metilação de DNA/genética , Bases de Dados como Assunto , Feminino , Perfilação da Expressão Gênica , Regulação da Expressão Gênica , Humanos , Gravidez , Doadores de Tecidos
6.
Int J Mol Sci ; 22(12)2021 Jun 15.
Artigo em Inglês | MEDLINE | ID: mdl-34204008

RESUMO

Assisted reproductive technologies impact transcriptome and epigenome of embryos and can result in long-term phenotypic consequences. Whole-genome DNA methylation profiles from individual bovine blastocysts in vivo- and in vitro-derived (using three sources of protein: reproductive fluids, blood serum and bovine serum albumin) were generated. The impact of in vitro culture on DNA methylation was analyzed, and sex-specific methylation differences at blastocyst stage were uncovered. In vivo embryos showed the highest levels of methylation (29.5%), close to those produced in vitro with serum, whilst embryos produced in vitro with reproductive fluids or albumin showed less global methylation (25-25.4%). During repetitive element analysis, the serum group was the most affected. DNA methylation differences between in vivo and in vitro groups were more frequent in the first intron than in CpGi in promoters. Moreover, hierarchical cluster analysis showed that sex produced a stronger bias in the results than embryo origin. For each group, distance between male and female embryos varied, with in vivo blastocyst showing a lesser distance. Between the sexually dimorphic methylated tiles, which were biased to X-chromosome, critical factors for reproduction, developmental process, cell proliferation and DNA methylation machinery were included. These results support the idea that blastocysts show sexually-dimorphic DNA methylation patterns, and the known picture about the blastocyst methylome should be reconsidered.


Assuntos
Blastocisto/metabolismo , Reprogramação Celular/genética , Meios de Cultura/farmacologia , Epigênese Genética/efeitos dos fármacos , Caracteres Sexuais , Animais , Blastocisto/efeitos dos fármacos , Bovinos , Cromossomos de Mamíferos/genética , Ilhas de CpG/genética , Metilação de DNA/efeitos dos fármacos , Metilação de DNA/genética , Feminino , Fertilização In Vitro , Ontologia Genética , Modelos Logísticos , Masculino , Anotação de Sequência Molecular , Análise de Componente Principal
7.
Nat Commun ; 12(1): 3438, 2021 06 08.
Artigo em Inglês | MEDLINE | ID: mdl-34103501

RESUMO

DNA methylation plays a fundamental role in the control of gene expression and genome integrity. Although there are multiple tools that enable its detection from Nanopore sequencing, their accuracy remains largely unknown. Here, we present a systematic benchmarking of tools for the detection of CpG methylation from Nanopore sequencing using individual reads, control mixtures of methylated and unmethylated reads, and bisulfite sequencing. We found that tools have a tradeoff between false positives and false negatives and present a high dispersion with respect to the expected methylation frequency values. We described various strategies to improve the accuracy of these tools, including a consensus approach, METEORE ( https://github.com/comprna/METEORE ), based on the combination of the predictions from two or more tools that shows improved accuracy over individual tools. Snakemake pipelines are also provided for reproducibility and to enable the systematic application of our analyses to other datasets.


Assuntos
Ilhas de CpG/genética , Metilação de DNA/genética , Sequenciamento por Nanoporos , Benchmarking , Proteína 9 Associada à CRISPR/metabolismo , Citosina/metabolismo , DNA/metabolismo , Escherichia coli/genética , Genoma Bacteriano , Curva ROC
8.
J Psychiatr Res ; 140: 214-220, 2021 08.
Artigo em Inglês | MEDLINE | ID: mdl-34118639

RESUMO

Psychiatric symptoms are interrelated and found to be largely captured by a general psychopathology factor (GPF). Although epigenetic mechanisms, such as DNA methylation (DNAm), have been linked to individual psychiatric outcomes, associations with GPF remain unclear. Using data from 440 children aged 10 years participating in the Generation R Study, we examined the associations of DNAm with both general and specific (internalizing, externalizing) factors of psychopathology. Genome-wide DNAm levels, measured in peripheral blood using the Illumina 450K array, were clustered into wider co-methylation networks ('modules') using a weighted gene co-expression network analysis. One co-methylated module associated with GPF after multiple testing correction, while none associated with the specific factors. This module comprised of 218 CpG probes, of which 198 mapped onto different genes. The CpG most strongly driving the association with GPF was annotated to FZD1, a gene that has been implicated in schizophrenia and wider neurological processes. Associations between the probes contained in the co-methylated module and GPF were supported in an independent sample of children from the Avon Longitudinal Study of Parents and Children (ALSPAC), as evidenced by significant correlations in effect sizes. These findings might contribute to improving our understanding of dynamic molecular processes underlying complex psychiatric phenotypes.


Assuntos
Metilação de DNA , Transtornos Mentais , Criança , Ilhas de CpG/genética , Epigênese Genética , Genoma , Estudo de Associação Genômica Ampla , Humanos , Estudos Longitudinais , Transtornos Mentais/genética
9.
Cancer Sci ; 112(9): 3699-3710, 2021 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-34115910

RESUMO

Pyrosequencing (PSQ) represents the golden standard for MGMT promoter status determination. Binary interpretation of results based on the threshold from the average of several CpGs tested would neglect the existence of the "gray zone". How to define the gray zone and reclassify patients in this subgroup remains to be elucidated. A consecutive cohort of 312 primary glioblastoma patients were enrolled. CpGs 74-81 in the promoter region of MGMT were tested by PSQ and the protein expression was assessed by immunohistochemistry (IHC). Receiver operating characteristic curves were constructed to calculate the area under the curves (AUC). Kaplan-Meier plots were used to estimate the survival rate of patients compared by the log-rank test. The optimal threshold of each individual CpG differed from 5% to 11%. Patients could be separated into the hypomethylated subgroup (all CpGs tested below the corresponding optimal thresholds, n = 126, 40.4%), hypermethylated subgroup (all CpGs tested above the corresponding optimal thresholds, n = 108, 34.6%), and the gray zone subgroup (remaining patients, n = 78, 25.0%). Patients in the gray zone harbored an intermediate prognosis. The IHC score instead of the average methylation levels could successfully predict the prognosis for the gray zone (AUC for overall survival, 0.653 and 0.519, respectively). Combining PSQ and IHC significantly improved the efficiency of survival prediction (AUC: 0.662, 0.648, and 0.720 for PSQ, IHC, and combined, respectively). Immunohistochemistry is a robust method to predict prognosis for patients in the gray zone defined by PSQ. Combining PSQ and IHC could significantly improve the predictive ability for clinical outcomes.


Assuntos
Neoplasias Encefálicas/genética , Neoplasias Encefálicas/metabolismo , Metilases de Modificação do DNA/genética , Metilases de Modificação do DNA/metabolismo , Enzimas Reparadoras do DNA/genética , Enzimas Reparadoras do DNA/metabolismo , Glioblastoma/genética , Glioblastoma/metabolismo , Regiões Promotoras Genéticas/genética , Proteínas Supressoras de Tumor/genética , Proteínas Supressoras de Tumor/metabolismo , Adolescente , Adulto , Idoso , Neoplasias Encefálicas/mortalidade , Neoplasias Encefálicas/patologia , Ilhas de CpG/genética , Metilação de DNA , Feminino , Seguimentos , Glioblastoma/mortalidade , Glioblastoma/patologia , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Imuno-Histoquímica/métodos , Masculino , Pessoa de Meia-Idade , Prognóstico , Intervalo Livre de Progressão , Estudos Retrospectivos , Taxa de Sobrevida , Adulto Jovem
10.
Nat Commun ; 12(1): 3987, 2021 06 28.
Artigo em Inglês | MEDLINE | ID: mdl-34183656

RESUMO

Here we examine the association between DNA methylation in circulating leukocytes and blood lipids in a multi-ethnic sample of 16,265 subjects. We identify 148, 35, and 4 novel associations among Europeans, African Americans, and Hispanics, respectively, and an additional 186 novel associations through a trans-ethnic meta-analysis. We observe a high concordance in the direction of effects across racial/ethnic groups, a high correlation of effect sizes between high-density lipoprotein and triglycerides, a modest overlap of associations with epigenome-wide association studies of other cardio-metabolic traits, and a largely non-overlap with lipid loci identified to date through genome-wide association studies. Thirty CpGs reached significance in at least 2 racial/ethnic groups including 7 that showed association with the expression of an annotated gene. CpGs annotated to CPT1A showed evidence of being influenced by triglycerides levels. DNA methylation levels of circulating leukocytes show robust and consistent association with blood lipid levels across multiple racial/ethnic groups.


Assuntos
Metilação de DNA/genética , Leucócitos/citologia , Lipídeos/sangue , Lipoproteínas HDL/sangue , Adulto , Afro-Americanos , Idoso , Carnitina O-Palmitoiltransferase/genética , Ilhas de CpG/genética , Epigênese Genética , Epigenoma/genética , Epigenômica , Grupo com Ancestrais do Continente Europeu , Feminino , Hispano-Americanos , Humanos , Masculino , Pessoa de Meia-Idade , Locos de Características Quantitativas/genética
11.
Nat Commun ; 12(1): 3582, 2021 06 11.
Artigo em Inglês | MEDLINE | ID: mdl-34117224

RESUMO

In mouse development, long-term silencing by CpG island DNA methylation is specifically targeted to germline genes; however, the molecular mechanisms of this specificity remain unclear. Here, we demonstrate that the transcription factor E2F6, a member of the polycomb repressive complex 1.6 (PRC1.6), is critical to target and initiate epigenetic silencing at germline genes in early embryogenesis. Genome-wide, E2F6 binds preferentially to CpG islands in embryonic cells. E2F6 cooperates with MGA to silence a subgroup of germline genes in mouse embryonic stem cells and in embryos, a function that critically depends on the E2F6 marked box domain. Inactivation of E2f6 leads to a failure to deposit CpG island DNA methylation at these genes during implantation. Furthermore, E2F6 is required to initiate epigenetic silencing in early embryonic cells but becomes dispensable for the maintenance in differentiated cells. Our findings elucidate the mechanisms of epigenetic targeting of germline genes and provide a paradigm for how transient repression signals by DNA-binding factors in early embryonic cells are translated into long-term epigenetic silencing during mouse development.


Assuntos
Ilhas de CpG/genética , Fator de Transcrição E2F6/genética , Fator de Transcrição E2F6/metabolismo , Desenvolvimento Embrionário/genética , Epigênese Genética , Células Germinativas/metabolismo , Animais , Sítios de Ligação , Sistemas CRISPR-Cas , Diferenciação Celular , Metilação de DNA , Inativação Gênica , Camundongos , Camundongos Knockout , Células-Tronco Embrionárias Murinas , Complexo Repressor Polycomb 1/metabolismo , RNA Interferente Pequeno
12.
Nat Commun ; 12(1): 3428, 2021 06 08.
Artigo em Inglês | MEDLINE | ID: mdl-34103526

RESUMO

Dysregulated extravillous trophoblast invasion and proliferation are known to increase the risk of recurrent spontaneous abortion (RSA); however, the underlying mechanism remains unclear. Herein, in our retrospective observational case-control study we show that villous samples from RSA patients, compared to healthy controls, display reduced succinate dehydrogenase complex iron sulfur subunit (SDHB) DNA methylation, elevated SDHB expression, and reduced succinate levels, indicating that low succinate levels correlate with RSA. Moreover, we find high succinate levels in early pregnant women are correlated with successful embryo implantation. SDHB promoter methylation recruited MBD1 and excluded c-Fos, inactivating SDHB expression and causing intracellular succinate accumulation which mimicked hypoxia in extravillous trophoblasts cell lines JEG3 and HTR8 via the PHD2-VHL-HIF-1α pathway; however, low succinate levels reversed this effect and increased the risk of abortion in mouse model. This study reveals that abnormal metabolite levels inhibit extravillous trophoblast function and highlights an approach for RSA intervention.


Assuntos
Aborto Habitual/metabolismo , Vilosidades Coriônicas/metabolismo , Ácido Succínico/metabolismo , Aborto Habitual/enzimologia , Aborto Habitual/genética , Animais , Estudos de Casos e Controles , Hipóxia Celular , Linhagem Celular Tumoral , Ilhas de CpG/genética , Metilação de DNA/genética , Proteínas de Ligação a DNA/metabolismo , Feminino , Regulação da Expressão Gênica , Glicólise , Humanos , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Metaboloma , Camundongos Endogâmicos C57BL , Gravidez , Regiões Promotoras Genéticas/genética , Ligação Proteica , Proteínas Proto-Oncogênicas c-fos/metabolismo , Fatores de Risco , Succinato Desidrogenase/genética , Succinato Desidrogenase/metabolismo , Fatores de Transcrição/metabolismo , Transcrição Genética , Trofoblastos/metabolismo , Trofoblastos/patologia
13.
Aging (Albany NY) ; 13(11): 14630-14650, 2021 06 04.
Artigo em Inglês | MEDLINE | ID: mdl-34086604

RESUMO

Telomere length (TL) and telomere shortening are biological indicators of aging, and epigenetic associates have been found for TL in adults. However, the role of epigenetic signatures in setting newborn TL and early life telomere dynamics is unknown. In the present study, based on 247 participating newborns from the ENVIRONAGE birth cohort, whole-genome DNA methylation, profiled on the Illumina MethylationEPIC BeadChip microarray, and TL were measured in cord blood. In a follow-up visit at a mean age of 4.58 years, leukocyte TL was evaluated. We combined an epigenome-wide association study and a statistical learning method with re-sampling to select CpGs and their two-way interactions to model baseline (cord blood) TL and early-life telomere attrition rate, where distinct epigenetic signatures were identified for the two outcomes. In addition, a stronger epigenetic regulation was suggested in setting newborn TL than that of telomere dynamics in early life: 47 CpGs and 7 between-CpG interactions explained 76% of the variance in baseline TLs, while 72% of the total variance in telomere attrition rate was explained by 31 CpGs and 5 interactions. Functional enrichment analysis based on the selected CpGs in the two models revealed GLUT4 translocation and immune cell signaling pathways, respectively. These CpGs and interactions, as well as the cellular pathways, are potential novel targets of further investigation of telomere biology and aging.


Assuntos
Epigênese Genética , Perfilação da Expressão Gênica , Homeostase do Telômero/genética , Encurtamento do Telômero/genética , Pré-Escolar , Ilhas de CpG/genética , Metilação de DNA/genética , Feminino , Estudo de Associação Genômica Ampla , Humanos , Recém-Nascido , Masculino , Modelos Genéticos
14.
Nat Commun ; 12(1): 3892, 2021 06 23.
Artigo em Inglês | MEDLINE | ID: mdl-34162876

RESUMO

The decline of brain function during aging is associated with epigenetic changes, including DNA methylation. Lifestyle interventions can improve brain function during aging, but their influence on age-related epigenetic changes is unknown. Using genome-wide DNA methylation sequencing, we here show that experiencing a stimulus-rich environment counteracts age-related DNA methylation changes in the hippocampal dentate gyrus of mice. Specifically, environmental enrichment prevented the aging-induced CpG hypomethylation at target sites of the methyl-CpG-binding protein Mecp2, which is critical to neuronal function. The genes at which environmental enrichment counteracted aging effects have described roles in neuronal plasticity, neuronal cell communication and adult hippocampal neurogenesis and are dysregulated with age-related cognitive decline in the human brain. Our results highlight the stimulating effects of environmental enrichment on hippocampal plasticity at the level of DNA methylation and give molecular insights into the specific aspects of brain aging that can be counteracted by lifestyle interventions.


Assuntos
Envelhecimento , Ilhas de CpG/genética , Metilação de DNA , Meio Ambiente , Hipocampo/metabolismo , Fatores Etários , Animais , Giro Denteado/metabolismo , Epigenômica/métodos , Feminino , Hipocampo/citologia , Humanos , Camundongos Endogâmicos C57BL , Neurogênese/genética , Plasticidade Neuronal/genética , Neurônios/metabolismo
15.
Nat Commun ; 12(1): 2655, 2021 05 11.
Artigo em Inglês | MEDLINE | ID: mdl-33976121

RESUMO

The epigenome has been shown to deteriorate with age, potentially impacting on ageing-related disease. tRNA, while arising from only ˜46 kb (<0.002% genome), is the second most abundant cellular transcript. tRNAs also control metabolic processes known to affect ageing, through core translational and additional regulatory roles. Here, we interrogate the DNA methylation state of the genomic loci of human tRNA. We identify a genomic enrichment for age-related DNA hypermethylation at tRNA loci. Analysis in 4,350 MeDIP-seq peripheral-blood DNA methylomes (16-82 years), identifies 44 and 21 hypermethylating specific tRNAs at study-and genome-wide significance, respectively, contrasting with none hypomethylating. Validation and replication (450k array and independent targeted Bisuphite-sequencing) supported the hypermethylation of this functional unit. Tissue-specificity is a significant driver, although the strongest consistent signals, also independent of major cell-type change, occur in tRNA-iMet-CAT-1-4 and tRNA-Ser-AGA-2-6. This study presents a comprehensive evaluation of the genomic DNA methylation state of human tRNA genes and reveals a discreet hypermethylation with advancing age.


Assuntos
Envelhecimento/genética , Ilhas de CpG/genética , Metilação de DNA , Genoma Humano/genética , RNA de Transferência/genética , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Animais , Criança , Pré-Escolar , Feminino , Humanos , Masculino , Camundongos , Pessoa de Meia-Idade , Neoplasias/genética , Neoplasias/patologia , Especificidade de Órgãos/genética , Adulto Jovem
16.
Mol Immunol ; 135: 217-225, 2021 07.
Artigo em Inglês | MEDLINE | ID: mdl-33932686

RESUMO

Ly49Q is an ITIM-bearing MHC class I receptor that is highly expressed in plasmacytoid dendritic cells (pDCs). Ly49Q is required for the TLR9-mediated IFN-I production in pDCs, although the mechanism is not fully understood. We here demonstrate that Ly49Q protects pDCs from pyroptotic cell death induced by CpG oligodeoxynucleotides (CpG). In the Ly49Q-deficient (Klra17-/-) mouse spleen, the number of ssDNA-positive pDCs increased significantly after CpG treatment, strongly suggesting that Klra17-/- pDCs were susceptible to CpG-induced cell death. In Klra17-/- bone-marrow-derived dendritic cells (BMDCs), CpG-induced cell death was accompanied by increased cathepsin B leakage from the vesicular compartments into the cytoplasm. Concurrently, IL-1ß secretion increased in the CpG-treated Klra17-/- BMDCs, strongly suggesting that the CpG-induced cell death in these cells is pyroptotic in nature. Consistent with these observations, inhibiting cathepsin B or caspase 1 in CpG-stimulated Klra17-/- BMDCs reversed the increase in cell death. Pyroptotic cell death and IL-1ß secretion were also observed in BMDCs derived from transgenic mice expressing an ITIM-less Ly49Q (Ly49Q-YF Tg). CpG also increased the IL-1ß production and cell death in B2m-/- BMDCs. These results suggest that Ly49Q and MHC class I play important roles for protecting pyroptosis-like cell death of DCs by influencing lysosome state.


Assuntos
Células Dendríticas/imunologia , Lisossomos/metabolismo , Subfamília A de Receptores Semelhantes a Lectina de Células NK/metabolismo , Oligodesoxirribonucleotídeos/farmacologia , Piroptose/imunologia , Animais , Caspase 1/metabolismo , Catepsina B/metabolismo , Membrana Celular/fisiologia , Células Cultivadas , Ilhas de CpG/genética , Interleucina-1beta/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Subfamília A de Receptores Semelhantes a Lectina de Células NK/genética , Oligodesoxirribonucleotídeos/genética
17.
PLoS Biol ; 19(5): e3001229, 2021 05.
Artigo em Inglês | MEDLINE | ID: mdl-34003819

RESUMO

DNA methylation, chromatin accessibility, and gene expression represent different levels information in biological process, but a comprehensive multiomics analysis of the mammalian heart is lacking. Here, we applied nucleosome occupancy and methylome sequencing, which detected DNA methylation and chromatin accessibility simultaneously, as well as RNA-seq, for multiomics analysis of the 4 chambers of adult and fetal human hearts, and adult mouse hearts. Our results showed conserved region-specific patterns in the mammalian heart at transcriptome and DNA methylation level. Adult and fetal human hearts showed distinct features in DNA methylome, chromatin accessibility, and transcriptome. Novel long noncoding RNAs were identified in the human heart, and the gene expression profiles of major cardiovascular diseases associated genes were displayed. Furthermore, cross-species comparisons revealed human-specific and mouse-specific differentially expressed genes between the atria and ventricles. We also reported the relationship among multiomics and found there was a bell-shaped relationship between gene-body methylation and expression in the human heart. In general, our study provided comprehensive spatiotemporal and evolutionary insights into the regulation of gene expression in the heart.


Assuntos
Coração/crescimento & desenvolvimento , Coração/fisiologia , Animais , Cromatina/metabolismo , Ilhas de CpG/genética , DNA/genética , Metilação de DNA/genética , Epigênese Genética/genética , Epigenômica/métodos , Expressão Gênica/genética , Perfilação da Expressão Gênica/métodos , Ventrículos do Coração/crescimento & desenvolvimento , Ventrículos do Coração/metabolismo , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Humanos , Camundongos , Nucleossomos/metabolismo , Especificidade de Órgãos/genética , RNA Longo não Codificante/metabolismo , Especificidade da Espécie , Transcriptoma/genética
18.
Forensic Sci Int Genet ; 53: 102521, 2021 07.
Artigo em Inglês | MEDLINE | ID: mdl-33933877

RESUMO

The analysis of DNA methylation levels of specific CpG sites is one of the most promising molecular techniques to estimate an individual's age. Numerous studies were published recently presenting age estimation models based on DNA methylation patterns from blood samples, with only a few using saliva or buccal swabs. The aim of this study was to identify age-dependent methylation of 88 CpG sites in eight different marker regions (PDE4C, ELOVL2, ITGA2B, ASPA, EDARADD, SST, KLF14 and SLC12A5) in buccal swab samples. A total of 141 buccal swabs from individuals with age ranging from 21 to 69 years were split into a training set (n = 95) and a validation set (n = 46). Samples of the training set were analyzed by pyrosequencing and markers with best age correlation were identified. Stepwise linear regression analysis was performed resulting in an age estimation model including three of the examined CpG sites and showing a mean absolute deviation of estimated from chronological age of 5.11 years. To allow easy implementation into forensic laboratories without the need for pyrosequencing equipment, a multiplex minisequencing reaction was developed, including the same CpG sites previously identified by pyrosequencing. An adjusted age estimation model was evaluated with a mean absolute deviation of estimated from chronological age of 5.16 years. The independent validation set of 46 buccal swab samples was used to test model performances. Mean absolute deviation of estimated from chronological age was 5.33 years and 6.44 years for the pyrosequencing model and the minisequencing model, respectively. Comparison of the two methods showed a high concordance of results, both, qualitatively and quantitatively. In conclusion, buccal swabs offer a suitable alternative to blood samples for molecular age estimation with the additional advantage of being collected non-invasively. Furthermore we showed that minisequencing offers a cost-effective and easy-to-integrate alternative to pyrosequencing for the analysis of methylation status of individual CpG sites.


Assuntos
Envelhecimento/genética , Ilhas de CpG/genética , Metilação de DNA , Genética Forense/métodos , Sequenciamento de Nucleotídeos em Larga Escala , Adulto , Idoso , Feminino , Marcadores Genéticos , Humanos , Masculino , Pessoa de Meia-Idade , Mucosa Bucal , Reação em Cadeia da Polimerase Multiplex , Saliva/química , Análise de Sequência de DNA , Adulto Jovem
19.
Nat Genet ; 53(6): 794-800, 2021 06.
Artigo em Inglês | MEDLINE | ID: mdl-33986537

RESUMO

Precise deposition of CpG methylation is critical for mammalian development and tissue homeostasis and is often dysregulated in human diseases. The localization of de novo DNA methyltransferase DNMT3A is facilitated by its PWWP domain recognizing histone H3 lysine 36 (H3K36) methylation1,2 and is normally depleted at CpG islands (CGIs)3. However, methylation of CGIs regulated by Polycomb repressive complexes (PRCs) has also been observed4-8. Here, we report that DNMT3A PWWP domain mutations identified in paragangliomas9 and microcephalic dwarfism10 promote aberrant localization of DNMT3A to CGIs in a PRC1-dependent manner. DNMT3A PWWP mutants accumulate at regions containing PRC1-mediated formation of monoubiquitylated histone H2A lysine 119 (H2AK119ub), irrespective of the amounts of PRC2-catalyzed formation of trimethylated histone H3 lysine 27 (H3K27me3). DNMT3A interacts with H2AK119ub-modified nucleosomes through a putative amino-terminal ubiquitin-dependent recruitment region, providing an alternative form of DNMT3A genomic targeting that is augmented by the loss of PWWP reader function. Ablation of PRC1 abrogates localization of DNMT3A PWWP mutants to CGIs and prevents aberrant DNA hypermethylation. Our study implies that a balance between DNMT3A recruitment by distinct reader domains guides de novo CpG methylation and may underlie the abnormal DNA methylation landscapes observed in select human cancer subtypes and developmental disorders.


Assuntos
Ilhas de CpG/genética , DNA (Citosina-5-)-Metiltransferases/metabolismo , Metilação de DNA/genética , Proteínas do Grupo Polycomb/metabolismo , Animais , Catálise , Linhagem Celular , DNA (Citosina-5-)-Metiltransferases/química , Predisposição Genética para Doença , Genoma Humano , Histonas/metabolismo , Humanos , Lisina/metabolismo , Camundongos , Mutação/genética , Nucleossomos/metabolismo , Domínios Proteicos , Ubiquitinação
20.
J Forensic Sci ; 66(4): 1524-1532, 2021 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-33942892

RESUMO

It has already been proposed that a combined use of different molecular and morphological markers of aging in multivariate models may result in a greater accuracy of age estimation. However, such an approach can be complex and expensive, and not every combination may be useful. The significance and usefulness of combined analyses of D-aspartic acid in dentine, pentosidine in dentine, DNA methylation in buccal swabs at five genomic regions (PDE4C, RPA2, ELOVL2, DDO, and EDARADD), and third molar mineralization were tested by investigating a sample of 90 oral surgery patients. Machine learning models for age estimation were trained and evaluated, and the contribution of each parameter to multivariate models was tested by assessment of the predictor importance. For models based on D-aspartic acid, pentosidine, and the combination of both, mean absolute errors (MAEs) of 2.93, 3.41, and 2.68 years were calculated, respectively. The additional inclusion of the five DNAm markers did not improve the results. The sole DNAm-based model revealed a MAE of 4.14 years. In individuals under 28 years of age, the combination of the DNAm markers with the third molar mineralization stages reduced the MAE from 3.85 to 2.81 years. Our findings confirm that the combination of parameters in multivariate models may be very useful for age estimation. However, the inclusion of many parameters does not necessarily lead to better results. It is a task for future research to identify the best selection of parameters for the different requirements in forensic practice.


Assuntos
Determinação da Idade pelos Dentes/métodos , Adolescente , Adulto , Idoso , Arginina/análogos & derivados , Arginina/metabolismo , Biomarcadores/metabolismo , Criança , Ilhas de CpG/genética , Nucleotídeo Cíclico Fosfodiesterase do Tipo 4/metabolismo , D-Aspartato Oxidase/metabolismo , Ácido D-Aspártico/metabolismo , Metilação de DNA , Dentina/metabolismo , Proteína de Domínio de Morte Associada a Edar/metabolismo , Elongases de Ácidos Graxos/metabolismo , Humanos , Lisina/análogos & derivados , Lisina/metabolismo , Aprendizado de Máquina , Pessoa de Meia-Idade , Dente Serotino/crescimento & desenvolvimento , Análise Multivariada , Proteína de Replicação A/metabolismo , Calcificação de Dente , Adulto Jovem
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