DNMT1
DNA (cytosine-5)-methyltransferase 1 (EC 2.1.1.37) (Dnmt1) (CXXC-type zinc finger protein 9) (DNA methyltransferase HsaI) (DNA MTase HsaI) (M.HsaI) (MCMT) [AIM] [CXXC9] [DNMT]
Publications[править]
Increased life expectancy in modern society comes at the cost of age-associated disabilities and diseases. Aged brains not only show reduced excitability and plasticity, but also a decline in inhibition. Age-associated defects in inhibitory circuits likely contribute to cognitive decline and age-related disorders. Molecular mechanisms that exert epigenetic control of gene expression contribute to age-associated neuronal impairments. Both DNA methylation, mediated by DNA methyltransferases (DNMTs), and histone modifications maintain neuronal function throughout lifespan. Here we provide evidence that DNMT1 function is implicated in the age-related loss of cortical inhibitory interneurons. [i]Dnmt1[/i] deletion in parvalbumin-positive interneurons attenuates their age-related decline in the cerebral cortex. Moreover, conditional [i]Dnmt1[/i]-deficient mice show improved somatomotor performance and reduced aging-associated transcriptional changes. A decline in the proteostasis network, responsible for the proper degradation and removal of defective proteins, is implicated in age- and disease-related neurodegeneration. Our data suggest that DNMT1 acts indirectly on interneuron survival in aged mice by modulating the proteostasis network during life-time.
Keywords
- DNA methylation
- GABA
- aging
- cerebral cortex
- inhibitory interneurons
- proteostasis
- synapse
- transcriptional control
To understand the role of the extensive senescence-associated 3D genome reorganization, we generated genome-wide chromatin interaction maps, epigenome, replication-timing, whole-genome bisulfite sequencing, and gene expression profiles from cells entering replicative senescence (RS) or upon oncogene-induced senescence (OIS). We identify senescence-associated heterochromatin domains (SAHDs). Differential intra- versus inter-SAHD interactions lead to the formation of senescence-associated heterochromatin foci (SAHFs) in OIS but not in RS. This OIS-specific configuration brings active genes located in genomic regions adjacent to SAHDs in close spatial proximity and favors their expression. We also identify DNMT1 as a factor that induces SAHFs by promoting HMGA2 expression. Upon DNMT1 depletion, OIS cells transition to a 3D genome conformation akin to that of cells in replicative senescence. These data show how multi-omics and imaging can identify critical features of RS and OIS and discover determinants of acute senescence and SAHF formation.
MeSH Terms
- Cells, Cultured
- Cellular Senescence
- Chromatin Assembly and Disassembly
- DNA (Cytosine-5-)-Methyltransferase 1
- DNA Methylation
- Fibroblasts
- Genome, Human
- Heterochromatin
- Humans
- In Situ Hybridization, Fluorescence
- Oncogenes
Keywords
- 3D genome architecture
- DNMT1
- Hi-C
- chromatin compartments
- gene regulation
- oncogene-induced senescence
- replicative senescence
- senescence
The role of epigenetics in endothelial cell senescence is a cutting-edge topic in ageing research. However, little is known of the relative contribution to pro-senescence signal propagation provided by microRNAs shuttled by extracellular vesicles (EVs) released from senescent cells. Analysis of microRNA and DNA methylation profiles in non-senescent (control) and senescent (SEN) human umbilical vein endothelial cells (HUVECs), and microRNA profiling of their cognate small EVs (sEVs) and large EVs demonstrated that SEN cells released a significantly greater sEV number than control cells. sEVs were enriched in miR-21-5p and miR-217, which target DNMT1 and SIRT1. Treatment of control cells with SEN sEVs induced a miR-21/miR-217-related impairment of DNMT1-SIRT1 expression, the reduction of proliferation markers, the acquisition of a senescent phenotype and a partial demethylation of the locus encoding for miR-21. MicroRNA profiling of sEVs from plasma of healthy subjects aged 40-100 years showed an inverse U-shaped age-related trend for miR-21-5p, consistent with senescence-associated biomarker profiles. Our findings suggest that miR-21-5p/miR-217 carried by SEN sEVs spread pro-senescence signals, affecting DNA methylation and cell replication.
Keywords
- Cellular senescence
- DNMT1
- SIRT1
- extracellular vesicles
- microRNAs
Inflammation and fibroproliferative diseases may be modulated by epigenetic changes. Therefore, we suggest that epigenetic mechanisms could be involved in equine endometrosis pathogenesis. DNA methylation is one of the methods to evaluate epigenetics, through the transcription of methyltransferases (DNMT1, DNMT3A, DNMT3B). The correlation between DNMTs and collagen (COL) transcripts was assessed for the different Kenney and Doig's (Current Therapy in Theriogenology. Philadelphia: WB Saunders; 1986) endometrium categories. Endometrial biopsies were randomly collected from cyclic mares. Histological classification (category I, n = 13; II A, n = 17; II B, n = 12; and III, n = 7) and evaluation of COL1A2, COL3A1 and DNMTs transcripts by qPCR, were performed. Data were analysed by one-way analysis of variance (ANOVA), Kruskal-Wallis test and Pearson correlation. As mares aged, there was an increase in endometrium fibrosis (p < .01), and in DNMT1 mRNA (p < .001). Considering DNMT3B transcripts for each category, there was an increase with fibrosis (p < .05). No changes were observed for DNMT1 and DNMT3A transcripts. However, DNMT3A mRNA levels were the highest in all categories (p < .01). In category I endometrium, a positive correlation was observed for transcripts of all DNMTs in both COLs (p < .01). In category IIA, this correlation was also maintained for all DNMTs transcripts in COL1A2 (p < .05), but only for DNMT3B in COL3A1 (p < .05). In category IIB, there was a positive correlation between DNMT3B and COL3A1 (p < .05). In category III, a positive correlation was only observed between DNMT3B and COL3A1 (p < .05). Our results suggest that there is a disturbance in COLs and DNMTs correlation during fibrosis.
MeSH Terms
- Aging
- Animals
- Collagen
- DNA (Cytosine-5-)-Methyltransferases
- DNA Methylation
- Endometritis
- Endometrium
- Female
- Fibrosis
- Horse Diseases
- Horses
- RNA, Messenger
Keywords
- DNA methylation
- collagen
- endometrium
- epigenetic
- fibrosis
- mare
To investigate the relationship among natural aging of epidermal cells, epigenetics, and SPRY1 methylation mechanism. Immunohistochemistry, reverse transcription-PCR (RT-PCR), and Western blot were used to detect the expression of DNA methyltransferase 1 (DNMT1) and Sprouty1 (SPRY1) in skin epithelial cells from different age groups. An aging model of HaCaT cells was constructed. In HaCaT cells and their aging groups, DNMT1 and SPRY1 expression were detected by RT-PCR and WB. SPRY1 methylation status in epidermal cells from different age groups and HaCaT cells were detected by Methylation-Specific PCR (MS-PCR). The expression of DNMT1 and SPRY1 in skin epithelial cells from natural aging groups decreased with age; there was no significant difference in the expression of DNMT1 in HaCaT cells and the different age groups. The expression of SPRY1 in HaCaT cells was lower than it was in the aging groups. The methylation status of SPRY1 gradually decreased as the age of skin epidermal cells increased, while the methylation status of SPRY1 was not different between HaCaT cells and the aging group. DNMT1 is involved in the regulation of natural aging of skin epidermal cells but has a nominal role in our induced aging model. SPRY1 is involved in natural aging and induced aging of skin epidermal cells. The regulation of SPRY1 methylation is involved in the natural senescence of skin epidermal cells, while the induced aging of epidermal cells is nominally involved in the mechanism of SPRY1 methylation.
Keywords
- SPRY1
- methylation
- natural aging
- skin epidermal aging
Demyelination of axons plays an important role in the pathology of many spinal cord diseases and injuries. Remyelination in demyelinated lesions is primarily performed by oligodendrocyte progenitor cells (OPCs), which generate oligodendrocytes in the developing and mature central nervous system. The efficiency of remyelination decreases with age. Many reports suggest that this decline in remyelination results from impaired OPC recruitment and differentiation during aging. Of the various molecular mechanisms involved in aging, changes in epigenetic modifications have received particular attention. Global DNA methylation is a major epigenetic modification that plays important roles in cellular senescence and organismal aging. Thus, we aimed to evaluate the dynamic changes in the global DNA methylation profiles of OPCs derived from rat spinal cords during the aging process. We separated and cultured OPCs from the spinal cords of neonatal, 4-month-old, and 16-month-old rats and investigated the age-related alterations of genomic DNA methylation levels by using quantitative colorimetric analysis. To determine the potential cause of dynamic changes in global DNA methylation, we further analyzed the activity of DNA methyltransferases (DNMTs) and the expression of DNMT1, DNMT3a, DNMT3b, TET1, TET2, TET3, MBD2, and MeCP2 in the OPCs from each group. Our results showed the genomic DNA methylation level and the activity of DNMTs from OPCs derived from rat spinal cords decreased gradually during aging, and OPCs from 16-month-old rats were characterized by global hypomethylation. During OPC aging, the mRNA and protein expression levels of DNMT3a, DNMT3b, and MeCP2 were significantly elevated; those of DNMT1 were significantly down-regulated; and no significant changes were observed in those for TET1, TET2, TET3, or MBD2. Our results indicated that global DNA hypomethylation in aged OPCs is correlated with DNMT1 downregulation. Together, these data provide important evidence for partly elucidating the mechanism of age-related impaired OPC recruitment and differentiation and assist in the development of new treatments for promoting efficient remyelination.
MeSH Terms
- Aging
- Animals
- Animals, Newborn
- Cell Differentiation
- Cells, Cultured
- DNA Methylation
- DNA-Cytosine Methylases
- Epigenesis, Genetic
- Gene Expression Regulation
- Oligodendrocyte Precursor Cells
- Rats
- Spinal Cord
Keywords
- Duchenne muscular dystrophy
- T2 mapping
- fat infiltration
- skeletal muscle
Myeloid leukemia treatment is quite successful nowadays; nevertheless the development of new therapies is still necessary. In the present study, we investigated the potential of epigenetic modulators EGCG (epigallocatechin-3-gallate) and BIX-01294 (N-(1-benzylpiperidin-4-yl)-6,7-dimethoxy-2-(4-methyl-1,4-diazepan-1-yl)quinazolin-4-amine) to alter epigenetic state and cause cellular senescence in acute and chronic myeloid leukemia NB4 and K562 cells. We have shown that after leukemia cell treatment with EGCG and BIX-01294 the proliferation and survival were inhibited of both cell lines; however, only NB4 cells underwent apoptosis. Both epigenetic modulators caused cell cycle arrest in G0/G1 phase as assessed by RT-qPCR (p53, p21, Rb) and flow cytometry analysis. Increased levels of ATM, HMGA2, phosphorylated ATM, and SA-β-galactosidase staining indicated that EGCG caused cellular senescence, whereas BIX-01294 did not. Immunoblot analysis of epigenetic players DNMT1, HP1α, H3K9me3, EZH2, and SUZ12 demonstrated beneficial epigenetic modulation by both agents with exception of mainly no epigenetic changes caused in K562 cells by EGCG. Therefore, we suggest EGCG as a promising epigenetic modulator for acute promyelocytic leukemia therapy and as a potential cellular senescence inducer in both acute and chronic myeloid leukemia treatment, whereas BIX-01294 could be beneficial as an epigenetic modifier for both myeloid leukemias treatment.
MeSH Terms
- Antineoplastic Agents
- Azepines
- Catechin
- Cellular Senescence
- Drug Screening Assays, Antitumor
- Epigenesis, Genetic
- G1 Phase Cell Cycle Checkpoints
- Gene Expression Regulation, Neoplastic
- Humans
- K562 Cells
- Leukemia, Myelogenous, Chronic, BCR-ABL Positive
- Leukemia, Promyelocytic, Acute
- Quinazolines
Keywords
- BIX-01294
- EGCG
- Epigenetic regulation
- Myeloid leukemia
- Senescence
Induction of cellular senescence represents a novel strategy to inhibit aberrant proliferation of cancer cells. Resveratrol is gaining attention for its cancer preventive and suppressive properties. Tumor suppressor gene DLC1 is shown to induce apoptosis, suppress migration and invasion in various cancer cells. However, the function of DLC1 in cancer cellular senescence is unclear. This study was designed to investigate the biological role of DLC1 in resveratrol induced cancer cellular senescence. Our results showed that resveratrol inhibited proliferation of cancer cell lines (MCF-7, MDA-MB-231 and H1299) and induced senescence along with increase of SA-β-gal activity and regulation of senescence-associated molecular markers p38MAPK, p-p38MAPK, p27, p21, Rb and p-Rb protein. The underlying mechanism was that resveratrol induced mitochondrial dysfunction with reduction of mitochondrial membrane potential, down-regulation of MT-ND1, MT-ND6 and ATPase8 in transcript level and down-regulation of PGC-1α in protein level to result in ROS production. With ROS elevation, resveratrol decreased DNMT1 and increased DLC1 expression significantly. However, after ROS scavenger NAC was added to the cancer cells treated by resveratrol, DNMT1, DLC1 and senescence-associated molecular markers were reversed. This reveals that resveratrol induced cancer cellular senescence through DLC1 in a ROS-dependent manner. Silencing DLC1 markedly attenuated SA-β-gal activity and p38MAPK, p27 and p21 protein levels, and increased Rb expression, indicating that resveratrol promoted senescence via targeting DLC1. Moreover, DLC1 promoted senescence through FoxO3a/NF-κB signaling mediated by SIRT1 after resveratrol treatment. Finally, resveratrol increased ROS production to induce DNA damage with p-CHK1 up-regulation and result in cancer cellular senescence. This is the first time to investigate resveratrol induced cancer cellular senescence by primarily targeting DLC1. Induction of cellular senescence by resveratrol may represent a novel anticancer mechanism.
MeSH Terms
- Cellular Senescence
- DNA Damage
- GTPase-Activating Proteins
- Genes, Mitochondrial
- Humans
- Oxidative Stress
- Reactive Oxygen Species
- Resveratrol
- Signal Transduction
- Tumor Suppressor Proteins
- p38 Mitogen-Activated Protein Kinases
Keywords
- Cellular senescence
- DLC1
- Mitochondrial dysfunction
- Reactive oxygen species
- Resveratrol
- SIRT1
In this study, we report the role of DNA methyltransferase 1 (DNMT1) in ultraviolet A (UVA)-induced senescence in human dermal fibroblasts (HDFs). We show that DNMT1 expression was significantly reduced during UVA-induced senescence, and this senescence could be alleviated or aggravated by the up- or down-regulation of DNMT1, respectively. Expression of the transcription factor zinc finger E-box binding homeobox 1(ZEB1) also decreased after UVA irradiation, following a UVA-induced increase of intracellular reactive oxygen species (ROS). We show that ZEB1 binds to the DMNT1 promoter and regulates its transcription, which, in turn, affects cellular senescence. These changes in DMNT1 and ZEB1 expression following UVA exposure were confirmed in matched skin specimens that had or had not been sun-exposed. On analyzing the promoter methylation of 24 senescence associated genes in these matched skin specimens, we discovered that p53 promoter methylation was significantly reduced in sun-exposed skin. [i]In vitro[/i] experiments confirmed that UVA irradiation reduced p53 promoter methylation, and DNMT1 up-regulation could reverse this effect. Collectively, down-regulation of ZEB1 caused by UVA induced ROS could transcriptionally inhibit DNMT1, leading to low methylation level of senescence related proteins p53 and increase its expression, eventually result in cellar senescence.
MeSH Terms
- Cellular Senescence
- Down-Regulation
- Fibroblasts
- Humans
- Reactive Oxygen Species
- Repressor Proteins
- Skin Aging
- Skin Physiological Phenomena
- Tumor Suppressor Protein p53
- Ultraviolet Rays
- Up-Regulation
- Zinc Finger E-box-Binding Homeobox 1
Keywords
- DNMT1
- UVA
- ZEB1
- methylation
- senescence
DNA methyltransferase 1 (DNMT1) is a major epigenetic regulator associated with many biological processes. However, the roles and mechanisms of DNMT1 in skin aging are incompletely understood. Here we explored the role of DNMT1 in human skin fibroblasts senescence and its related regulatory mechanisms. DNMT1 expression decreased in passage-aged fibroblasts and DNMT1 silencing in young fibroblasts induced the senescence phenotype. MiR-217 is predicted to target DNMT1 mRNA and miR-217 expression increased in passage-aged fibroblasts. MiR-217 directly targeted the 3'-untranslated region (3'-UTR) of DNMT1 in HEK 293T cells and inhibited DNMT1 expression in fibroblasts. MiR-217 overexpression induced a senescence phenotype in young fibroblasts, and miR-217 downregulation in old HSFs partially reversed the senescence phenotype. However, these effects could be significantly rescued by regulating DNMT1 expression in fibroblasts. After regulating miR-217 levels, we analyzed changes in the promoter methylation levels of 24 senescent-associated genes, finding that 6 genes were significantly altered, and verified p16 and phosphorylated retinoblastoma (pRb) protein levels. Finally, an inverse correlation between DNMT1 and miR-217 expression was observed in skin tissues and different-aged fibroblasts. Together, these findings revealed that miR-217 promotes fibroblasts senescence by suppressing DNMT1-mediated methylation of p16 and pRb by targeting the DNMT1 3'-UTR.
MeSH Terms
- 3' Untranslated Regions
- Age Factors
- Aged
- Cell Line
- Cellular Senescence
- Child
- Child, Preschool
- Cyclin-Dependent Kinase Inhibitor p16
- DNA (Cytosine-5-)-Methyltransferase 1
- DNA Methylation
- Fibroblasts
- Gene Expression Regulation
- Gene Silencing
- Genes, Retinoblastoma
- Humans
- Infant
- Infant, Newborn
- MicroRNAs
- Promoter Regions, Genetic
- RNA Interference
- Skin
Keywords
- DNMT1
- miR-217
- senescence
- skin aging
Oxidative stress can induce cell dysfunction and lead to a broad range of degenerative alterations, including carcinogenesis, aging, and other oxidative stress-related conditions. To avoid undergoing carcinogenesis in response to oxidative stress, cells trigger a succession of checkpoint responses, including premature senescence and apoptosis. Increasing evidence indicates that H O , an important cause of oxidative stress, functions as an important physiological regulator of intracellular signaling pathways that participate in regulation of cell premature senescence and apoptosis. However, the precise mechanisms underlying this process remain to be studied extensively. In this study, we describe the importance of Pim-1 kinase in this checkpoint response to oxidative stress. Pim-1 binds to and phosphorylates the transcription factor high mobility group box transcription factor 1 (HBP1), activating it. H O enhances the interaction between Pim-1 and HBP1 and promotes HBP1 accumulation. In turn, HBP1 rapidly and selectively up-regulates Pim-1 expression in H O -stimulated cells, thereby creating a Pim-1-HBP1 positive feedback loop that regulates H O -induced premature senescence and apoptosis. Furthermore, the Pim-1-HBP1 positive feedback loop exerts its effect by regulating the senescence markers DNMT1 and p16 and the apoptosis marker Bax. The Pim-1-HBP1 axis thus constitutes a novel checkpoint pathway critical for the inhibition of tumorigenesis.
MeSH Terms
- Apoptosis
- Cell Cycle Checkpoints
- Cellular Senescence
- Cyclin-Dependent Kinase Inhibitor p16
- DNA (Cytosine-5-)-Methyltransferase 1
- DNA (Cytosine-5-)-Methyltransferases
- Gene Expression Regulation, Enzymologic
- HeLa Cells
- High Mobility Group Proteins
- Humans
- Hydrogen Peroxide
- Oxidative Stress
- Proto-Oncogene Proteins c-pim-1
- Repressor Proteins
- Up-Regulation
Keywords
- HBP1
- Pim-1
- apoptosis
- cellular senescence
- gene transcription
- hydrogen peroxide
- phosphorylation
- positive feedback loop
Skin aging is a complicated physiological process and epigenetic feature, including microRNA-mediated regulation and DNA methylation, have been shown to contribute to this process. DNA methylation is regulated by DNA methyltransferase, of which DNA methyltransferase 1 (DNMT1) is the most abundantly known. But evidence supporting its role in skin aging remains scarce, and no report regards its specifical upstream-regulating molecules in the process of skin aging so far. Here, we found that DNMT1 expression was markedly higher in young human skin fibroblasts (HSFs) than that in passage-aged HSFs, and DNMT1 knockdown significantly induced the senescence phenotype in young HSFs. We predicted the upstream miRNAs which could regulate DNMT1 with miRNA databases and found miR-377 had high homology with a sequence in the 3'-UTR of human DNMT1 mRNA. We confirmed that miR-377 was a potential regulator of DNMT1 by luciferase reporter assays. miR-377 expression in passage-aged HSFs was markedly higher than that in the young HSFs. miR-377 overexpression promoted senescence in young HSFs, and inhibition of miR-377 reduced senescence in passage-aged HSFs. Moreover, these functions were mediated by targeting DNMT1. Microfluidic PCR and next-generation bisulfite sequencing of 24 senescent-associated genes' promoters revealed alterations of the promoter methylation levels of FoxD3, p53, and UTF1 in HSFs treated with miR-377 mimics or inhibitors. We also verified that the miR-377-mediated changes in p53 expression could be reversed by regulation of DNMT1 in HSFs. Similarly, there was a negative correlation between miR-377 and DNMT1 expression in young and photoaged HSFs, HSFs, or skin tissues from UV-unexposed areas of different aged donors. Our results highlight a novel role for miR-377-DNMT1-p53 axis in HSF senescence. These findings shed new light on the mechanisms of skin aging and identify future opportunities for its therapeutic prevention.
MeSH Terms
- Aging
- DNA (Cytosine-5-)-Methyltransferase 1
- DNA (Cytosine-5-)-Methyltransferases
- DNA Methylation
- Fibroblasts
- Gene Expression Regulation
- Gene Knockdown Techniques
- High-Throughput Nucleotide Sequencing
- Humans
- MicroRNAs
- Promoter Regions, Genetic
- Skin
- Tumor Suppressor Protein p53
Insulin-producing pancreatic β cells in mice can slowly regenerate from glucagon-producing α cells in settings like β cell loss, but the basis of this conversion is unknown. Moreover, it remains unclear if this intra-islet cell conversion is relevant to diseases like type 1 diabetes (T1D). We show that the α cell regulators Aristaless-related homeobox (Arx) and DNA methyltransferase 1 (Dnmt1) maintain α cell identity in mice. Within 3 months of Dnmt1 and Arx loss, lineage tracing and single-cell RNA sequencing revealed extensive α cell conversion into progeny resembling native β cells. Physiological studies demonstrated that converted α cells acquire hallmark β cell electrophysiology and show glucose-stimulated insulin secretion. In T1D patients, subsets of glucagon-expressing cells show loss of DNMT1 and ARX and produce insulin and other β cell factors, suggesting that DNMT1 and ARX maintain α cell identity in humans. Our work reveals pathways regulated by Arx and Dnmt1 that are sufficient for achieving targeted generation of β cells from adult pancreatic α cells.
MeSH Terms
- Adult
- Aging
- Calcium Signaling
- Cell Lineage
- Child
- Child, Preschool
- DNA (Cytosine-5-)-Methyltransferase 1
- DNA (Cytosine-5-)-Methyltransferases
- Diabetes Mellitus, Type 1
- Electrophysiological Phenomena
- Female
- Gene Deletion
- Gene Expression Regulation
- Glucagon
- Glucagon-Secreting Cells
- Glucose
- Homeodomain Proteins
- Humans
- Insulin
- Insulin Secretion
- Insulin-Secreting Cells
- Male
- Sequence Analysis, RNA
- Single-Cell Analysis
- Transcription Factors
- Young Adult
As senescence develops, cells sequentially acquire diverse senescent phenotypes along with simultaneous multistage gene reprogramming. It remains unclear what acts as the key regulator of the collective changes in gene expression at initiation of senescent reprogramming. Here we analyzed time series gene expression profiles obtained in two different senescence models in human diploid fibroblasts: replicative senescence and H O -induced senescence. Our results demonstrate that suppression of DNA methyltransferase 1 (DNMT1)-mediated DNA methylation activity was an initial event prior to the display of senescent phenotypes. We identified seven DNMT1-interacting proteins, ubiquitin-like with PHD and ring finger domains 1 (UHRF1), EZH2, CHEK1, SUV39H1, CBX5, PARP1, and HELLS (also known as LSH (lymphoid-specific helicase) 1), as being commonly down-regulated at the same time point as DNMT1 in both senescence models. Knockdown experiments revealed that, among the DNMT1-interacting proteins, only UHRF1 knockdown suppressed DNMT1 transcription. However, UHRF1 overexpression alone did not induce DNMT1 expression, indicating that UHRF1 was essential but not sufficient for DNMT1 transcription. Although UHRF1 knockdown effectively induced senescence, this was significantly attenuated by DNMT1 overexpression, clearly implicating the UHRF1/DNMT1 axis in senescence. Bioinformatics analysis further identified WNT5A as a downstream effector of UHRF1/DNMT1-mediated senescence. Senescence-associated hypomethylation was found at base pairs -1569 to -1363 from the transcription start site of the WNT5A gene in senescent human diploid fibroblasts. As expected, WNT5A overexpression induced senescent phenotypes. Overall, our results indicate that decreased UHRF1 expression is a key initial event in the suppression of DNMT1-mediated DNA methylation and in the consequent induction of senescence via increasing WNT5A expression.
MeSH Terms
- CCAAT-Enhancer-Binding Proteins
- Cellular Senescence
- DNA (Cytosine-5-)-Methyltransferase 1
- DNA (Cytosine-5-)-Methyltransferases
- DNA Methylation
- Fibroblasts
- Gene Expression Profiling
- Gene Expression Regulation
- HEK293 Cells
- Histones
- Humans
- Hydrogen Peroxide
- Male
- Oligonucleotide Array Sequence Analysis
- Phenotype
- Promoter Regions, Genetic
- Protein Binding
- Protein Domains
- RNA, Small Interfering
- Ubiquitin-Protein Ligases
- Wnt-5a Protein
- beta-Galactosidase
Keywords
- DNA methylation
- cellular senescence
- gene expression
- gene regulation
- microarray
Despite well-documented evidence for lifespan extension by methionine restriction (MR), underlying mechanisms remain unknown. As methionine can alter S-adenosylmethionine (SAM) and S-adenosylhomocysteine (SAH), the substrate and product of DNA methyltransferase-1 (DNMT1), we hypothesized that MR diet alters DNA methylation. Young (8-week-old) and adult (1-year-old) male C57BL/6J mice were fed diets with different levels of methionine (0.12%-MR, 0.84%-CD) for 12weeks. Functional indicators of DNA methylation, including global methylation (GM), gene-specific methylation (GSM) and LINE-1 methylation; and biochemical factors affecting DNA methylation, SAH, SAM, and DNMT1 were assessed in different tissues. MR altered DNA methylation depending on the age of intervention. While MR had no effect on hepatic GM in young animals, it increased GM by 27% over CD in adults (p<0.01). In comparison with young animals, hepatic GM levels were 17% lower in CD adults (p<0.05), but not different in MR adults. The MR-induced increase in hepatic GM was associated with a 38% decrease in SAH levels in adults (p<0.001), with SAH and GM levels being negatively correlated (r =0.33, p<0.001). No changes were observed in DNMT protein levels in liver. In adipose tissue, MR caused a 6% decline in GM in adults (p<0.05), a corresponding 2-fold increase in SAH (p<0.05), and a 2-fold decrease in DNMT1 (p<0.01). MR caused both increases and decreases in GSM of liver and adipose. No changes were observed in LINE-1. Together, these findings provide evidence for protective effects of MR diet on hepatic DNA hypomethylation in adults, apparently mediated by SAH. These findings also indicate that altered DNA methylation might be playing a role in benefits conferred by MR diet.
MeSH Terms
- Aging
- Animals
- Caloric Restriction
- DNA Methylation
- Diet
- Liver
- Male
- Methionine
- Mice
- Mice, Inbred C57BL
- Time Factors
Keywords
- Aging
- Caloric restriction
- DNA methylation
- Methionine
- One-carbon cycle
- Sulfur amino acids
The present work was designed to study the mechanisms associated with Nurr1 modulation following early life permethrin (PERM) treatment during rat's life span. Here we demonstrate that PERM exposure in rats, at a dose close to No Observed Adverse Effect Level (NOAEL) for 15days during neonatal brain development leads to its accumulation long after exposure. In striatum from adolescent rats we detected an increase in DNA methyltransferases (DNMTs) such as DNMT1, DNMT3a, Tyrosine hydroxylase, monomeric and aggregated α-synuclein protein levels. Adult rats showed enhanced DNMT3b and α-synuclein aggregation compared to the control group, while with aging a significant decrease in all biomarkers studied was observed. No changes in Nurr1 promoter methylation in adolescent, adult and old rats were found. In silico studies showed clear evidence of a strong binding interaction between PERM and its metabolite 3-phenoxybenzoic acid with the nuclear orphan receptor Nurr1. These findings suggest that an additional interference with the dopaminergic neuron pathway could occur in situ during PERM accumulation in brain. Therefore, Nurr1 modulation in early life PERM-treated rats, depends on age-related adaptive responses in animals.
MeSH Terms
- Aging
- Animals
- Benzoates
- Binding Sites
- Corpus Striatum
- DNA Methylation
- DNA Modification Methylases
- Male
- Molecular Docking Simulation
- Molecular Dynamics Simulation
- Nuclear Receptor Subfamily 4, Group A, Member 2
- Permethrin
- Promoter Regions, Genetic
- Protein Multimerization
- Rats, Wistar
- alpha-Synuclein
Keywords
- DNMTs
- Nurr1 promoter methylation
- early life permethrin exposure
- molecular docking
- rat
- α-synuclein
This study was conducted in order to explore age-related changes in the production of Th1 and Th2 cytokines and determine the corresponding status of DNA methylation. The plasma IL-4 and IFN-γ levels and expression of Th-related cytokines and transcription factors in CD4 splenocytes were observed in mice at different weeks of age. The DNA methylation levels of IL-4 and IFN-γ promoters and the related regulatory regions in CD4 splenocytes of mice at different weeks of age were analyzed. The DNA methyltransferase (DNMT) levels in CD4 splenocytes of mice were analyzed. Changes in plasma IL-4 and IFN-γ levels after 5-AZA injection were evaluated. Plasma IL-4 and IL-4 expression in CD4 splenocytes declined with increasing age, while the IFN-γ expression levels increased. Th-related transcription factors showed no differences in mice at different weeks of age. The DNMT1 and DNMT3b mRNA expression did not show significant changes in CD4 splenocytes, whereas the DNMT3a mRNA expression increased with age. DNA methylation in the IL-4 promoter was increased, while DNA methylation in the IFN-γ promoter was decreased. The methylation of RSH7, CNS-1, and HSV increased significantly with age. Age-related changes in DNA methylation may be associated with the shift in Th1/Th2 balance.
MeSH Terms
- Aging
- Animals
- Cytokines
- DNA Methylation
- Interferon-gamma
- Interleukin-4
- Mice
- Promoter Regions, Genetic
- Regulatory Sequences, Nucleic Acid
- Th1 Cells
- Th1-Th2 Balance
- Th2 Cells
Keywords
- T helper cell
- epigenetics
- methylation
Senescence is a natural anticancer defense program disabled in tumor cells. We discovered that deregulated CDK4 (cyclin dependant kinase 4) and CDK6 activities contribute to senescence bypass during tumorigenesis and that their inhibition restores the senescence response in tumor cells. CDK4 and CDK6 phosphorylate RB1/RB, preventing its inhibitory interaction with the E2Fs, the cell cycle transcription factors. However, we also found that CDK4 interacts and phosphorylates the DNMT1 (DNA methyltransferase 1) protein protecting it from macroautophagy/autophagy-mediated protein degradation. This discovery highlights a new epigenetic component of CDK4-CDK6 signaling that could be exploited in cancer treatment.
MeSH Terms
- Autophagy
- Cellular Senescence
- Clinical Trials as Topic
- Cyclin-Dependent Kinase 4
- Cyclin-Dependent Kinase 6
- DNA (Cytosine-5-)-Methyltransferase 1
- DNA (Cytosine-5-)-Methyltransferases
- Humans
- Models, Biological
- Neoplasms
- Protein Kinase Inhibitors
- Proteolysis
Keywords
- CDK4
- DNMT1
- PML
- autophagy
- cancer
- epigenetic
- senescence
Early life lead (Pb) exposure results in a latent increase in Alzheimer's disease (AD)-related proteins, and cognitive deficits late in life in both rodents and primates. This study was conducted to investigate if these late life changes were accompanied by epigenetic alterations. Western blot analysis and RT-PCR were used to measure Deoxyribonucleic acid methylation regulators (DNMT1, DNMT3a, MeCP2, MAT2A) and histone proteins (H3K9Ac, H3K4me2, H3K27me3). Cerebral levels of DNMT1 and MeCP2 were significantly reduced in mice exposed to Pb early in life, whereas the expression of DNMT3a was not altered. Levels of MAT2a were increased in the Pb-exposed mice across the lifespan. H3K9Ac and H3K4me2, involved in gene activation, were decreased, whereas the repressive mark H3K27me3 was elevated. Epigenetic modifiers are affected by the developmental exposure to Pb and may play a role in mediating the latent increases in AD-related proteins in the brain.
Keywords
- Aging
- Alzheimer's disease
- Epigenetics
- Lead (Pb)
- Lifespan
Aging is associated with alterations in the content and patterns of DNA methylation virtually throughout the entire human lifespan. Reasons for these variations are not well understood. However, several lines of evidence suggest that the epigenetic instability in aging may be traced back to the alteration of the expression of DNA methyltransferases. Here, the association of the expression of DNA methyltransferases DNMT1 and DNMT3B with age has been analysed in the context of the MARK-AGE study, a large-scale cross-sectional study of the European general population. Using peripheral blood mononuclear cells, we assessed the variation of DNMT1 and DNMT3B gene expression in more than two thousand age-stratified women and men (35-75 years) recruited across eight European countries. Significant age-related changes were detected for both transcripts. The level of DNMT1 gradually dropped with aging but this was only observed up to the age of 64 years. By contrast, the expression of DNMT3B decreased linearly with increasing age and this association was particularly evident in females. We next attempted to trace the age-related changes of both transcripts to the influence of different variables that have an impact on changes of their expression in the population, including demographics, dietary and health habits, and clinical parameters. Our results indicate that age affects the expression of DNMT1 and DNMT3B as an almost independent variable in respect of all other variables evaluated.
MeSH Terms
- Adult
- Aged
- Aging
- Body Mass Index
- DNA (Cytosine-5-)-Methyltransferase 1
- DNA (Cytosine-5-)-Methyltransferases
- European Continental Ancestry Group
- Female
- Gene Expression Regulation, Developmental
- Gene Expression Regulation, Enzymologic
- Gene Ontology
- Humans
- Leukocytes, Mononuclear
- Life Style
- Male
- Middle Aged
- RNA, Messenger
- Regression Analysis
- Risk Factors
Keywords
- DNA methylation
- DNMT1
- DNMT3B
- aging
SIRT1 is likely to play a role in the extension in healthspan induced by dietary restriction. Actions of SIRT1 are pleiotropic, and effects on healthspan may include effects on DNA methylation. Polycomb group protein target genes (PCGTs) are suppressed by epigenetic mechanisms in stem cells, partly through the actions of the polycomb repressive complexes (PRCs), and have been shown previously to correspond with loci particularly susceptible to age-related changes in DNA methylation. We hypothesised that SIRT1 would affect DNA methylation particularly at PCGTs. To map the sites in the genome where SIRT1 affects DNA methylation, we altered SIRT1 expression in human intestinal (Caco-2) and vascular endothelial (HuVEC) cells by transient transfection with an expression construct or with siRNA. DNA was enriched for the methylated fraction then sequenced (HuVEC) or hybridised to a human promoter microarray (Caco-2). The profile of genes where SIRT1 manipulation affected DNA methylation was enriched for PCGTs in both cell lines, thus supporting our hypothesis. SIRT1 knockdown affected the mRNA for none of seven PRC components nor for DNMT1 or DNMT3b. We thus find no evidence that SIRT1 affects DNA methylation at PCGTs by affecting the expression of these gene transcripts. EZH2, a component of PRC2 that can affect DNA methylation through association with DNA methyltransferases (DNMTs), did not co-immunoprecipitate with SIRT1, and SIRT1 knockdown did not affect the expression of EZH2 protein. Thus, it is unlikely that the effects of SIRT1 on DNA methylation at PCGTs are mediated through direct intermolecular association with EZH2 or through effects in its expression. SIRT1 affects DNA methylation across the genome, but particularly at PCGTs. Although the mechanism through which SIRT1 has these effects is yet to be uncovered, this action is likely to contribute to extended healthspan, for example under conditions of dietary restriction.
MeSH Terms
- Aging
- Caco-2 Cells
- DNA (Cytosine-5-)-Methyltransferase 1
- DNA (Cytosine-5-)-Methyltransferases
- DNA Methylation
- DNA-Binding Proteins
- Enhancer of Zeste Homolog 2 Protein
- Epigenesis, Genetic
- Gene Expression Regulation
- Humans
- Polycomb Repressive Complex 2
- Polycomb-Group Proteins
- Promoter Regions, Genetic
- Sirtuin 1
We discovered the expression level of miR-148a significantly decreased in pancreatic cancer tissues whereas that of DNMT1 increased. In ASPC-1 cancer cells, the overexpression of miR-148a led to a decreased level of DNMT1 and reduced the proliferation and metastasis of ASPC-1 cells. Moreover, the increased expression of miR-148a arrested the UTR methylation of p27, giving rise to an increased level of p27. Interestingly, it was shown that the DNMT1 inhibition enhanced the expression of miR-148a. In vivo studies demonstrated that the tumorigenesis of ASPC-1 was significantly arrested by either the overexpression of miR-148a or the inhibition of DNMT1.
MeSH Terms
- Animals
- Cell Line, Tumor
- Cyclin-Dependent Kinase Inhibitor p27
- DNA (Cytosine-5-)-Methyltransferase 1
- DNA (Cytosine-5-)-Methyltransferases
- DNA Methylation
- Gene Expression Regulation, Neoplastic
- Humans
- Mice
- MicroRNAs
- Molecular Targeted Therapy
- Neoplasm Metastasis
- Neoplasm Transplantation
- Pancreatic Neoplasms
- Untranslated Regions
Keywords
- DNMT1
- Pancreatic cancer
- Senescence
- miR-148a
- p27
Methyltransferase expression and DNA methylation are linked to aging and age-related disease. We utilized 3-, 12-, and 24-month-old Ames dwarf and their wild-type siblings to examine the genotype and age-related differences in the expression of methyltransferase enzymes related to DNA methylation in the liver, glycine-N-methyltransferase and DNA methyltransferase (DNMT). We found that DNMT proteins and transcripts are differentially expressed in dwarf mice compared with wild-type siblings that can be attributed to age and/or genotype. However, DNMT1 protein expression is drastically reduced compared with wild-type controls at every age. DNMT3a protein levels coincide with differences observed in DNMT activity. Growth hormone appears to modulate expression of DNMT1 and 3a in dwarf liver tissue and primary hepatocytes. Therefore, growth hormone may contribute to age-related processes, DNA methylation, and, ultimately, longevity.
MeSH Terms
- Animals
- Colorimetry
- DNA (Cytosine-5-)-Methyltransferases
- DNA Methylation
- Dwarfism, Pituitary
- Glycine N-Methyltransferase
- Growth Hormone
- Hepatocytes
- Immunoblotting
- Longevity
- Methyltransferases
- Mice
- Mice, Inbred Strains
- Repressor Proteins
Keywords
- Aging
- Ames dwarf mice
- DNA methylation
- DNA methyltransferase
- Glycine-N-methyltransferase
- Liver.
Regular exercise improves learning and memory, including during aging process. Interestingly, the imbalance of epigenetic mechanisms has been linked to age-related cognitive deficits. However, studies about epigenetic alterations after exercise during the aging process are rare. In this preliminary study we investigated the effect of aging and exercise on DNA methyltransferases (DNMT1 and DNMT3b) and H3-K9 methylation levels in hippocampus from 3 and 20-months aged Wistar rats. The animals were submitted to two exercise protocols: single session or chronic treadmill protocol. DNMT1 and H3-K9 methylation levels were decreased in hippocampus from aged rats. The single exercise session decreased both DNMT3b and DNMT1 levels in young adult rats, without any effect in the aged group. Both exercise protocols reduced H3-K9 methylation levels in young adult rats, while the single session reversed the changes on H3-K9 methylation levels induced by aging. Together, these results suggest that an imbalance on DNMTs and H3-K9 methylation levels might be linked to the brain aging process and that the outcome to exercise seems to vary through lifespan.
MeSH Terms
- Age Factors
- Aging
- Animals
- Biomarkers
- DNA (Cytosine-5-)-Methyltransferase 1
- DNA (Cytosine-5-)-Methyltransferases
- DNA Methylation
- Epigenesis, Genetic
- Hippocampus
- Histones
- Lysine
- Male
- Physical Exertion
- Rats
- Rats, Wistar
DNA methylation is a major control program that modulates gene expression in a plethora of organisms. Gene silencing through methylation occurs through the activity of DNA methyltransferases, enzymes that transfer a methyl group from S-adenosyl-L-methionine to the carbon 5 position of cytosine. DNA methylation patterns are established by the de novo DNA methyltransferases (DNMTs) DNMT3A and DNMT3B and are subsequently maintained by DNMT1. Aging and age-related diseases include defined changes in 5-methylcytosine content and are generally characterized by genome-wide hypomethylation and promoter-specific hypermethylation. These changes in the epigenetic landscape represent potential disease biomarkers and are thought to contribute to age-related pathologies, such as cancer, osteoarthritis, and neurodegeneration. Some diseases, such as a hereditary form of sensory neuropathy accompanied by dementia, are directly caused by methylomic changes. Epigenetic modifications, however, are reversible and are therefore a prime target for therapeutic intervention. Numerous drugs that specifically target DNMTs are being tested in ongoing clinical trials for a variety of cancers, and data from finished trials demonstrate that some, such as 5-azacytidine, may even be superior to standard care. DNMTs, demethylases, and associated partners are dynamically shaping the methylome and demonstrate great promise with regard to rejuvenation.
MeSH Terms
- Aging
- Animals
- DNA Methylation
- DNA Modification Methylases
- Epigenesis, Genetic
- Humans
- Rejuvenation
The expression of 5-lipoxygenase (5-LOX) is affected by aging and regulated by epigenetic mechanisms including DNA methylation. We used methylation-sensitive restriction endonucleases (AciI, BstUI, HpaII, and HinP1I) to assess 5-LOX DNA methylation in brain and heart tissue samples from young (2 months) and old (22 months) mice. We also measured mRNA content for 5-LOX and the DNA methyltransferases DNMT1 and DNMT3a. In young mice, the 5-LOX mRNA content was significantly greater in the heart compared to the brain; 5-LOX DNA methylation was lower, except in the AciI assay in which it was higher in the heart. Aging decreased 5-LOX mRNA content in the heart and increased it in the brain. Aging also increased 5-LOX DNA methylation and this effect was site- (i.e., enzyme) and tissue-specific. Generally, DNMT1 and DNMT3a mRNA content was lower in the brain regions compared to the heart; the only effect of aging was observed in the mRNA content of DNMT3a, which was decreased in the heart of old mice. These results indicate a complex tissue-specific and aging-dependent interplay between the DNA methylation system and 5-LOX mRNA content. Interpretation of this data must take into account that the tissue samples contained a mixture of various cell types.
MeSH Terms
- Aging
- Animals
- Arachidonate 5-Lipoxygenase
- Brain
- DNA (Cytosine-5-)-Methyltransferase 1
- DNA (Cytosine-5-)-Methyltransferases
- DNA Methylation
- Down-Regulation
- Epigenesis, Genetic
- Gene Silencing
- Inflammation
- Leukotrienes
- Lipoxins
- Male
- Mice
- Mice, Inbred C57BL
- Myocardium
- Promoter Regions, Genetic
- RNA, Messenger
- Restriction Mapping
Estrogens play a critical role in brain development by acting on areas that express estrogen receptors. In the rodent cortex, estrogen receptor alpha (ER alpha) mRNA expression is high early in postnatal development but declines starting at postnatal day (PND) 10 and is virtually absent in the adult cortex. The mechanisms controlling this regulation are largely unknown. Methylation is important for gene silencing during development in many tissues, including the brain. In the present study, we examined the methylation status of ER alpha 5' untranslated exons during early postnatal development in male and female mice using methylation-specific PCR and pyrosequencing. Several regions of ER alpha promoter displayed a significant increase in methylation at PND 18 and 25 compared with PND 4. DNA methyltransferases (DNMT) are important for the initiation and maintenance of methylation. Real-time PCR showed that DNMT3A, the de novo DNMT peaked at PND 10 and was decreased by PND 25. DNMT1, which is important for maintenance of methylation, increased across development and stayed high in adult cortex. The methyl-CpG-binding protein 2 (MeCP2) is also important for stabilization of methylation. A chromatin immunoprecipitation assay showed a correlation between association of MeCP2 with ER alpha promoter and the increase in methylation and decrease in ER alpha expression after PND 10. In mice containing a mutant MeCP2 protein, ER alpha mRNA expression and promoter methylation patterns across development were different compared with wild-type mice. These data suggest that methylation of ER alpha promoters regulates ER alpha mRNA expression in the cortex during postnatal development in a MeCP2-dependent fashion.
MeSH Terms
- Aging
- Animals
- Cerebral Cortex
- DNA
- DNA (Cytosine-5-)-Methyltransferase 1
- DNA (Cytosine-5-)-Methyltransferases
- DNA Methylation
- DNA Primers
- Estrogen Receptor alpha
- Female
- Gene Expression Regulation, Developmental
- Male
- Methyl-CpG-Binding Protein 2
- Mice
- Mice, Inbred C57BL
- Polymerase Chain Reaction
- RNA, Messenger
- Sex Characteristics
With increasing age, T cells gain expression of killer immunoglobulin-like receptors (KIRs) that transmit negative signals and dampen the immune response. KIR expression is induced in CD4 and CD8 T cells by CpG DNA demethylation suggesting epigenetic control. To define the mechanisms that underlie the age-associated preferential KIR expression in CD8 T cells, we examined KIR2DL3 promoter methylation patterns. With age, CD8 T cells developed a patchy and stochastic promoter demethylation even in cells that did not express the KIR2DL3-encoded CD158b protein; complete demethylation of the minimal KIR2DL3 promoter was characteristic for CD158b-expressing cells. In contrast, the promoter in CD4 T cells was fully methylated irrespective of age. The selectivity for CD8 T cells correlated with lower DNMT1 recruitment to the KIR2DL3 promoter which further diminished with age. In contrast, binding of the polycomb protein EZH2 known to be involved in DNMT1 recruitment was not different. Our data suggest that CD8 T cells endure increasing displacement of DNMT1 from the KIR promoter with age, possibly because of an active histone signature. The ensuing partial demethylation lowers the threshold for transcriptional activation and renders CD8 T cells more susceptible to express KIR, thereby contributing to the immune defect in the elderly.
MeSH Terms
- Adult
- Aged
- Aged, 80 and over
- Aging
- CD4-Positive T-Lymphocytes
- CD8-Positive T-Lymphocytes
- CpG Islands
- DNA (Cytosine-5-)-Methyltransferase 1
- DNA (Cytosine-5-)-Methyltransferases
- DNA Methylation
- DNA-Binding Proteins
- Enhancer of Zeste Homolog 2 Protein
- Epigenesis, Genetic
- Female
- Humans
- Male
- Polycomb Repressive Complex 2
- Promoter Regions, Genetic
- Receptors, KIR2DL3
- Transcription Factors
Epigenetic alterations of DNA play key roles in determining gene structure and expression. Methylation of the 5-position of cytosine is thought to be the most common modification of the genome in mammals. Studies have generally shown that hypermethylation in gene regulatory regions is associated with inactivation and reduced transcription and that alteration in established methylation patterns during development can affect embryonic viability. Changes in methylation have also been associated with aging and cellular senescence as well as tumorogenesis. DNA methyltransferase 1 (DNMT1) is thought to play an important role in maintaining already established methylation patterns during DNA replication and catalyzes the transfer of a methyl moiety from S-adenosyl-L-methionine (SAM) to the 5-position of cytosines in the CpG dinucleotide. Several studies illustrate changes in activity and transcription of DNMT1 during aging and here we show a comprehensive method of detection of DNMT1 mRNA transcription from senescing cells in culture.
MeSH Terms
- Aging
- Animals
- Cell Culture Techniques
- Cell Transformation, Neoplastic
- Cells, Cultured
- Cellular Senescence
- DNA (Cytosine-5-)-Methyltransferase 1
- DNA (Cytosine-5-)-Methyltransferases
- DNA Methylation
- Epigenesis, Genetic
- Gene Expression Regulation
- Humans
- RNA, Messenger
- Transcription, Genetic
- Xenopus