Detection of DNA methyltransferase activity using allosteric molecular beacons

The Analyst ◽  
2016 ◽  
Vol 141 (2) ◽  
pp. 579-584 ◽  
Author(s):  
Weiting Zhang ◽  
Xiaolong Zu ◽  
Yanling Song ◽  
Zhi Zhu ◽  
Chaoyong James Yang
Keyword(s):  
Dna Methylation ◽  
Dna Methyltransferase ◽  
Molecular Beacon ◽  
Molecular Beacons ◽  
Sensitive Detection ◽  
Methyltransferase Activity ◽  
Mtase Activity ◽  
Adenine Methylation ◽  
Methylation Patterns

Abnormal DNA methylation patterns caused by altered DNA methyltransferase (MTase) activity are closely associated with cancer. Herein, using DNA adenine methylation methyltransferase (Dam MTase) as a model analyte, we designed an allosteric molecular beacon (aMB) for sensitive detection of Dam MTase activity.

A glucometer-based strategy for sensitive DNA methyltransferase activity detection via a polymerization nicking reaction and enzyme amplification

2017 ◽  
Vol 9 (20) ◽  
pp. 2933-2938 ◽  
Author(s):  
Ying Chen ◽  
Hongchao Yi
Keyword(s):  
Dna Methyltransferase ◽  
Sensitive Detection ◽  
Activity Detection ◽  
Methyltransferase Activity ◽  
Mtase Activity ◽  
Enzyme Amplification ◽  
Signal Output

The polymerization nicking reaction (PNR) and invertase enzyme amplify the glucometer signal output to achieve simple and sensitive detection of DNA methyltransferase (MTase) activity.

Abstract 40: Disturbed Flow Alters Genomewide DNA Methylation Patterns, Regulating Endothelial Gene Expression and Atherosclerosis

2014 ◽  
Vol 34 (suppl_1) ◽  
Author(s):  
Jessilyn Dunn ◽  
Haiwei Qiu ◽  
Soyeon Kim ◽  
Daudi Jjingo ◽  
Ryan Hoffman ◽  
...  
Keyword(s):  
Gene Expression ◽  
Dna Methylation ◽  
Dna Methyltransferase ◽  
Methylation Status ◽  
Western Diet ◽  
Dependent Manner ◽  
Gene Promoters ◽  
Genome Wide ◽  
Methylation Patterns ◽  
Endothelial Gene Expression

Atherosclerosis preferentially occurs in arterial regions of disturbed blood flow (d-flow), which alters gene expression, endothelial function, and atherosclerosis. Here, we show that d-flow regulates genome-wide DNA methylation patterns in a DNA methyltransferase (DNMT)-dependent manner. We found that d-flow induced expression of DNMT1, but not DNMT3a or DNMT3b, in mouse arterial endothelium in vivo and in cultured endothelial cells by oscillatory shear (OS) compared to unidirectional laminar shear in vitro. The DNMT inhibitor 5-Aza-2’deoxycytidine (5Aza) or DNMT1 siRNA significantly reduced OS-induced endothelial inflammation. Moreover, 5Aza reduced lesion formation in two atherosclerosis models using ApoE-/- mice (western diet for 3 months and the partial carotid ligation model with western diet for 3 weeks). To identify the 5Aza mechanisms, we conducted two genome-wide studies: reduced representation bisulfite sequencing (RRBS) and transcript microarray using endothelial-enriched gDNA and RNA, respectively, obtained from the partially-ligated left common carotid artery (LCA exposed to d-flow) and the right contralateral control (RCA exposed to s-flow) of mice treated with 5Aza or vehicle. D-flow induced DNA hypermethylation in 421 gene promoters, which was significantly prevented by 5Aza in 335 genes. Systems biological analyses using the RRBS and the transcriptome data revealed 11 mechanosensitive genes whose promoters were hypermethylated by d-flow but rescued by 5Aza treatment. Of those, five genes contain hypermethylated cAMP-response-elements in their promoters, including the transcription factors HoxA5 and Klf3. Their methylation status could serve as a mechanosensitive master switch in endothelial gene expression. Our results demonstrate that d-flow controls epigenomic DNA methylation patterns in a DNMT-dependent manner, which in turn alters endothelial gene expression and induces atherosclerosis.

scholarly journals Fetal and Neonatal Exposure to the Endocrine Disruptor Methoxychlor Causes Epigenetic Alterations in Adult Ovarian Genes

Endocrinology ◽  
2009 ◽  
Vol 150 (10) ◽  
pp. 4681-4691 ◽  
Author(s):  
Aparna Mahakali Zama ◽  
Mehmet Uzumcu
Keyword(s):  
Dna Methylation ◽  
Dna Methyltransferase ◽  
Ovarian Function ◽  
Endocrine Disrupting Chemicals ◽  
Neonatal Exposure ◽  
Dna Hypermethylation ◽  
Promoter Regions ◽  
Altered Expression ◽  
Arbitrarily Primed Pcr ◽  
Methylation Patterns

Abstract Exposure to endocrine-disrupting chemicals during development could alter the epigenetic programming of the genome and result in adult-onset disease. Methoxychlor (MXC) and its metabolites possess estrogenic, antiestrogenic, and antiandrogenic activities. Previous studies showed that fetal/neonatal exposure to MXC caused adult ovarian dysfunction due to altered expression of key ovarian genes including estrogen receptor (ER)-β, which was down-regulated, whereas ERα was unaffected. The objective of the current study was to evaluate changes in global and gene-specific methylation patterns in adult ovaries associated with the observed defects. Rats were exposed to MXC (20 μg/kg·d or 100 mg/kg·d) between embryonic d 19 and postnatal d 7. We performed DNA methylation analysis of the known promoters of ERα and ERβ genes in postnatal d 50–60 ovaries using bisulfite sequencing and methylation-specific PCRs. Developmental exposure to MXC led to significant hypermethylation in the ERβ promoter regions (P < 0.05), whereas the ERα promoter was unaffected. We assessed global DNA methylation changes using methylation-sensitive arbitrarily primed PCR and identified 10 genes that were hypermethylated in ovaries from exposed rats. To determine whether the MXC-induced methylation changes were associated with increased DNA methyltransferase (DNMT) levels, we measured the expression levels of Dnmt3a, Dnmt3b, and Dnmt3l using semiquantitative RT-PCR. Whereas Dnmt3a and Dnmt3l were unchanged, Dnmt3b expression was stimulated in ovaries of the 100 mg/kg MXC group (P < 0.05), suggesting that increased DNMT3B may cause DNA hypermethylation in the ovary. Overall, these data suggest that transient exposure to MXC during fetal and neonatal development affects adult ovarian function via altered methylation patterns.

scholarly journals Structural insights into CpG-specific DNA methylation by human DNA methyltransferase 3B

2020 ◽  
Vol 48 (7) ◽  
pp. 3949-3961 ◽  
Author(s):  
Chien-Chu Lin ◽  
Yi-Ping Chen ◽  
Wei-Zen Yang ◽  
James C K Shen ◽  
Hanna S Yuan
Keyword(s):  
Dna Methylation ◽  
Dna Methyltransferase ◽  
Cytosine Methylation ◽  
Catalytic Domain ◽  
De Novo ◽  
Water Channel ◽  
Dna Methyltransferases ◽  
Structural Basis ◽  
Cpg Sites ◽  
Methylation Patterns

Abstract DNA methyltransferases are primary enzymes for cytosine methylation at CpG sites of epigenetic gene regulation in mammals. De novo methyltransferases DNMT3A and DNMT3B create DNA methylation patterns during development, but how they differentially implement genomic DNA methylation patterns is poorly understood. Here, we report crystal structures of the catalytic domain of human DNMT3B–3L complex, noncovalently bound with and without DNA of different sequences. Human DNMT3B uses two flexible loops to enclose DNA and employs its catalytic loop to flip out the cytosine base. As opposed to DNMT3A, DNMT3B specifically recognizes DNA with CpGpG sites via residues Asn779 and Lys777 in its more stable and well-ordered target recognition domain loop to facilitate processive methylation of tandemly repeated CpG sites. We also identify a proton wire water channel for the final deprotonation step, revealing the complete working mechanism for cytosine methylation by DNMT3B and providing the structural basis for DNMT3B mutation-induced hypomethylation in immunodeficiency, centromere instability and facial anomalies syndrome.

scholarly journals Persistent epigenetic memory impedes rescue of the telomeric phenotype in human ICF iPSCs following DNMT3B correction

eLife ◽  
2019 ◽  
Vol 8 ◽  
Author(s):  
Shir Toubiana ◽  
Miriam Gagliardi ◽  
Mariarosaria Papa ◽  
Roberta Manco ◽  
Maty Tzukerman ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Dna Methyltransferase ◽  
De Novo ◽  
Pharmacological Intervention ◽  
Icf Syndrome ◽  
Genome Wide ◽  
Mammalian Genomes ◽  
Induced Pluripotent ◽  
Methylation Patterns

DNA methyltransferase 3B (DNMT3B) is the major DNMT that methylates mammalian genomes during early development. Mutations in human DNMT3B disrupt genome-wide DNA methylation patterns and result in ICF syndrome type 1 (ICF1). To study whether normal DNA methylation patterns may be restored in ICF1 cells, we corrected DNMT3B mutations in induced pluripotent stem cells from ICF1 patients. Focusing on repetitive regions, we show that in contrast to pericentromeric repeats, which reacquire normal methylation, the majority of subtelomeres acquire only partial DNA methylation and, accordingly, the ICF1 telomeric phenotype persists. Subtelomeres resistant to de novo methylation were characterized by abnormally high H3K4 trimethylation (H3K4me3), and short-term reduction of H3K4me3 by pharmacological intervention partially restored subtelomeric DNA methylation. These findings demonstrate that the abnormal epigenetic landscape established in ICF1 cells restricts the recruitment of DNMT3B, and suggest that rescue of epigenetic diseases with genome-wide disruptions will demand further manipulation beyond mutation correction.

Sensitive detection of DNA methyltransferase activity by transcription-mediated duplex-specific nuclease-assisted cyclic signal amplification

2015 ◽  
Vol 51 (73) ◽  
pp. 13968-13971 ◽  
Author(s):  
Yan Zhang ◽  
Wen-jing Xu ◽  
Ya-ping Zeng ◽  
Chun-yang Zhang
Keyword(s):  
Signal Amplification ◽  
Dna Methyltransferase ◽  
Sensitive Detection ◽  
Methyltransferase Activity

We develop transcription-mediated duplex-specific nuclease-assisted cyclic signal amplification for sensitive detection of DNA methyltransferase activity.

scholarly journals Single-ribonucleotide repair-mediated ligation-dependent cycling signal amplification for sensitive and specific detection of DNA methyltransferase

2018 ◽  
Vol 9 (28) ◽  
pp. 6053-6061 ◽  
Author(s):  
Li-juan Wang ◽  
Xiao Han ◽  
Chen-chen Li ◽  
Chun-yang Zhang
Keyword(s):  
Signal Amplification ◽  
Dna Methyltransferase ◽  
Sensitive Detection ◽  
Specific Detection ◽  
Mtase Activity

Specific and sensitive detection of DNA MTase activity can be achieved by a single-ribonucleotide repair-mediated ligation-dependent cycling signal amplification approach.

scholarly journals DNA Methylation Changes in Human Papillomavirus-Driven Head and Neck Cancers

Cells ◽  
2020 ◽  
Vol 9 (6) ◽  
pp. 1359 ◽  
Author(s):  
Chameera Ekanayake Weeramange ◽  
Kai Dun Tang ◽  
Sarju Vasani ◽  
Julian Langton-Lockton ◽  
Liz Kenny ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Human Papillomavirus ◽  
Head And Neck ◽  
Dna Methyltransferase ◽  
Long Control Region ◽  
Cancer Types ◽  
E6 And E7 ◽  
Hpv Oncoproteins ◽  
Cellular Dna ◽  
Methylation Patterns

Disruption of DNA methylation patterns is one of the hallmarks of cancer. Similar to other cancer types, human papillomavirus (HPV)-driven head and neck cancer (HNC) also reveals alterations in its methylation profile. The intrinsic ability of HPV oncoproteins E6 and E7 to interfere with DNA methyltransferase activity contributes to these methylation changes. There are many genes that have been reported to be differentially methylated in HPV-driven HNC. Some of these genes are involved in major cellular pathways, indicating that DNA methylation, at least in certain instances, may contribute to the development and progression of HPV-driven HNC. Furthermore, the HPV genome itself becomes a target of the cellular DNA methylation machinery. Some of these methylation changes appearing in the viral long control region (LCR) may contribute to uncontrolled oncoprotein expression, leading to carcinogenesis. Consistent with these observations, demethylation therapy appears to have significant effects on HPV-driven HNC. This review article comprehensively summarizes DNA methylation changes and their diagnostic and therapeutic indications in HPV-driven HNC.

scholarly journals Sensitive detection of methylated DNA and methyltransferase activity based on the lighting up of FAM-labeled DNA quenched fluorescence by gold nanoparticles

RSC Advances ◽  
2019 ◽  
Vol 9 (21) ◽  
pp. 12063-12069 ◽  
Author(s):  
Mohammad Ali Karimi ◽  
Mehdi Dadmehr ◽  
Morteza Hosseini ◽  
Behnaz Korouzhdehi ◽  
Fatemeh Oroojalian
Keyword(s):  
Dna Methylation ◽  
Gold Nanoparticles ◽  
Sensitive Detection ◽  
Fluorescence Recovery ◽  
Methyltransferase Activity ◽  
Au Nps ◽  
Methylated Dna ◽  
Novel Method

A novel method for detection of DNA methylation based on fluorescence recovery of FAM labeled DNA/Au NPs was introduced.

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