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https://www.readbyqxmd.com/read/29224170/affinity-based-enrichment-techniques-for-the-genome-wide-analysis-of-5-hydroxymethylcytosine
#1
John P Thomson, Richard R Meehan
Since its initial characterization in 2009 there has been a great degree of interest in comparative profiling of 5-hydroxymethylcytosine (5hmC) nucleotides in vertebrate DNA. Through a host of genome-wide studies the distribution of 5hmC has been mapped in a range of cell lines, tissue types and organisms; the majority of which have been generated through affinity-based methods for 5hmC enrichment. Although recent advances in the field have resulted in the ability to investigate the levels of both methylated and hydroxymethylated cytosines at single base resolution, such studies are still relatively cost-prohibitive as well as technically challenging...
2018: Methods in Molecular Biology
https://www.readbyqxmd.com/read/29224169/multiplexing-for-oxidative-bisulfite-sequencing-oxbs-seq
#2
Kristina Kirschner, Felix Krueger, Anthony R Green, Tamir Chandra
DNA modifications, especially methylation, are known to play a crucial part in many regulatory processes in the cell. Recently, 5-hydroxymethylcytosine (5hmC) was discovered, a DNA modification derived as an intermediate of 5-methylcytosine (5mC) oxidation. Efforts to gain insights into function of this DNA modification are underway and several methods were recently described to assess 5hmC levels using sequencing approaches. Here we integrate adaptation based multiplexing and high-efficiency library prep into the oxidative Bisulfite Sequencing (oxBS-seq) workflow reducing the starting amount and cost per sample to identify 5hmC levels genome-wide...
2018: Methods in Molecular Biology
https://www.readbyqxmd.com/read/29224168/tet-assisted-bisulfite-sequencing-tab-seq
#3
Miao Yu, Dali Han, Gary C Hon, Chuan He
5-Hydroxymethylcytosine (5hmC) is a modified form of cytosine, which has recently been found in mammalian cells and tissues. 5hmC is derived from 5-methylcytosine (5mC) by Ten-eleven translocation (TET) family protein-mediated oxidation and may regulate gene expression. Numerous affinity-based profiling methods have been developed to help understand the exact function of 5hmC in the genome. However, these methods have a relatively low resolution (~100 bp) without quantitative information of the modification percentage on each site...
2018: Methods in Molecular Biology
https://www.readbyqxmd.com/read/29224139/antibody-based-detection-of-global-nuclear-dna-methylation-in-cells-tissue-sections-and-mammalian-embryos
#4
Nathalie Beaujean, Juliette Salvaing, Nur Annies Abd Hadi, Sari Pennings
Immunostaining is widely used in cell biology for the in situ detection of proteins in fixed cells. The method is based on the specificity of antibodies for recognizing and binding to a selected target, combined with immunolabeling techniques for microscopic imaging. Antibodies with high specificities for modified nucleotides have also been widely developed, and among those, antibodies that recognize modified cytosine: 5-methylcytosine (5mC), and more recently, its derivates 5-hydroxymethylcytosine (5hmC), 5-formylcytosine (5fC) and 5-carboxylcytosine (5caC)...
2018: Methods in Molecular Biology
https://www.readbyqxmd.com/read/29224138/quantification-of-global-dna-methylation-levels-by-mass-spectrometry
#5
Agustin F Fernandez, Luis Valledor, Fernando Vallejo, Maria Jesús Cañal, Mario F Fraga
Global DNA methylation was classically considered the relative percentage of 5-methylcysine (5mC) with respect to total cytosine (C). Early approaches were based on the use of high-performance separation technologies and UV detection. However, the recent development of protocols using mass spectrometry for the detection has increased sensibility and permitted the precise identification of peak compounds based on their molecular masses. This allows work to be conducted with much less genomic DNA starting material and also to quantify 5-hydroxymethyl-cytosine (5hmC), a recently identified form of methylated cytosine that could play an important role in active DNA demethylation...
2018: Methods in Molecular Biology
https://www.readbyqxmd.com/read/29223816/cortex-and-hippocampus-dna-epigenetic-response-to-a-long-term-arsenic-exposure-via-drinking-water
#6
Xiaoyan Du, Meiping Tian, Xiaoxue Wang, Jie Zhang, Qingyu Huang, Liangpo Liu, Heqing Shen
The neurotoxicity of arsenic is a serious health problem, especially for children. DNA epigenetic change may be an important pathogenic mechanism, but the molecular pathway remains obscure. In this study, the weaned male Sprague-Dawly (SD) rats were treated with arsenic trioxide via drinking water for 6 months, simulating real developmental exposure situation of children. Arsenic exposure impaired the cognitive abilities, and altered the expression of neuronal activity-regulated genes. Total arsenic concentrations of cortex and hippocampus tissues were significantly increased in a dose-dependent manner...
December 7, 2017: Environmental Pollution
https://www.readbyqxmd.com/read/29220513/integrating-5hmc-and-gene-expression-data-to-infer-regulatory-mechanisms
#7
Cristina Mitrea, Priyanga Wijesinghe, Greg Dyson, Adéle Kruger, Douglas M Ruden, Sorin Draghici, Aliccia Bollig-Fischer
Motivation: Epigenetic mechanisms are known to play a major role in breast cancer. However, the role of 5-hydroxymethylcytosine (5hmC) remains understudied. We hypothesize that 5hmC mediates redox regulation of gene expression in an aggressive subtype known as triple negative breast cancer (TNBC). To address this, our objective was to highlight genes that may be the target of this process by identifying redox-regulated, antioxidant-sensitive, gene-localized 5hmC changes associated with mRNA changes in TNBC cells...
December 6, 2017: Bioinformatics
https://www.readbyqxmd.com/read/29203910/tet-proteins-safeguard-bivalent-promoters-from-de-novo-methylation-in-human-embryonic-stem-cells
#8
Nipun Verma, Heng Pan, Louis C Doré, Abhijit Shukla, Qing V Li, Bobbie Pelham-Webb, Virginia Teijeiro, Federico González, Andrei Krivtsov, Chan-Jung Chang, Eirini P Papapetrou, Chuan He, Olivier Elemento, Danwei Huangfu
TET enzymes oxidize 5-methylcytosine (5mC) to 5-hydroxymethylcytosine (5hmC), which can lead to DNA demethylation. However, direct connections between TET-mediated DNA demethylation and transcriptional output are difficult to establish owing to challenges in distinguishing global versus locus-specific effects. Here we show that TET1, TET2 and TET3 triple-knockout (TKO) human embryonic stem cells (hESCs) exhibit prominent bivalent promoter hypermethylation without an overall corresponding decrease in gene expression in the undifferentiated state...
December 4, 2017: Nature Genetics
https://www.readbyqxmd.com/read/29186571/oxygen-gradients-can-determine-epigenetic-asymmetry-and-cellular-differentiation-via-differential-regulation-of-tet-activity-in-embryonic-stem-cells
#9
Simon Burr, Anna Caldwell, Mei Chong, Matteo Beretta, Stephen Metcalf, Matthew Hancock, Matthew Arno, Sucharitha Balu, Valeria Leon Kropf, Rajesh K Mistry, Ajay M Shah, Giovanni E Mann, Alison C Brewer
Graded levels of molecular oxygen (O2) exist within developing mammalian embryos and can differentially regulate cellular specification pathways. During differentiation, cells acquire distinct epigenetic landscapes, which determine their function, however the mechanisms which regulate this are poorly understood. The demethylation of 5-methylcytosine (5mC) is achieved via successive oxidation reactions catalysed by the Ten-Eleven-Translocation (Tet) enzymes, yielding the 5-hydroxymethylcytosine (5hmC) intermediate...
November 24, 2017: Nucleic Acids Research
https://www.readbyqxmd.com/read/29179435/genome-wide-5-hydroxymethylcytosine-patterns-in-human-spermatogenesis-are-associated-with-semen-quality
#10
Olga A Efimova, Anna A Pendina, Andrei V Tikhonov, Sergey E Parfenyev, Irina D Mekina, Evgeniia M Komarova, Mariia A Mazilina, Eugene V Daev, Olga G Chiryaeva, Ilona A Galembo, Mikhail I Krapivin, Oleg S Glotov, Irina S Stepanova, Svetlana A Shlykova, Igor Yu Kogan, Alexander M Gzgzyan, Tatyana V Kuznetzova, Vladislav S Baranov
We performed immunofluorescent analysis of DNA hydroxymethylation and methylation in human testicular spermatogenic cells from azoospermic patients and ejaculated spermatozoa from sperm donors and patients from infertile couples. In contrast to methylation which was present throughout spermatogenesis, hydroxymethylation was either high or almost undetectable in both spermatogenic cells and ejaculated spermatozoa. On testicular cytogenetic preparations, 5-hydroxymethylcytosine was undetectable in mitotic and meiotic chromosomes, and was present exclusively in interphase spermatogonia Ad and in a minor spermatid population...
October 24, 2017: Oncotarget
https://www.readbyqxmd.com/read/29175454/selective-inhibition-of-ctcf-binding-by-ias-directs-tet-mediated-reprogramming-of-5-hydroxymethylation-patterns-in-ias-transformed-cells
#11
Matthew Rea, Tyler Gripshover, Yvonne Fondufe-Mittendorf
Methylation at cytosine (5mC) is a fundamental epigenetic DNA modification recently associated with iAs-mediated carcinogenesis. In contrast, the role of 5-hydroxymethylcytosine (5hmC), the oxidation product of 5mC in iAs-mediated carcinogenesis is unknown. Here we assess the hydroxymethylome in iAs-transformed cells, showing that dynamic modulation of hydroxymethylated DNA is associated with specific transcriptional networks. Moreover, this pathologic iAs-mediated carcinogenesis is characterized by a shift toward a higher hydroxymethylation pattern genome-wide...
November 21, 2017: Toxicology and Applied Pharmacology
https://www.readbyqxmd.com/read/29158345/stable-oxidative-cytosine-modifications-accumulate-in-cardiac-mesenchymal-cells-from-type2-diabetes-patients-rescue-by-alpha-ketoglutarate-and-tet-tdg-functional-reactivation
#12
Francesco Spallotta, Chiara Cencioni, Sandra Atlante, Davide Garella, Mattia Cocco, Mattia Mori, Raffaella Mastrocola, Carsten Künne, Stefan Günther, Simona Nanni, Valerio Azzimato, Sven Zukunft, Angela Kornberger, Duran Sueruen, Frank Schnutgen, Harald von Melchner, Antonella Di Stilo, Manuela Aragno, Maarten Braspenning, Wim Van Criekinge, Miles J De Blasio, Rebecca H Ritchie, Germana Zaccagnini, Fabio Martelli, Antonella Farsetti, Ingrid Fleming, Thomas Braun, Andres Beiras-Fernandez, Bruno Botta, Massimo Collino, Massimo Bertinaria, Andreas M Zeiher, Carlo Gaetano
Rationale: Human cardiac mesenchymal cells (CMSCs) are a therapeutically-relevant primary cell population. Diabetes compromises CMSC function as consequence of metabolic alterations and incorporation of stable epigenetic changes. Objective: To investigate the role of α-ketoglutarate (αKG) in the epi-metabolic control of DNA demethylation in CMSCs. Methods and Results: Quantitative global analysis, methylated and hydroxymethylated DNA sequencing and gene specific GC methylation detection revealed an accumulation of 5mC, 5hmC and 5fC in the genomic DNA of human CMSCs isolated from diabetic (D) donors (D-CMSCs)...
November 20, 2017: Circulation Research
https://www.readbyqxmd.com/read/29156803/tet1-inhibits-cell-proliferation-by-inducing-rassf5-expression
#13
Bo-Tai Li, Chao Yu, Ying Xu, Sheng-Bing Liu, Heng-Yu Fan, Wei-Wei Pan
Tet methylcytosine dioxygenases (TETs) catalyze the oxidative reactions of 5-methylcytosine to 5-hydroxymethylcytosine (5hmC). However, TET1 roles in ovarian cancer cell growth are unknown. Here, we show that ectopic expression of TET1 increased 5hmC levels, and inhibited proliferation and colony formation in ovarian cancer cell lines. Furthermore, in vitro and in vivo functional studies demonstrated that TET1 overexpression is necessary for the suppression of ovarian cancer growth, whereas depletion of TET1 expression had the opposite effect...
October 17, 2017: Oncotarget
https://www.readbyqxmd.com/read/29156615/clinical-significance-of-measuring-global-hydroxymethylation-of-white-blood-cell-dna-in-prostate-cancer-comparison-to-psa-in-a-pilot-exploratory-study
#14
Alin Grelus, Dragos V Nica, Imola Miklos, Valerica Belengeanu, Ioan Ioiart, Cristina Popescu
This is the first study investigating the clinical relevance of 5-hydroxymethylcytosine (5hmC) in genomic DNA from white blood cells (WBC) in the context of prostate cancer (PCa) and other prostate pathologies. Using an enzyme-linked immunosorbent assay, we identified significantly different distributions of patients with low and elevated 5hmC content in WBC DNA across controls and patients with prostate cancer (PCa), atypical small acinar proliferation (ASAP), and benign prostatic hyperplasia (BPH). The measured values were within the normal range for most PCa patients, while the latter category was predominant for ASAP...
November 20, 2017: International Journal of Molecular Sciences
https://www.readbyqxmd.com/read/29111854/the-effect-of-morphine-upon-dna-methylation-in-ten-regions-of-the-rat-brain
#15
Timothy M Barrow, Hyang-Min Byun, Xinyan Li, Chris Smart, Yong-Xiang Wang, Yacong Zhang, Andrea A Baccarelli, Liqiong Guo
Morphine is one of the most effective analgesics in medicine. However, its use is associated with the development of tolerance and dependence. Recent studies demonstrating epigenetic changes in the brain after exposure to opiates have provided insight into mechanisms possibly underlying addiction. In this study, we sought to identify epigenetic changes in ten regions of the rat brain following acute and chronic morphine exposure. We analyzed DNA methylation of six nuclear-encoded genes implicated in brain function (Bdnf, Comt, Il1b, Il6, Nr3c1, and Tnf) and three mitochondrially-encoded genes (Mtco1, Mtco2, and Mtco3), and measured global 5-methylcytosine (5mC) and 5-hydroxymethylcytosine (5hmC) levels...
November 7, 2017: Epigenetics: Official Journal of the DNA Methylation Society
https://www.readbyqxmd.com/read/29092933/integrating-dna-methylation-and-hydroxymethylation-data-with-the-mint-pipeline
#16
Raymond G Cavalcante, Snehal Patil, Yongseok Park, Laura S Rozek, Maureen A Sartor
DNA methylation (5mC) plays important roles in mammalian development, oncogenesis, treatment response, and responses to the environment. DNA hydroxymethylation (5hmC) is also an informative epigenetic mark with distinct roles in regulation and cancer. Gold-standard, widely used technologies (bisulfite conversion, followed by deep sequencing) cannot distinguish between 5mC and 5hmC. Therefore, additional experiments are required to differentiate the two marks, and in silico methods are needed to analyze, integrate, and interpret these data...
November 1, 2017: Cancer Research
https://www.readbyqxmd.com/read/29072898/the-epstein-barr-virus-b-zip-protein-zta-recognizes-specific-dna-sequences-containing-5-methylcytosine-and-5-hydroxymethylcytosine
#17
Desiree Tillo, Sreejana Ray, Khund-Sayeed Syed, Mary Rose Gaylor, Ximiao He, Jun Wang, Nima Assad, Stewart R Durell, Aleksey Porollo, Matthew T Weirauch, Charles Vinson
The Epstein-Barr virus (EBV) B-ZIP transcription factor Zta binds to many DNA sequences containing methylated CG dinucleotides. Using protein binding microarrays (PBMs), we analyzed the sequence specific DNA binding of Zta to four kinds of double-stranded DNA (dsDNA): (1) DNA containing cytosine in both strands, (2) DNA with 5-methylcytosine (5mC) in one strand and cytosine in the second strand, (3) DNA with 5-hydroxymethylcytosine (5hmC) in one strand and cytosine in the second strand, and (4) DNA in which both cytosines in all CG dinucleotides contain 5mC...
November 10, 2017: Biochemistry
https://www.readbyqxmd.com/read/29069286/dna-hydroxymethylation-levels-are-altered-in-blood-cells-from-down-syndrome-persons-enrolled-in-the-mark-age-project
#18
Fabio Ciccarone, Elisabetta Valentini, Marco Malavolta, Michele Zampieri, Maria Giulia Bacalini, Roberta Calabrese, Tiziana Guastafierro, Anna Reale, Claudio Franceschi, Miriam Capri, Nicolle Breusing, Tilman Grune, María Moreno-Villanueva, Alexander Bürkle, Paola Caiafa
Down syndrome (DS) is caused by the presence of part or an entire extra copy of chromosome 21, a phenomenon that can cause a wide spectrum of clinically defined phenotypes of the disease. Most of the clinical signs of DS are typical of the ageing process including dysregulation of immune system. Beyond the causative genetic defect, DS persons display epigenetic alterations, particularly aberrant DNA methylation patterns that can contribute to the heterogeneity of the disease. In the present work we investigated the levels of 5-hydroxymethylcytosine (5hmC) and of the TET dioxygenase enzymes, which are involved in DNA demethylation processes and are often deregulated in pathological conditions as well as in ageing...
October 21, 2017: Journals of Gerontology. Series A, Biological Sciences and Medical Sciences
https://www.readbyqxmd.com/read/29036642/striking-a-balance-regulation-of-transposable-elements-by-zfp281-and-mll2-in-mouse-embryonic-stem-cells
#19
Qian Dai, Yang Shen, Yan Wang, Xin Wang, Joel Celio Francisco, Zhuojuan Luo, Chengqi Lin
Transposable elements (TEs) compose about 40% of the murine genome. Retrotransposition of active TEs such as LINE-1 (L1) tremendously impacts genetic diversification and genome stability. Therefore, transcription and transposition activities of retrotransposons are tightly controlled. Here, we show that the Krüppel-like zinc finger protein Zfp281 directly binds and suppresses a subset of retrotransposons, including the active young L1 repeat elements, in mouse embryonic stem (ES) cells. In addition, we find that Zfp281-regulated L1s are highly enriched for 5-hydroxymethylcytosine (5hmC) and H3K4me3...
September 25, 2017: Nucleic Acids Research
https://www.readbyqxmd.com/read/29028987/in-silico-structural-modelling-of-multiple-epigenetic-marks-on-dna
#20
Konrad Krawczyk, Samuel Demharter, Bernhard Knapp, Charlotte M Deane, Peter Minary
Summary: There are four known epigenetic cytosine modifications in mammals: 5mC, 5hmC, 5fC and 5caC. The biological effects of 5mC are well understood but the roles of the remaining modifications remain elusive. Experimental and computational studies suggest that a single epigenetic mark has little structural effect but six of them can radically change the structure of DNA to a new form, F-DNA. Investigating the collective effect of multiple epigenetic marks requires the ability to interrogate all possible combinations of epigenetic states (e...
September 25, 2017: Bioinformatics
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