scholarly journals Changes of DNA methylation and hydroxymethylation in plant protoplast cultures.

2013 ◽  
Vol 60 (1) ◽  
Author(s):  
Pavla Moricová ◽  
Vladan Ondřej ◽  
Božena Navrátilová ◽  
Lenka Luhová

Cytosine methylation patterns in higher eukaryotes are important in gene regulation. Along with 5-methylcytosine (5-mC), a newly discovered constituent of mammalian DNA, 5-hydroxymethylcytosine (5-hmC), is the other modified base in higher organisms. In this study we detected 5-hmC in plant protoplast DNA and demonstrated its increasing content during the first 72 hrs. of protoplast cultivation. In contrast to 5-hmC, the amount of 5-mC decreased during protoplast cultivation. It was also found that 5-hmC did not primarily arise as a product of oxidative DNA damage following protoplast culture.

1996 ◽  
Vol 318 (1) ◽  
pp. 21-23 ◽  
Author(s):  
Harparkash KAUR ◽  
Barry HALLIWELL

Oxidative DNA damage is thought to be an important contributor to cancer development and to be affected by dietary constituents, so its accurate measurement is important. DNA methylation is recognized as an important mechanism affecting gene expression. In the present paper we describe an HPLC-with-electrochemical-detection procedure that allows rapid and sensitive measurement of four oxidized (2,6-diamino-4-hydroxy-5-formamidopyrimidine, 5-hydroxyuracil, 8-hydroxyguanine, 8-hydroxyadenine) and three methylated (7-methylguanine, 1-methylguanine, O6-methylguanine) bases in acid hydrolysates of DNA. Guanine was also detected, but was clearly separated from the other bases.


Author(s):  
S. Lewis ◽  
L. Ross ◽  
S.A. Bain ◽  
E. Pahita ◽  
S.A. Smith ◽  
...  

AbstractCytosine methylation is an ancient epigenetic modification yet its function and extent within genomes is highly variable across eukaryotes. In mammals, methylation controls transposable elements and regulates the promoters of genes. In insects, DNA methylation is generally restricted to a small subset of transcribed genes, with both intergenic regions and transposable elements (TEs) depleted of methylation. The evolutionary origin and the function of these methylation patterns are poorly understood. Here we characterise the evolution of DNA methylation across the arthropod phylum. While the common ancestor of the arthropods had low levels of TE methylation and did not methylate promoters, both of these functions have evolved independently in centipedes and mealybugs. In contrast, methylation of the exons of a subset of transcribed genes is ancestral and widely conserved across the phylum, but has been lost in specific lineages. Remarkably the same set of genes are likely to be methylated in all species that retained exon-enriched methylation. We show that these genes have characteristic patterns of expression correlating to broad transcription initiation sites and well-positioned nucleosomes, providing new insights into potential mechanisms driving methylation patterns over hundreds of millions of years.Author SummaryAnimals develop from a single cell to form a complex organism with many specialised cells. Almost all of the fantastic variety of cells must have the same sequence of DNA, and yet they have distinct identities that are preserved even when they divide. This remarkable process is achieved by turning different sets of genes on or off in different types of cell using molecular mechanisms known as “epigenetic gene regulation”.Surprisingly, though all animals need epigenetic gene regulation, there is a huge diversity in the mechanisms that they use. Characterising and explaining this diversity is crucial in understanding the functions of epigenetic pathways, many of which have key roles in human disease. We studied how one particular type of epigenetic regulation, known as DNA methylation, has evolved within arthropods. Arthropods are an extraordinarily diverse group of animals ranging from horseshoe crabs to fruit flies. We discovered that the levels of DNA methylation and where it is found within the genome changes rapidly throughout arthropod evolution. Nevertheless, there are some features of DNA methylation that seem to be the same across most arthropods-in particular we found that there is a tendency for a similar set of genes to acquire methylation of DNA in most arthropods, and that this is conserved over 350 million years. We discovered that these genes have distinct features that might explain how methylation gets targeted. Our work provides important new insights into the evolution of DNA methylation and gives some new hints to its essential functions.


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

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.


2019 ◽  
Author(s):  
Luis Busto-Moner ◽  
Julien Morival ◽  
Arjang Fahim ◽  
Zachary Reitz ◽  
Timothy L. Downing ◽  
...  

AbstractDNA methylation is a heritable epigenetic modification that plays an essential role in mammalian development. Genomic methylation patterns are dynamically maintained, with DNA methyltransferases mediating inheritance of methyl marks onto nascent DNA over cycles of replication. A recently developed experimental technique employing immunoprecipitation of bromodeoxyuridine labeled nascent DNA followed by bisulfite sequencing (Repli-BS) measures post-replication temporal evolution of cytosine methylation, thus enabling genome-wide monitoring of methylation maintenance. In this work, we combine statistical analysis and stochastic mathematical modeling to analyze Repli-BS data from human embryonic stem cells. We estimate site-specific kinetic rate constants for the restoration of methyl marks on >10 million uniquely mapped cytosines within the CpG (cytosine-phosphate-guanine) dinucleotide context across the genome using Maximum Likelihood Estimation. We find that post-replication remethylation rate constants span approximately two orders of magnitude, with half-lives of per-site recovery of steady-state methylation levels ranging from shorter than ten minutes to five hours and longer. Furthermore, we find that kinetic constants of maintenance methylation are correlated among neighboring CpG sites. Stochastic mathematical modeling provides insight to the biological mechanisms underlying the inference results, suggesting that enzyme processivity and/or collaboration can produce the observed kinetic correlations. Our combined statistical/mathematical modeling approach expands the utility of genomic datasets and disentangles heterogeneity in methylation patterns arising from replication-associated temporal dynamics versus stable cell-to-cell differences.


Cells ◽  
2020 ◽  
Vol 9 (1) ◽  
pp. 225 ◽  
Author(s):  
Zhongliang Jiang ◽  
Yanhao Lai ◽  
Jill M. Beaver ◽  
Pawlos S. Tsegay ◽  
Ming-Lang Zhao ◽  
...  

DNA damage and base excision repair (BER) are actively involved in the modulation of DNA methylation and demethylation. However, the underlying molecular mechanisms remain unclear. In this study, we seek to understand the mechanisms by exploring the effects of oxidative DNA damage on the DNA methylation pattern of the tumor suppressor breast cancer 1 (BRCA1) gene in the human embryonic kidney (HEK) HEK293H cells. We found that oxidative DNA damage simultaneously induced DNA demethylation and generation of new methylation sites at the CpGs located at the promoter and transcribed regions of the gene ranging from −189 to +27 in human cells. We demonstrated that DNA damage-induced demethylation was mediated by nucleotide misincorporation by DNA polymerase β (pol β). Surprisingly, we found that the generation of new DNA methylation sites was mediated by coordination between pol β and the de novo DNA methyltransferase, DNA methyltransferase 3b (DNMT3b), through the interaction between the two enzymes in the promoter and encoding regions of the BRCA1 gene. Our study provides the first evidence that oxidative DNA damage can cause dynamic changes in DNA methylation in the BRCA1 gene through the crosstalk between BER and de novo DNA methylation.


2006 ◽  
Vol 70 (3) ◽  
pp. 830-856 ◽  
Author(s):  
Josep Casadesús ◽  
David Low

SUMMARY Like many eukaryotes, bacteria make widespread use of postreplicative DNA methylation for the epigenetic control of DNA-protein interactions. Unlike eukaryotes, however, bacteria use DNA adenine methylation (rather than DNA cytosine methylation) as an epigenetic signal. DNA adenine methylation plays roles in the virulence of diverse pathogens of humans and livestock animals, including pathogenic Escherichia coli, Salmonella, Vibrio, Yersinia, Haemophilus, and Brucella. In Alphaproteobacteria, methylation of adenine at GANTC sites by the CcrM methylase regulates the cell cycle and couples gene transcription to DNA replication. In Gammaproteobacteria, adenine methylation at GATC sites by the Dam methylase provides signals for DNA replication, chromosome segregation, mismatch repair, packaging of bacteriophage genomes, transposase activity, and regulation of gene expression. Transcriptional repression by Dam methylation appears to be more common than transcriptional activation. Certain promoters are active only during the hemimethylation interval that follows DNA replication; repression is restored when the newly synthesized DNA strand is methylated. In the E. coli genome, however, methylation of specific GATC sites can be blocked by cognate DNA binding proteins. Blockage of GATC methylation beyond cell division permits transmission of DNA methylation patterns to daughter cells and can give rise to distinct epigenetic states, each propagated by a positive feedback loop. Switching between alternative DNA methylation patterns can split clonal bacterial populations into epigenetic lineages in a manner reminiscent of eukaryotic cell differentiation. Inheritance of self-propagating DNA methylation patterns governs phase variation in the E. coli pap operon, the agn43 gene, and other loci encoding virulence-related cell surface functions.


2013 ◽  
Vol 33 (6) ◽  
pp. 1139-1148 ◽  
Author(s):  
Hao Chen ◽  
Honghui Ma ◽  
Hiroyuki Inuzuka ◽  
Jianbo Diao ◽  
Fei Lan ◽  
...  

UHRF1 (ubiquitin-like, with PHD and RING finger domains 1) is a critical epigenetic player involved in the maintenance of DNA methylation patterns during DNA replication. Dysregulation of the UHRF1 level is implicated in cancer onset, metastasis, and tumor recurrence. Previous studies demonstrated that UHRF1 can be stabilized through USP7-mediated deubiquitylation, but the mechanism through which UHRF1 is ubiquitylated is still unknown. Here we show that proteasomal degradation of UHRF1 is mediated by the SCFβ-TrCPE3 ligase. Through bioinformatic and mutagenesis studies, we identified a functional DSG degron in the UHRF1 N terminus that is necessary for UHRF1 stability regulation. We further show that UHRF1 physically interacts with β-TrCP1 in a manner dependent on phosphorylation of serine 108 (S108UHRF1) within the DSG degron. Furthermore, we demonstrate that S108UHRF1phosphorylation is catalyzed by casein kinase 1 delta (CK1δ) and is important for the recognition of UHRF1 by SCFβ-TrCP. Importantly, we demonstrate that UHRF1 degradation is accelerated in response to DNA damage, coincident with enhanced S108UHRF1phosphorylation. Taken together, our data identify SCFβ-TrCPas a bona fide UHRF1 E3 ligase important for regulating UHRF1 steady-state levels both under normal conditions and in response to DNA damage.


2018 ◽  
Author(s):  
Komivi Dossa ◽  
Marie Ali Mmadi ◽  
Rong Zhou ◽  
Qi Zhou ◽  
Mei Yang ◽  
...  

AbstractDNA methylation is a heritable epigenetic mechanism that participates in gene regulation under abiotic stresses in plants. Sesame (Sesamum indicum L.) is typically considered a drought-tolerant crop but highly susceptible to waterlogging, a property attributed to its presumed origin in Africa or India. Understanding DNA methylation patterns in sesame under drought and waterlogging conditions can provide insights into the regulatory mechanisms underlying its contrasting responses to these principal abiotic stresses. Here, we combined Methylation-Sensitive Amplified Polymorphism and transcriptome analyses to profile cytosine methylation patterns, gene expression alteration, and their interplay in drought-tolerant and waterlogging-tolerant sesame genotypes under control, stress and recovery conditions. Our data showed that drought stress strongly induced de novo methylation (DNM) whereas most of the loci were demethylated (DM) during the recovery phase. In contrast, waterlogging decreased the level of methylation under stress but during the recovery phase, both DM and DNM were concomitantly deployed. In both stresses, the differentially expressed genes (DEGs) were highly correlated with the methylation patterns. We observed that DM was associated with the up-regulation of the DEGs while DNM was correlated with the down-regulation of the DEGs. In addition, we sequenced 44 differentially methylated regions of which 90% overlapped with the promoters and coding sequences of the DEGs. Altogether, we demonstrated that sesame has divergent epigenetic programs that respond to drought and waterlogging stresses. Our results also highlighted the possible interplay among DNA methylation and gene expression, which may modulate the contrasting responses to drought and waterlogging in sesame.


2020 ◽  
Vol 48 (8) ◽  
pp. 4081-4099 ◽  
Author(s):  
Alex I Finnegan ◽  
Somang Kim ◽  
Hu Jin ◽  
Michael Gapinske ◽  
Wendy S Woods ◽  
...  

Abstract Cytosine methylation is a ubiquitous modification in mammalian DNA generated and maintained by several DNA methyltransferases (DNMTs) with partially overlapping functions and genomic targets. To systematically dissect the factors specifying each DNMT’s activity, we engineered combinatorial knock-in of human DNMT genes in Komagataella phaffii, a yeast species lacking endogenous DNA methylation. Time-course expression measurements captured dynamic network-level adaptation of cells to DNMT3B1-induced DNA methylation stress and showed that coordinately modulating the availability of S-adenosyl methionine (SAM), the essential metabolite for DNMT-catalyzed methylation, is an evolutionarily conserved epigenetic stress response, also implicated in several human diseases. Convolutional neural networks trained on genome-wide CpG-methylation data learned distinct sequence preferences of DNMT3 family members. A simulated annealing interpretation method resolved these preferences into individual flanking nucleotides and periodic poly(A) tracts that rotationally position highly methylated cytosines relative to phased nucleosomes. Furthermore, the nucleosome repeat length defined the spatial unit of methylation spreading. Gene methylation patterns were similar to those in mammals, and hypo- and hypermethylation were predictive of increased and decreased transcription relative to control, respectively, in the absence of mammalian readers of DNA methylation. Introducing controlled epigenetic perturbations in yeast thus enabled characterization of fundamental genomic features directing specific DNMT3 proteins.


2019 ◽  
Vol 63 (6) ◽  
pp. 649-661 ◽  
Author(s):  
Oscar Ortega-Recalde ◽  
Timothy Alexander Hore

Abstract Cytosine methylation is a DNA modification that is critical for vertebrate development and provides a plastic yet stable information module in addition to the DNA code. DNA methylation memory establishment, maintenance and erasure is carefully balanced by molecular machinery highly conserved among vertebrates. In mammals, extensive erasure of epigenetic marks, including 5-methylcytosine (5mC), is a hallmark of early embryo and germline development. Conversely, global cytosine methylation patterns are preserved in at least some non-mammalian vertebrates over comparable developmental windows. The evolutionary mechanisms which drove this divergence are unknown, nevertheless a direct consequence of retaining epigenetic memory in the form of 5mC is the enhanced potential for transgenerational epigenetic inheritance (TEI). Given that DNA methylation dynamics remains underexplored in most vertebrate lineages, the extent of information transferred to offspring by epigenetic modification might be underestimated.


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