scholarly journals Human mitochondrial DNA is extensively methylated in a non-CpG context

2019 ◽  
Vol 47 (19) ◽  
pp. 10072-10085 ◽  
Author(s):  
Vibha Patil ◽  
Cyrille Cuenin ◽  
Felicia Chung ◽  
Jesus R Rodriguez Aguilera ◽  
Nora Fernandez-Jimenez ◽  
...  
Keyword(s):  
Gene Expression ◽  
Mitochondrial Dna ◽  
Dna Methyltransferase ◽  
Molecular Mechanisms ◽  
Dna Methyltransferases ◽  
Functional Link ◽  
Functional Consequences ◽  
Single Base Pair ◽  
Methylation Patterns ◽  
Global Reduction

Abstract Mitochondrial dysfunction plays critical roles in cancer development and related therapeutic response; however, exact molecular mechanisms remain unclear. Recently, alongside the discovery of mitochondrial-specific DNA methyltransferases, global and site-specific methylation of the mitochondrial genome has been described. Investigation of any functional consequences however remains unclear and debated due to insufficient evidence of the quantitative degree and frequency of mitochondrial DNA (mtDNA) methylation. This study uses WGBS to provide the first quantitative report of mtDNA methylation at single base pair resolution. The data show that mitochondrial genomes are extensively methylated predominantly at non-CpG sites. Importantly, these methylation patterns display notable differences between normal and cancer cells. Furthermore, knockdown of DNA methyltransferase enzymes resulted in a marked global reduction of mtDNA methylation levels, indicating these enzymes may be associated with the establishment and/or maintenance of mtDNA methylation. DNMT3B knockdown cells displayed a comparatively pronounced global reduction in mtDNA methylation with concomitant increases in gene expression, suggesting a potential functional link between methylation and gene expression. Together these results demonstrate reproducible, non-random methylation patterns of mtDNA and challenge the notion that mtDNA is lowly methylated. This study discusses key differences in methodology that suggest future investigations must allow for techniques that assess both CpG and non-CpG methylation.

scholarly journals Selective Anchoring of DNA Methyltransferases 3A and 3B to Nucleosomes Containing Methylated DNA

2009 ◽  
Vol 29 (19) ◽  
pp. 5366-5376 ◽  
Author(s):  
Shinwu Jeong ◽  
Gangning Liang ◽  
Shikhar Sharma ◽  
Joy C. Lin ◽  
Si Ho Choi ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Dna Methyltransferase ◽  
Molecular Mechanisms ◽  
Dna Methyltransferases ◽  
Micrococcal Nuclease ◽  
Nuclear Antigen ◽  
Heterochromatin Protein 1 ◽  
Histone Deacetylase 1 ◽  
Methylation Patterns

ABSTRACT Proper DNA methylation patterns are essential for mammalian development and differentiation. DNA methyltransferases (DNMTs) primarily establish and maintain global DNA methylation patterns; however, the molecular mechanisms for the generation and inheritance of methylation patterns are still poorly understood. We used sucrose density gradients of nucleosomes prepared by partial and maximum micrococcal nuclease digestion, coupled with Western blot analysis to probe for the interactions between DNMTs and native nucleosomes. This method allows for analysis of the in vivo interactions between the chromatin modification enzymes and their actual nucleosomal substrates in the native state. We show that little free DNA methyltransferase 3A and 3B (DNMT3A/3B) exist in the nucleus and that almost all of the cellular contents of DNMT3A/3B, but not DNMT1, are strongly anchored to a subset of nucleosomes. This binding of DNMT3A/3B does not require the presence of other well-known chromatin-modifying enzymes or proteins, such as proliferating cell nuclear antigen, heterochromatin protein 1, methyl-CpG binding protein 2, Enhancer of Zeste homolog 2, histone deacetylase 1, and UHRF1, but it does require an intact nucleosomal structure. We also show that nucleosomes containing methylated SINE and LINE elements and CpG islands are the main sites of DNMT3A/3B binding. These data suggest that inheritance of DNA methylation requires cues from the chromatin component in addition to hemimethylation.

scholarly journals Mapping chromatin accessibility and active regulatory elements reveals pathological mechanisms in human gliomas

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Karolina Stępniak ◽  
Magdalena A. Machnicka ◽  
Jakub Mieczkowski ◽  
Anna Macioszek ◽  
Bartosz Wojtaś ◽  
...  
Keyword(s):  
Gene Expression ◽  
Chromatin Structure ◽  
Molecular Mechanisms ◽  
Genome Mapping ◽  
Malignant Gliomas ◽  
Regulatory Elements ◽  
Chromatin Accessibility ◽  
Multiple Tumor ◽  
Genes Encoding ◽  
Methylation Patterns

AbstractChromatin structure and accessibility, and combinatorial binding of transcription factors to regulatory elements in genomic DNA control transcription. Genetic variations in genes encoding histones, epigenetics-related enzymes or modifiers affect chromatin structure/dynamics and result in alterations in gene expression contributing to cancer development or progression. Gliomas are brain tumors frequently associated with epigenetics-related gene deregulation. We perform whole-genome mapping of chromatin accessibility, histone modifications, DNA methylation patterns and transcriptome analysis simultaneously in multiple tumor samples to unravel epigenetic dysfunctions driving gliomagenesis. Based on the results of the integrative analysis of the acquired profiles, we create an atlas of active enhancers and promoters in benign and malignant gliomas. We explore these elements and intersect with Hi-C data to uncover molecular mechanisms instructing gene expression in gliomas.

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 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 DNA methyltransferase 1 (DNMT1) function is implicated in the age-related loss of cortical interneurons

2020 ◽  
Author(s):  
Anne Hahn ◽  
Cathrin Bayer ◽  
Daniel Pensold ◽  
Jessica Tittelmeier ◽  
Lisa Marx-Blümel ◽  
...  
Keyword(s):  
Dna Methyltransferase ◽  
Molecular Mechanisms ◽  
Life Time ◽  
Modern Society ◽  
Dna Methyltransferases ◽  
Control Of Gene Expression ◽  
Inhibitory Circuits ◽  
Age Related ◽  
Transcriptional Changes ◽  
The Cost

AbstractIncreased 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. Deletion of Dnmt1 in parvalbumin-positive interneurons attenuates their age-related decline in the cerebral cortex. Moreover, DNMT1-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 suggested to be essentially 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.

scholarly journals A RID-like cytosine methyltransferase homologue controls sexual development in the fungusPodospora anserina

2019 ◽  
Author(s):  
P Grognet ◽  
H Timpano ◽  
F Carlier ◽  
J Aït-Benkhali ◽  
V Berteaux-Lecellier ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Sexual Reproduction ◽  
Sexual Development ◽  
Genomic Dna ◽  
Dna Methyltransferase ◽  
Dna Methyltransferases ◽  
Female Sterility ◽  
Developmental Pathway ◽  
Important Group ◽  
Methylation Patterns

AbstractDNA methyltransferases are ubiquitous enzymes conserved in bacteria, plants and opisthokonta. These enzymes, which methylate cytosines, are involved in numerous biological processes, notably development. In mammals and higher plants, methylation patterns established and maintained by the cytosine DNA methyltransferases (DMTs) are essential to zygotic development. In fungi, some members of an extensively conserved fungal-specific DNA methyltransferase class are both mediators of the Repeat Induced Point mutation (RIP) genome defense system and key players of sexual reproduction. Yet, no DNA methyltransferase activity of these purified RID (RIP deficient) proteins could be detectedin vitro. These observations led us to explore how RID-like DNA methyltransferase encoding genes would play a role during sexual development of fungi showing very little genomic DNA methylation, if any.To do so, we used the model ascomycete fungusP. anserina. We identified thePaRidgene, encoding a RID-like DNA methyltransferase and constructed knocked-out ΔPaRiddefective mutants. Crosses involvingP. anserinaΔPaRidmutants are sterile. Our results show that, although gametes are readily formed and fertilization occurs in a ΔPaRidbackground, sexual development is blocked just before the individualization of the dikaryotic cells leading to meiocytes. Complementation of ΔPaRidmutants with ectopic alleles ofPaRid, including GFP-tagged, point-mutated, inter-specific and chimeric alleles, demonstrated that the catalytic motif of the putative PaRid methyltransferase is essential to ensure proper sexual development and that the expression of PaRid is spatially and temporally restricted. A transcriptomic analysis performed on mutant crosses revealed an overlap of the PaRid-controlled genetic network with the well-known mating-types gene developmental pathway common to an important group of fungi, the Pezizomycotina.Author SummarySexual reproduction is considered to be essential for long-term persistence of eukaryotic species. Sexual reproduction is controlled by strict mechanisms governing which haploids can fuse (mating) and which developmental paths the resulting zygote will follow. In mammals, differential genomic DNA methylation patterns of parental gametes, known as ‘DNA methylation imprints’ are essential to zygotic development, while in plants, global genomic demethylation often results in female-sterility. Although animal and fungi are evolutionary related, little is known about epigenetic regulation of gene expression and development in multicellular fungi. Here, we report on a gene of the model fungusPodospora anserina, encoding a protein called PaRid that looks like a DNA methyltrasferase. We showed that expression of the catalytically functional version of the PaRid protein is required in the maternal parental strain to form zygotes. By establishing the transcriptional profile ofPaRidmutant strain, we identified a set of PaRid direct and/or indirect target genes. Half of them are also targets of a mating-type transcription factor known to be a major regulator of sexual development. So far, there was no other example of identified RID targets shared with a well-known developmental pathway that is common to an important group of fungi, the Pezizomycotina

scholarly journals GuidingNet: revealing transcriptional cofactor and predicting binding for DNA methyltransferase by network regularization

2020 ◽  
Author(s):  
Lixin Ren ◽  
Caixia Gao ◽  
Zhana Duren ◽  
Yong Wang
Keyword(s):  
Dna Methyltransferase ◽  
Dna Methyltransferases ◽  
Chromatin Accessibility ◽  
Protein Interaction Data ◽  
Genomic Locus ◽  
Protein Protein Interaction ◽  
Chromatin Regulators ◽  
Regularized Methods ◽  
Single Data ◽  
Methylation Patterns

Abstract The DNA methyltransferases (DNMTs) (DNMT3A, DNMT3B and DNMT3L) are primarily responsible for the establishment of genomic locus-specific DNA methylation patterns, which play an important role in gene regulation and animal development. However, this important protein family’s binding mechanism, i.e. how and where the DNMTs bind to genome, is still missing in most tissues and cell lines. This motivates us to explore DNMTs and TF’s cooperation and develop a network regularized logistic regression model, GuidingNet, to predict DNMTs’ genome-wide binding by integrating gene expression, chromatin accessibility, sequence and protein–protein interaction data. GuidingNet accurately predicted methylation experimental data validated DNMTs’ binding, outperformed single data source based and sparsity regularized methods and performed well in within and across tissue prediction for several DNMTs in human and mouse. Importantly, GuidingNet can reveal transcription cofactors assisting DNMTs for methylation establishment. This provides biological understanding in the DNMTs’ binding specificity in different tissues and demonstrate the advantage of network regularization. In addition to DNMTs, GuidingNet achieves good performance for other chromatin regulators’ binding. GuidingNet is freely available at https://github.com/AMSSwanglab/GuidingNet.

scholarly journals Cooperativity between DNA Methyltransferases in the Maintenance Methylation of Repetitive Elements

2002 ◽  
Vol 22 (2) ◽  
pp. 480-491 ◽  
Author(s):  
Gangning Liang ◽  
Matilda F. Chan ◽  
Yoshitaka Tomigahara ◽  
Yvonne C. Tsai ◽  
Felicidad A. Gonzales ◽  
...  
Keyword(s):  
Dna Methyltransferase ◽  
De Novo ◽  
Es Cells ◽  
Repetitive Sequences ◽  
Embryonic Stem ◽  
Dna Methyltransferases ◽  
Dna Methyltransferase 1 ◽  
Gene Knockouts ◽  
Methylation Patterns ◽  
Maintenance Methylation

ABSTRACT We used mouse embryonic stem (ES) cells with systematic gene knockouts for DNA methyltransferases to delineate the roles of DNA methyltransferase 1 (Dnmt1) and Dnmt3a and -3b in maintaining methylation patterns in the mouse genome. Dnmt1 alone was able to maintain methylation of most CpG-poor regions analyzed. In contrast, both Dnmt1 and Dnmt3a and/or Dnmt3b were required for methylation of a select class of sequences which included abundant murine LINE-1 promoters. We used a novel hemimethylation assay to show that even in wild-type cells these sequences contain high levels of hemimethylated DNA, suggestive of poor maintenance methylation. We showed that Dnmt3a and/or -3b could restore methylation of these sequences to pretreatment levels following transient exposure of cells to 5-aza-CdR, whereas Dnmt1 by itself could not. We conclude that ongoing de novo methylation by Dnmt3a and/or Dnmt3b compensates for inefficient maintenance methylation by Dnmt1 of these endogenous repetitive sequences. Our results reveal a previously unrecognized degree of cooperativity among mammalian DNA methyltransferases in ES cells.

Imprinted and DNA methyltransferase gene expression in the endometrium during the pre- and peri-implantation period in cattle

2017 ◽  
Vol 29 (9) ◽  
pp. 1729 ◽  
Author(s):  
A. M. O'Doherty ◽  
L. C. O'Shea ◽  
O. Sandra ◽  
P. Lonergan ◽  
T. Fair ◽  
...  
Keyword(s):  
Gene Expression ◽  
Dna Methyltransferase ◽  
Expression Profiles ◽  
Dna Methyltransferases ◽  
Imprinted Genes ◽  
Early Stages ◽  
Methyltransferase Gene ◽  
Implantation Period ◽  
Epigenetic Processes

The endometrium plays a key role in providing an optimal environment for attachment of the preimplantation embryo during the early stages of pregnancy. Investigations over the past 2 decades have demonstrated that vital epigenetic processes occur in the embryo during the preimplantation stages of development. However, few studies have investigated the potential role of imprinted genes and their associated modulators, the DNA methyltransferases (DNMTs), in the bovine endometrium during the pre- and peri-implantation period. Therefore, in the present study we examined the expression profiles of the DNMT genes (3A, 3A2 and 3B) and a panel of the most comprehensively studied imprinted genes in the endometrium of cyclic and pregnant animals. Intercaruncular (Days 5, 7, 13, 16 and 20) and caruncular (Days 16 and 20) regions were analysed for gene expression changes, with protein analysis also performed for DNMT3A, DNMT3A2 and DNMT3B on Days 16 and 20. An overall effect of day was observed for expression of several of the imprinted genes. Tissue-dependent gene expression was detected for all genes at Day 20. Differences in DNMT protein abundance were mostly observed in the intercaruncular regions of pregnant heifers at Day 16 when DNMT3A, DNMT3A2 and DNMT3B were all lower when compared with cyclic controls. At Day 20, DNMT3A2 expression was lower in the pregnant caruncular samples compared with cyclic animals. This study provides evidence that epigenetic mechanisms in the endometrium may be involved with implantation of the embryo during the early stages of pregnancy in cattle.

scholarly journals DNA Methylation Profiling of Pediatric AML Reveals That Hypomethylation of MN1 Is Characteristic of Inv(16) AML and a Driver of MN1 Overexpression

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 867-867
Author(s):  
Nicole S.D. Larmonie ◽  
Marry M. van den Heuvel-Eibrink ◽  
Askar Obulkasim ◽  
Valerie de Haas ◽  
Dirk Reinhardt ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Dna Methyltransferase ◽  
Cpg Island ◽  
Dna Methyltransferases ◽  
Genetic Abnormalities ◽  
Pediatric Aml ◽  
Methylation Patterns ◽  
Dnmt3b Expression ◽  
Methylation Profiling ◽  
Epigenetic Processes

Abstract Primary refractory and relapsed pediatric acute myeloid leukemia (AML) still lead to a significant number of childhood cancer deaths, despite the current chemotherapeutic regimens. AML leukemogenesis is driven by collaborative genetic abnormalities that induce hematopoietic maturation arrest and cell proliferation. Particular AML-associated maturation inhibiting aberrations are known to target chromatin regulators, thus directly influencing the transcriptional program of leukemic cells. Therapies targeting epigenetic processes, e.g. with hypomethylation-inducing agents, are therefore becoming an attractive therapeutic strategy in adult AML. AML biology in children is not equivalent to that of adults, thus methylation patterns seen in adult AML cannot be extrapolated to pediatric AML. Therefore there is a need to unravel the mechanism behind changes in epigenetic processes as the result of AML-causing genetic abnormalities in order to develop new drugs for pediatric AML. We hypothesized that pediatric AML samples have distinct DNA-methylation patterns which may provide a rationale for treatment with demethylating agents in specific pediatric AML subtypes. Furthermore, these differences in methylation could be characteristic for AML subgroups and that particular methylation patterns drive the expression of specific genes which may play a key role in the tumorigenesis of these AML leukemias. We performed genome-wide CpG-island methylation profiling on a representative and molecularly characterized cohort of pediatric patients with de novo AML. Empirical Bayes Wilcoxon rank-sum test showed that AML patients carrying inv(16)(p13;q22) (n=9) have distinct DNA methylation patterns when compared to non-inv(16) AML patients (n=143) (consisting mainly of MLL-rearranged, t(8;21), t(15;17), t(8;16) AML and AML cases with a normal karyotype). The MN1 gene ranked as most significantly differentially methylated in inv(16) AML compared to non-inv(16) AML, with inv(16) AML cases having significantly (p=2x10-6) lower methylation levels compared to non-inv(16) AML cases. Hypomethylation of specific regions of the MN1-associated CpG-island was confirmed by methylation specific PCR and bisulfite sequencing. Subsequent gene expression (GEP) data on 294 pediatric AML patients showed that MN1 was 8 fold higher expressed in patients carrying inv(16) compared to all other patients (9.9, n=35 vs 6.9, n=259, p<0.001). Furthermore, integrating GEP and methylation array data showed that MN1 expression negatively correlated (ρs= 0.82, p=0.011) with methylation levels, which is in agreement with the biological assumption of methylation and gene expression. Since genes known to regulate DNA methylation have frequently been shown to be mutated in adult AML we determined whether a decreased expression of DNA methyltransferases, DNMT1, DNMT2, DNMT3A, DNMT3B, could be the cause of a hypometylated MN1 locus in inv(16) AML. Our findings show that only DNMT3B expression was significantly (p=8x10-15) lower in inv(16) cases compared to non-inv(16) cases. To test whether hypomethylation of the MN1 CpG-island and the overexpression of MN1 is the result of decreased DNMT1 expression, HL60 cells which express negligible levels of MN1 were treated with the DNMT1 inhibitor Decitabine. This showed that treatment of HL60 cells with Decitabine led to increase of MN1 transcript levels, however, not as high as those observed in patient samples. This suggests that DNMT1 activity may not be the only DNA methyltransferase influencing expression of MN1 in inv(16) patients. Interestingly, we observed a high (ρs= 0.42) correlation between MN1 methylation and DNMT3B expression, which suggests DNMT3B could be an important DNA methyltransferase involved in regulating MN1expression. Overall we show that pediatric AML patients carrying and inv(16) have a characteristic DNA methylation pattern compared to other AML patients carrying specific cytogenetic aberrations. Furthermore, our data suggest that hypomethylation of the MN1 gene is an underlying mechanism for high MN1 expression in inv(16)(p13;q22) patients possibly regulated by multiple DNA methyltransferases. Disclosures No relevant conflicts of interest to declare.

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