scholarly journals Inference of DNA methylation patterns in molluscs

2021 ◽  
Vol 376 (1825) ◽  
Author(s):  
Lisa Männer ◽  
Tilman Schell ◽  
Panagiotis Provataris ◽  
Martin Haase ◽  
Carola Greve
Keyword(s):  
Dna Methylation ◽  
De Novo ◽  
Morphological Diversity ◽  
Dna Methyltransferases ◽  
Animal Kingdom ◽  
Future Directions ◽  
Protein Coding ◽  
General Picture ◽  
Methylation Patterns ◽  
De Novo Methylation

Mollusca are the second largest and arguably most diverse phylum of the animal kingdom. This is in sharp contrast to our very limited knowledge concerning epigenetic mechanisms including DNA methylation in this invertebrate group. Here, we inferred DNA methylation patterns by analysing the normalized dinucleotide CG content in protein-coding sequences and identified DNA methyltransferases (DNMT1 and 3) in published transcriptomes and genomes of 140 species across all eight classes of molluscs. Given the evolutionary age and morphological diversity of molluscs, we expected to find evidence for diverse methylation patterns. Our inferences suggest that molluscs possess substantial levels of DNA methylation in gene bodies as a rule. Yet, we found deviations from this general picture with regard to (i) the CpG observed/expected distributions indicating a reduction in DNA methylation in certain groups and (ii) the completeness of the DNMT toolkit. Reductions were evident in Caudofoveata, Solenogastres, Polyplacophora, Monoplacophora, as well as Scaphopoda. Heterobranchia and Oegopsida were remarkable as they lacked DNMT3, usually responsible for de novo methylation, yet showed signs of DNA methylation. Our survey may serve as guidance for direct empirical analyses of DNA methylation in molluscs. This article is part of the Theo Murphy meeting issue ‘Molluscan genomics: broad insights and future directions for a neglected phylum’.

scholarly journals Establishment and Maintenance of Genomic Methylation Patterns in Mouse Embryonic Stem Cells by Dnmt3a and Dnmt3b

2003 ◽  
Vol 23 (16) ◽  
pp. 5594-5605 ◽  
Author(s):  
Taiping Chen ◽  
Yoshihide Ueda ◽  
Jonathan E. Dodge ◽  
Zhenjuan Wang ◽  
En Li
Keyword(s):  
Dna Methylation ◽  
De Novo ◽  
Es Cells ◽  
Embryonic Stem ◽  
Single Copy ◽  
Dna Methyltransferases ◽  
Methylation Patterns ◽  
Genomic Methylation ◽  
De Novo Methylation

ABSTRACT We have previously shown that the DNA methyltransferases Dnmt3a and Dnmt3b carry out de novo methylation of the mouse genome during early postimplantation development and of maternally imprinted genes in the oocyte. In the present study, we demonstrate that Dnmt3a and Dnmt3b are also essential for the stable inheritance, or “maintenance,” of DNA methylation patterns. Inactivation of both Dnmt3a and Dnmt3b in embryonic stem (ES) cells results in progressive loss of methylation in various repeats and single-copy genes. Interestingly, introduction of the Dnmt3a, Dnmt3a2, and Dnmt3b1 isoforms back into highly demethylated mutant ES cells restores genomic methylation patterns; these isoforms appear to have both common and distinct DNA targets, but they all fail to restore the maternal methylation imprints. In contrast, overexpression of Dnmt1 and Dnmt3b3 failed to restore DNA methylation patterns due to their inability to catalyze de novo methylation in vivo. We also show that hypermethylation of genomic DNA by Dnmt3a and Dnmt3b is necessary for ES cells to form teratomas in nude mice. These results indicate that genomic methylation patterns are determined partly through differential expression of different Dnmt3a and Dnmt3b isoforms.

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 Comprehensive structure-function characterization of DNMT3B and DNMT3A reveals distinctive de novo DNA methylation mechanisms

2020 ◽  
Author(s):  
Linfeng Gao ◽  
Max Emperle ◽  
Yiran Guo ◽  
Sara A Grimm ◽  
Wendan Ren ◽  
...  
Keyword(s):  
Dna Methylation ◽  
De Novo ◽  
Target Selection ◽  
Dna Methyltransferases ◽  
Recognition Mechanism ◽  
Layered Substrate ◽  
Structural Enzymology ◽  
Catalytic Loop ◽  
Methylation Patterns

AbstractMammalian DNA methylation patterns are established by two de novo DNA methyltransferases DNMT3A and DNMT3B, which exhibit both redundant and distinctive methylation activities. However, the related molecular basis remains undetermined. Through comprehensive structural, enzymology and cellular characterization of DNMT3A and DNMT3B, we here report a multi-layered substrate-recognition mechanism underpinning their divergent genomic methylation activities. A hydrogen bond in the catalytic loop of DNMT3B causes a lower CpG specificity than DNMT3A, while the interplay of target recognition domain and homodimeric interface fine-tunes the distinct target selection between the two enzymes, with Lysine 777 of DNMT3B acting as a unique sensor of the +1 flanking base. The divergent substrate preference between DNMT3A and DNMT3B provides an explanation for site-specific epigenomic alterations seen in ICF syndrome with DNMT3B mutations. Together, this study reveals crucial and distinctive substrate-readout mechanisms of the two DNMT3 enzymes, implicative of their differential roles during development and pathogenesis.

scholarly journals DNA Methylation Density Influences the Stability of an Epigenetic Imprint and Dnmt3a/b-Independent De Novo Methylation

2002 ◽  
Vol 22 (21) ◽  
pp. 7572-7580 ◽  
Author(s):  
Matthew C. Lorincz ◽  
Dirk Schübeler ◽  
Shauna R. Hutchinson ◽  
David R. Dickerson ◽  
Mark Groudine
Keyword(s):  
Dna Methylation ◽  
Transcriptional Activation ◽  
Transcriptional Repression ◽  
De Novo ◽  
Embryonic Stem ◽  
Dna Methyltransferases ◽  
The Stability ◽  
Murine Erythroleukemia ◽  
De Novo Methylation

ABSTRACT DNA methylation plays an important role in transcriptional repression. To gain insight into the dynamics of demethylation and de novo methylation, we introduced a proviral reporter, premethylated at different densities, into a defined chromosomal site in murine erythroleukemia cells and monitored the stability of the introduced methylation and reporter gene expression. A high density of methylation was faithfully propagated in vivo. In contrast, a low level of methylation was not stable, with complete demethylation and associated transcriptional activation or maintenance-coupled de novo methylation and associated silencing occurring with equal probability. Deletion of the proviral enhancer increased the probability of maintenance-coupled de novo methylation, suggesting that this enhancer functions in part to antagonize such methylation. The DNA methyltransferases (MTases) Dnmt3a and Dnmt3b are thought to be the sole de novo MTases in the mammalian genome. To determine whether these enzymes are responsible for maintenance-coupled de novo methylation, the unmethylated or premethylated proviral reporter was introduced into DNA MTase-deficient embryonic stem cells. These studies revealed the presence of a Dnmt3a/Dnmt3b-independent de novo methyltransferase activity that is stimulated by the presence of preexisting methylation.

scholarly journals Comparative analysis of DNA methylation patterns in transgenic Drosophila overexpressing mouse DNA methyltransferases

2004 ◽  
Vol 378 (3) ◽  
pp. 763-768 ◽  
Author(s):  
Cora MUND ◽  
Tanja MUSCH ◽  
Martin STRÖDICKE ◽  
Birte ASSMANN ◽  
En LI ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Mammalian Cells ◽  
Cytosine Methylation ◽  
De Novo ◽  
Epigenetic Modification ◽  
Dna Methyltransferases ◽  
Significant Activity ◽  
Cpg Dinucleotides ◽  
Methylation Patterns ◽  
Transgenic Drosophila

DNA methyltransferases (Dnmts) mediate the epigenetic modification of eukaryotic genomes. Mammalian DNA methylation patterns are established and maintained by co-operative interactions among the Dnmt proteins Dnmt1, Dnmt3a and Dnmt3b. Owing to their simultaneous presence in mammalian cells, the activities of individual Dnmt have not yet been determined. This includes a fourth putative Dnmt, namely Dnmt2, which has failed to reveal any activity in previous assays. We have now established transgenic Drosophila strains that allow for individual overexpression of all known mouse Dnmts. Quantitative analysis of genomic cytosine methylation levels demonstrated a robust Dnmt activity for the de novo methyltransferases Dnmt3a and Dnmt3b. In addition, we also detected a weak but significant activity for Dnmt2. Subsequent methylation tract analysis by genomic bisulphite sequencing revealed that Dnmt3 enzymes preferentially methylated CpG dinucleotides in a processive manner, whereas Dnmt2 methylated isolated cytosine residues in a non-CpG dinucleotide context. Our results allow a direct comparison of the activities of mammalian Dnmts and suggest a significant functional specialization of these enzymes.

scholarly journals Flanking sequence preference modulates de novo DNA methylation in the mouse genome

2020 ◽  
Author(s):  
Izaskun Mallona ◽  
Ioana Mariuca Ilie ◽  
Ino Dominiek Karemaker ◽  
Stefan Butz ◽  
Massimiliano Manzo ◽  
...  
Keyword(s):  
Dna Methylation ◽  
De Novo ◽  
Cell Types ◽  
Dna Methyltransferases ◽  
Side Chain ◽  
Flanking Sequence ◽  
Base Pairs ◽  
Sequence Preference ◽  
Dynamics Simulations ◽  
Methylation Patterns

Abstract Mammalian de novo DNA methyltransferases (DNMT) are responsible for the establishment of cell-type-specific DNA methylation in healthy and diseased tissues. Through genome-wide analysis of de novo methylation activity in murine stem cells we uncover that DNMT3A prefers to methylate CpGs followed by cytosines or thymines, while DNMT3B predominantly methylates CpGs followed by guanines or adenines. These signatures are further observed at non-CpG sites, resembling methylation context observed in specialised cell types, including neurons and oocytes. We further show that these preferences result from structural differences in the catalytic domains of the two de novo DNMTs and are not a consequence of differential recruitment to the genome. Molecular dynamics simulations suggest that, in case of human DNMT3A, the preference is due to favourable polar interactions between the flexible Arg836 side chain and the guanine that base-pairs with the cytosine following the CpG. By exchanging arginine to a lysine, the corresponding side chain in DNMT3B, the sequence preference is reversed, confirming the requirement for arginine at this position. This context-dependent enzymatic activity provides additional insights into the complex regulation of DNA methylation patterns.

scholarly journals Flanking sequence preference modulates de novo DNA methylation in the mouse genome

2020 ◽  
Author(s):  
Izaskun Mallona ◽  
Ioana Mariuca Ilie ◽  
Massimiliano Manzo ◽  
Amedeo Caflisch ◽  
Tuncay Baubec
Keyword(s):  
Dna Methylation ◽  
De Novo ◽  
Cell Types ◽  
Dna Methyltransferases ◽  
Flanking Sequence ◽  
Base Pairs ◽  
Genome Wide ◽  
Dynamics Simulations ◽  
Methylation Patterns ◽  
Different Cell Types

AbstractMammalian de novo DNA methyltransferases (DNMT) are responsible for the establishment of cell-type-specific DNA methylation in healthy and diseased tissues. Through genome-wide analysis of de novo methylation activity in murine stem cells we uncover that DNMT3A prefers to methylate CpGs followed by cytosines or thymines, while DNMT3B predominantly methylates CpGs followed by guanines or adenines. These signatures are further observed at non-CpG sites, resembling methylation context observed in specialised cell types, including neurons and oocytes. We further show that these preferences are not mediated by the differential recruitment of the two de novo DNMTs to the genome but are resulting from structural differences in their catalytic domains. Molecular dynamics simulations suggest that, in case of DNMT3A, the preference is due to favourable polar interactions between the flexible Arg836 side chain and the guanine that base-pairs with the cytosine following the CpG. This context-dependent de novo DNA methylation provides additional insights into the complex regulation of methylation patterns in different cell types.

scholarly journals Interplay between Histone and DNA Methylation Seen through Comparative Methylomes in Rare Mendelian Disorders

2021 ◽  
Vol 22 (7) ◽  
pp. 3735
Author(s):  
Guillaume Velasco ◽  
Damien Ulveling ◽  
Sophie Rondeau ◽  
Pauline Marzin ◽  
Motoko Unoki ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Catalytic Domain ◽  
De Novo ◽  
Mutation Screening ◽  
Histone H3 ◽  
Regulatory Elements ◽  
Germline Mutations ◽  
Dna Methyltransferases ◽  
Mendelian Disorders ◽  
Epigenetic Machinery

DNA methylation (DNAme) profiling is used to establish specific biomarkers to improve the diagnosis of patients with inherited neurodevelopmental disorders and to guide mutation screening. In the specific case of mendelian disorders of the epigenetic machinery, it also provides the basis to infer mechanistic aspects with regard to DNAme determinants and interplay between histone and DNAme that apply to humans. Here, we present comparative methylomes from patients with mutations in the de novo DNA methyltransferases DNMT3A and DNMT3B, in their catalytic domain or their N-terminal parts involved in reading histone methylation, or in histone H3 lysine (K) methylases NSD1 or SETD2 (H3 K36) or KMT2D/MLL2 (H3 K4). We provide disease-specific DNAme signatures and document the distinct consequences of mutations in enzymes with very similar or intertwined functions, including at repeated sequences and imprinted loci. We found that KMT2D and SETD2 germline mutations have little impact on DNAme profiles. In contrast, the overlapping DNAme alterations downstream of NSD1 or DNMT3 mutations underlines functional links, more specifically between NSD1 and DNMT3B at heterochromatin regions or DNMT3A at regulatory elements. Together, these data indicate certain discrepancy with the mechanisms described in animal models or the existence of redundant or complementary functions unforeseen in humans.

Decreased expression of DNA methyltransferases in the testes of patients with non-obstructive azoospermia leads to changes in global DNA methylation levels

2019 ◽  
Vol 31 (8) ◽  
pp. 1386 ◽  
Author(s):  
Fatma Uysal ◽  
Gokhan Akkoyunlu ◽  
Saffet Ozturk
Keyword(s):  
Dna Methylation ◽  
Male Infertility ◽  
De Novo ◽  
Biological Effects ◽  
Testicular Biopsy ◽  
Round Spermatid ◽  
Dna Methyltransferases ◽  
Global Dna Methylation ◽  
Obstructive Azoospermia ◽  
Spermatogenic Cells

DNA methylation plays key roles in epigenetic regulation during mammalian spermatogenesis. DNA methyltransferases (DNMTs) function in de novo and maintenance methylation processes by adding a methyl group to the fifth carbon atom of the cytosine residues within cytosine–phosphate–guanine (CpG) and non-CpG dinucleotide sites. Azoospermia is one of the main causes of male infertility, and is classified as obstructive (OA) or non-obstructive (NOA) azoospermia based on histopathological characteristics. The molecular background of NOA is still largely unknown. DNA methylation performed by DNMTs is implicated in the transcriptional regulation of spermatogenesis-related genes. The aim of the present study was to evaluate the cellular localisation and expression levels of the DNMT1, DNMT3A and DNMT3B proteins, as well as global DNA methylation profiles in testicular biopsy samples obtained from men with various types of NOA, including hypospermatogenesis (hyposperm), round spermatid (RS) arrest, spermatocyte (SC) arrest and Sertoli cell-only (SCO) syndrome. In the testicular biopsy samples, DNMT1 expression and global DNA methylation levels decreased gradually from the hyposperm to SCO groups (P<0.05). DNMT3A expression was significantly decreased in the RS arrest, SC arrest and SCO groups compared with the hyposperm group (P<0.05). DNMT3B expression was significantly lower in the RS arrest and SCO groups than in the hyposperm group (P<0.05). Although both DNMT1 and DNMT3A were localised in the cytoplasm and nucleus of the spermatogenic cells, staining for DNMT3B was more intensive in the nucleus of spermatogenic cells. In conclusion, the findings suggest that significant changes in DNMT expression and global DNA methylation levels in spermatogenic cells may contribute to development of male infertility in the NOA groups. Further studies are needed to determine the molecular biological effects of the altered DNMT expression and DNA methylation levels on development of male infertility.

A targeted-replacement system for identification of signals for de novo methylation in Neurospora crassa

1994 ◽  
Vol 14 (11) ◽  
pp. 7059-7067
Author(s):  
V P Miao ◽  
M J Singer ◽  
M R Rountree ◽  
E U Selker
Keyword(s):  
Dna Methylation ◽  
Neurospora Crassa ◽  
De Novo ◽  
Gene Replacement ◽  
Position Effects ◽  
Efficient Gene ◽  
Bona Fide ◽  
Replacement System ◽  
Methylation Signal ◽  
De Novo Methylation

Transformation of eukaryotic cells can be used to test potential signals for DNA methylation. This approach is not always reliable, however, because of chromosomal position effects and because integration of multiple and/or rearranged copies of transforming DNA can influence DNA methylation. We developed a robust system to evaluate the potential of DNA fragments to function as signals for de novo methylation in Neurospora crassa. The requirements of the system were (i) a location in the N. crassa genome that becomes methylated only in the presence of a bona fide methylation signal and (ii) an efficient gene replacement protocol. We report here that the am locus fulfills these requirements, and we demonstrate its utility with the identification of a 2.7-kb fragment from the psi 63 locus as a new portable signal for de novo methylation.

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