scholarly journals Genomic profiling of PML bodies reveals transcriptional regulation by PML bodies through the DNMT3A exclusion

2019 ◽  
Author(s):  
Misuzu Kurihara ◽  
Kagayaki Kato ◽  
Chiaki Sanbo ◽  
Shuji Shigenobu ◽  
Yasuyuki Ohkawa ◽  
...  
Keyword(s):  
Transcriptional Regulation ◽  
Dna Methyltransferase ◽  
De Novo ◽  
Embryonic Stem ◽  
Nuclear Space ◽  
Gene Promoters ◽  
Chromatin Regulators ◽  
Pml Bodies ◽  
Nuclear Structures ◽  
Nuclear Spaces

AbstractThe promyelocytic leukaemia (PML) body is a phase-separated nuclear structure involved in various biological processes, including senescence, and tumour suppression1. PML bodies consist of various proteins, including PML proteins and several chromatin regulators2,3and physically associate with chromatin4,5, implying their crucial roles in particular genome functions. However, their roles in transcriptional regulation are largely unknown. Here, we developed APEX-mediated chromatin labelling and purification (ALaP), to identify the genomic regions associated with PML bodies. We find that PML bodies associate with active regulatory regions across the genome and prominently with a ∼300 kb of the short arm of the Y chromosome (YS300) in mouse embryonic stem cells (mESCs). The association with YS300 is essential for the transcriptional activities of neighbouring Y-linked cluster genes. Mechanistically, we show that PML bodies play a novel role in 3D nuclear organization by providing specific nuclear spaces that the de novo DNA methyltransferase DNMT3A cannot access, which results in the robust maintenance of the hypo-methylated states at the Y-linked gene promoters. Our study underscores a new mechanism for gene regulation in the 3D-nuclear space and provides insights into the functional properties of nuclear structures for genome functions.

De novo DNA cytosine methyltransferase activities in mouse embryonic stem cells

Development ◽  
1996 ◽  
Vol 122 (10) ◽  
pp. 3195-3205 ◽  
Author(s):  
H. Lei ◽  
S.P. Oh ◽  
M. Okano ◽  
R. Juttermann ◽  
K.A. Goss ◽  
...  
Keyword(s):  
Stem Cells ◽  
Embryonic Stem Cells ◽  
Mammalian Cells ◽  
Dna Methyltransferase ◽  
De Novo ◽  
Embryonic Stem ◽  
Cytosine Methyltransferase ◽  
Methyl Cytosine ◽  
Methylation Activity ◽  
De Novo Methylation

It has been a controversial issue as to how many DNA cytosine methyltransferase mammalian cells have and whether de novo methylation and maintenance methylation activities are encoded by a single gene or two different genes. To address these questions, we have generated a null mutation of the only known mammalian DNA methyltransferase gene through homologous recombination in mouse embryonic stem cells and found that the development of the homozygous embryos is arrested prior to the 8-somite stage. Surprisingly, the null mutant embryonic stem cells are viable and contain low but stable levels of methyl cytosine and methyltransferase activity, suggesting the existence of a second DNA methyltransferase in mammalian cells. Further studies indicate that de novo methylation activity is not impaired by the mutation as integrated provirus DNA in MoMuLV-infected homozygous embryonic stem cells become methylated at a similar rate as in wild-type cells. Differentiation of mutant cells results in further reduction of methyl cytosine levels, consistent with the de novo methylation activity being down regulated in differentiated cells. These results provide the first evidence that an independently encoded DNA methyltransferase is present in mammalian cells which is capable of de novo methylating cellular and viral DNA in vivo.

scholarly journals DNA methyltransferase-3–dependent nonrandom template segregation in differentiating embryonic stem cells

2013 ◽  
Vol 203 (1) ◽  
pp. 73-85 ◽  
Author(s):  
Christian Elabd ◽  
Wendy Cousin ◽  
Robert Y. Chen ◽  
Marc S. Chooljian ◽  
Joey T. Pham ◽  
...  
Keyword(s):  
Stem Cells ◽  
Embryonic Stem Cells ◽  
Molecular Mechanism ◽  
Cell Fate ◽  
Dna Methyltransferase ◽  
Daughter Cell ◽  
De Novo ◽  
Embryonic Stem ◽  
Fundamental Property ◽  
Dna Strands

Asymmetry of cell fate is one fundamental property of stem cells, in which one daughter cell self-renews, whereas the other differentiates. Evidence of nonrandom template segregation (NRTS) of chromosomes during asymmetric cell divisions in phylogenetically divergent organisms, such as plants, fungi, and mammals, has already been shown. However, before this current work, asymmetric inheritance of chromatids has never been demonstrated in differentiating embryonic stem cells (ESCs), and its molecular mechanism has remained unknown. Our results unambiguously demonstrate NRTS in asymmetrically dividing, differentiating human and mouse ESCs. Moreover, we show that NRTS is dependent on DNA methylation and on Dnmt3 (DNA methyltransferase-3), indicating a molecular mechanism that regulates this phenomenon. Furthermore, our data support the hypothesis that retention of chromatids with the “old” template DNA preserves the epigenetic memory of cell fate, whereas localization of “new” DNA strands and de novo DNA methyltransferase to the lineage-destined daughter cell facilitates epigenetic adaptation to a new cell fate.

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.

scholarly journals Bivalent chromatin protects reversibly repressed genes from irreversible silencing

2020 ◽  
Author(s):  
Dhirendra Kumar ◽  
Raja Jothi
Keyword(s):  
Dna Methylation ◽  
Regulatory Mechanism ◽  
De Novo ◽  
Embryonic Stem ◽  
Cell Types ◽  
List Type ◽  
Gene Promoters ◽  
Dna Hypermethylation ◽  
Rapid Activation ◽  
Bivalent Gene

ABSTRACTBivalent chromatin is characterized by the simultaneous presence of H3K4me3 and H3K27me3, histone modifications generally associated with transcriptionally active and repressed chromatin, respectively. Prevalent in embryonic stem cells, bivalency is postulated to poise lineage-controlling developmental genes for rapid activation during embryogenesis while maintaining a transcriptionally repressed state in the absence of activation cues, but its function in development and disease remains a mystery. Here we show that bivalency does not poise genes for rapid activation but protects reversibly repressed genes from irreversible silencing. We find that H3K4me3 at bivalent gene promoters—a product of the underlying DNA sequence—persists in nearly all cell types irrespective of gene expression and confers protection from de novo DNA methylation. Accordingly, loss of H3K4me3 at bivalent promoters is strongly associated with aberrant hypermethylation and irreversible silencing in adult human cancers. Bivalency may thus represent a distinct regulatory mechanism for maintaining epigenetic plasticity.HIGHLIGHTSBivalent chromatin does not poise genes for rapid activationH3K4me3 at bivalent promoters is not instructive for transcription activationH3K4me3 at bivalent promoters protects reversibly repressed genes from de novo DNA methylationLoss of H3K4me3/bivalency is associated with aberrant DNA hypermethylation in cancer

scholarly journals 278 Jak/Stat3 SIGNALING PROMOTES SOMATIC CELL REPROGRAMMING BY EPIGENETIC REGULATION

2013 ◽  
Vol 25 (1) ◽  
pp. 287
Author(s):  
Y. Tang ◽  
Y. Luo ◽  
Z. Jiang ◽  
M. Carter ◽  
X. (Cindy) Tian
Keyword(s):  
Stem Cells ◽  
Dna Sequences ◽  
Histone Deacetylases ◽  
Dna Methyltransferase ◽  
De Novo ◽  
Embryonic Stem ◽  
Dna Demethylation ◽  
Janus Kinase ◽  
Cell Reprogramming ◽  
Open Chromatin

Although leukemia inhibitory factor (LIF) maintains the ground state pluripotency of mouse embryonic stem cells and induced pluripotent stem cells (iPSC) by activating the Janus kinase/signal transducer and activator of transcription 3 (Jak/Stat3) pathway, the mechanism remains unclear. Stat3 has only been shown to promote complete reprogramming of epiblast and neural stem cells, and the partially reprogrammed cells (pre-iPSC). We investigated if and how Jak/Stat3 activation promotes reprogramming of terminally differentiated mouse embryonic fibroblasts (MEF). We demonstrated that activated Stat3 not only promotes but is essential for the pluripotency establishment in MEF cell reprogramming. We further demonstrated that, during reprogramming, inhibiting Jak/Stat3 activity blocks demethylation of Oct4 and Nanog regulatory DNA sequences in induced cells, which are marked by suppressed endogenous pluripotent gene expression. These are correlated with significant upregulation of DNA methyltransferase (Dnmt) 1 and histone deacetylases (HDAC) expression, as well as the increased expression of lysine-specific histone demethylase 2 and methyl-CpG binding protein 2. Inhibiting Jak/Stat3 also blocks the expression of Dnmt3L, which is correlated with the failure of retroviral transgene silencing. Furthermore, Dnmt or HDAC inhibitor but not overexpression of Nanog rescues the reprogramming arrested by Jak/Stat3 inhibition or LIF deprivation. Finally, we demonstrated that LIF/Stat3 signal also represents the prerequisite for complete reprogramming of pre-iPSC. We conclude that Jak/Stat3 activity plays a fundamental role in promoting the establishment of pluripotency at the epigenetic level, by facilitating DNA demethylation/de novo methylation and open-chromatin formation during late stage reprogramming.

scholarly journals DNMT3B shapes the mCA landscape and regulates mCG for promoter bivalency in human embryonic stem cells

2019 ◽  
Vol 47 (14) ◽  
pp. 7460-7475 ◽  
Author(s):  
Hong Kee Tan ◽  
Chan-Shuo Wu ◽  
Jia Li ◽  
Zi Hui Tan ◽  
Jordan R Hoffman ◽  
...  
Keyword(s):  
Stem Cells ◽  
Embryonic Stem Cells ◽  
Human Embryonic Stem Cells ◽  
Dna Methyltransferase ◽  
De Novo ◽  
Embryonic Stem ◽  
Distribution Profile ◽  
Pwwp Domain ◽  
The Common ◽  
On Chip

Abstract DNMT3B is known as a de novo DNA methyltransferase. However, its preferential target sites for DNA methylation are largely unknown. Our analysis on ChIP-seq experiment in human embryonic stem cells (hESC) revealed that DNMT3B, mCA and H3K36me3 share the same genomic distribution profile. Deletion of DNMT3B or its histone-interacting domain (PWWP) demolished mCA in hESCs, suggesting that PWWP domain of DNMT3B directs the formation of mCA landscape. In contrast to the common presumption that PWWP guides DNMT3B-mediated mCG deposition, we found that deleting PWWP does not affect the mCG landscape. Nonetheless, DNMT3B knockout led to the formation of 2985 de novo hypomethylated regions at annotated promoter sites. Upon knockout, most of these promoters gain the bivalent marks, H3K4me3 and H3K27me3. We call them spurious bivalent promoters. Gene ontology analysis associated spurious bivalent promoters with development and cell differentiation. Overall, we found the importance of DNMT3B for shaping the mCA landscape and for maintaining the fidelity of the bivalent promoters in hESCs.

scholarly journals Transcriptional regulation of the human DNA methyltransferase 3A and 3B genes by Sp3 and Sp1 zinc finger proteins

2005 ◽  
Vol 385 (2) ◽  
pp. 557-564 ◽  
Author(s):  
Artit JINAWATH ◽  
Satoshi MIYAKE ◽  
Yuka YANAGISAWA ◽  
Yoshimitsu AKIYAMA ◽  
Yasuhito YUASA
Keyword(s):  
Transcriptional Regulation ◽  
Mrna Expression ◽  
Binding Sites ◽  
Dna Methyltransferase ◽  
De Novo ◽  
Fold Increase ◽  
Mrna Levels ◽  
Expression Levels ◽  
Dna Methyltransferase 3A ◽  
Mrna Expression Levels

The DNMT3A (DNA methyltransferase 3A) and DNMT3B genes encode putative de novo methyltransferases and show complex transcriptional regulation in the presence of three and two different promoters respectively. All promoters of DNMT3A and DNMT3B lack typical TATA sequences adjacent to their transcription start sites and contain several Sp1-binding sites. The importance of these Sp1-binding sites was demonstrated by using a GC-rich DNA-binding protein inhibitor, mithramycin A, i.e. on the basis of decrease in the promoter activities and mRNA expression levels of DNMT3A and DNMT3B. Overexpression of Sp1 and Sp3 up-regulated the promoter activities of these two genes. The physical binding of Sp1 and Sp3 to DNMT3A and DNMT3B promoters was confirmed by a gel shift assay. Interestingly, Sp3 overexpression in HEK-293T cells (human embryonic kidney 293T cells) resulted in 3.3- and 4.0-fold increase in DNMT3A and DNMT3B mRNA expression levels respectively by quantitative reverse transcriptase–PCR, whereas Sp1 overexpression did not. Furthermore, an antisense oligonucleotide to Sp3 significantly decreased the mRNA levels of DNMT3A and DNMT3B. These results indicate the functional importance of Sp proteins, particularly Sp3, in the regulation of DNMT3A and DNMT3B gene expression.

scholarly journals Decoding the function of bivalent chromatin in development and cancer

2021 ◽  
pp. gr.275736.121
Author(s):  
Dhirendra Kumar ◽  
Senthilkumar Cinghu ◽  
Andrew J Oldfield ◽  
Pengyi Yang ◽  
Raja Jothi
Keyword(s):  
Dna Methylation ◽  
Regulatory Mechanism ◽  
De Novo ◽  
Embryonic Stem ◽  
Cell Types ◽  
Gene Promoters ◽  
Genome Wide ◽  
Silent Genes ◽  
Aberrant Dna Methylation ◽  
Rapid Activation

Bivalent chromatin is characterized by the simultaneous presence of H3K4me3 and H3K27me3, histone modifications generally associated with transcriptionally active and repressed chromatin, respectively. Prevalent in embryonic stem cells (ESCs), bivalency is postulated to poise/prime lineage-controlling developmental genes for rapid activation during embryogenesis while maintaining a transcriptionally repressed state in the absence of activation cues; however, this hypothesis remains to be directly tested. Most gene promoters DNA-hypermethylated in adult human cancers are bivalently marked in ESCs, and it was speculated that bivalency predisposes them for aberrant de novo DNA methylation and irreversible silencing in cancer, but evidence supporting this model is largely lacking. Here we show that bivalent chromatin does not poise genes for rapid activation but protects promoters from de novo DNA methylation. Genome-wide studies in differentiating ESCs reveal that activation of bivalent genes is no more rapid than that of other transcriptionally silent genes, challenging the premise that H3K4me3 is instructive for transcription. H3K4me3 at bivalent promoters, a product of the underlying DNA sequence, persists in nearly all cell types irrespective of gene expression and confers protection from de novo DNA methylation. Bivalent genes in ESCs that are frequent targets of aberrant hypermethylation in cancer are particularly strongly associated with loss of H3K4me3/bivalency in cancer. Altogether, our findings suggest that bivalency protects reversibly repressed genes from irreversible silencing and that loss of H3K4me3 may make them more susceptible to aberrant DNA methylation in diseases such as cancer. Bivalency may thus represent a distinct regulatory mechanism for maintaining epigenetic plasticity.

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