Non-germ Line Restoration of Genomic Imprinting for a Small Subset of Imprinted Genes in Ubiquitin-like PHD and RING Finger Domain-Containing 1 (Uhrf1) Null Mouse Embryonic Stem Cells

Qi, Shankang; Wang, Zhiqiang; Li, Pishun; Wu, Qihan; Shi, Tieliu; Li, Jiwen; Wong, Jiemin

doi:10.1074/jbc.m114.626697

Cited by 5 publications

(6 citation statements)

References 46 publications

(52 reference statements)

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“…During the course of writing, a paper from the Wong group investigating the behaviour of UHRF1 rescue cells found that a number of imprinted genes showed gains in methylation in that system too [55]. Methylation gain was only seen at some of the imprinted loci, and there was no clear link to the location of the gDMR, the presence of antisense transcripts or the type of imprint.…”

Section: Discussionmentioning

confidence: 99%

Widespread recovery of methylation at gametic imprints in hypomethylated mouse stem cells following rescue with DNMT3A2

Thakur

Mackin

Irwin

et al. 2016

Epigenetics & Chromatin

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BackgroundImprinted loci are paradigms of epigenetic regulation and are associated with a number of genetic disorders in human. A key characteristic of imprints is the presence of a gametic differentially methylated region (gDMR). Previous studies have indicated that DNA methylation lost from gDMRs could not be restored by DNMT1, or the de novo enzymes DNMT3A or 3B in stem cells, indicating that imprinted regions must instead undergo passage through the germline for reprogramming. However, previous studies were non-quantitative, were unclear on the requirement for DNMT3A/B and showed some inconsistencies. In addition, new putative gDMR has recently been described, along with an improved delineation of the existing gDMR locations. We therefore aimed to re-examine the dependence of methylation at gDMRs on the activities of the methyltransferases in mouse embryonic stem cells (ESCs).ResultsWe examined the most complete current set of imprinted gDMRs that could be assessed using quantitative pyrosequencing assays in two types of ESCs: those lacking DNMT1 (1KO) and cells lacking a combination of DNMT3A and DNMT3B (3abKO). We further verified results using clonal analysis and combined bisulfite and restriction analysis. Our results showed that loss of methylation was approximately equivalent in both cell types. 1KO cells rescued with a cDNA-expressing DNMT1 could not restore methylation at the imprinted gDMRs, confirming some previous observations. However, nearly all gDMRs were remethylated in 3abKO cells rescued with a DNMT3A2 expression construct (3abKO + 3a2). Transcriptional activity at the H19/Igf2 locus also tracked with the methylation pattern, confirming functional reprogramming in the latter.ConclusionsThese results suggested (1) a vital role for DNMT3A/B in methylation maintenance at imprints, (2) that loss of DNMT1 and DNMT3A/B had equivalent effects, (3) that rescue with DNMT3A2 can restore imprints in these cells. This may provide a useful system in which to explore factors influencing imprint reprogramming.Electronic supplementary materialThe online version of this article (doi:10.1186/s13072-016-0104-2) contains supplementary material, which is available to authorized users.

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Section: Discussionmentioning

confidence: 99%

Widespread recovery of methylation at gametic imprints in hypomethylated mouse stem cells following rescue with DNMT3A2

Thakur

Mackin

Irwin

et al. 2016

Epigenetics & Chromatin

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“…UHRF1 specifically binds to hemimethylated DNA via its SRA domain [120][121][122][123] and recognizes the N-terminal tails of histone H3 di-and tri-methylated at lysine 9 (H3K9me2/me3) via combined binding of its tandem Tudor domain (TTD) and its plant homeodomain (PHD) [124][125][126][127] . These specific chromatin interactions of UHRF1 are necessary for the recruitment of DNMT1 to replicating chromatin and DNA methylation maintenance, since mutations preventing histone binding in any of the domains abolished DNA methylation by DNMT1 in cells 125,127,128 . In addition to its role in targeting of DNMT1, UHRF1 was also shown to stimulate the catalytic activity of DNMT1 through direct interaction 129,130 .…”

Section: Recruitment Of Dnmt1 To Replicating Chromatinmentioning

confidence: 99%

Target specificity of mammalian DNA methylation and demethylation machinery

2018

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DNA methylation is an essential epigenetic modification for mammalian embryonic development and biology. The DNA methylation pattern across the genome, together with other epigenetic signals, is responsible for the transcriptional profile of a cell and thus preservation of the cell's identity. Equally, the family of TET enzymes which triggers the initiation of the DNA demethylation cycle plays a vital role in the early embryonic development and a lack of these enzymes at later stages leads to a diseased state and dysregulation of the epigenome. DNA methylation has long been considered a very stable modification; however, it has become increasingly clear that for the establishment and maintenance of the methylation pattern, both generation of DNA methylation and its removal are important, and that a delicate balance of ongoing DNA methylation and demethylation shapes the final epigenetic methylation pattern of the cell. Although this epigenetic mark has been investigated in great detail, it still remains to be fully understood how specific DNA methylation imprints are precisely generated, maintained, read or erased in the genome. Here, we provide a biochemist's view on how both DNA methyltransferases and TET enzymes are recruited to specific genomic loci, and how their chromatin interactions, as well as their intrinsic sequence specificities and molecular mechanisms, contribute to the methylation pattern of the cell.

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“…Despite intense work in the DNA methylation field, its role in regulating differentially expressed genes is controversial (Gutierrez-Arcelus et al, 2013). Many studies in organisms across the evolutionary spectrum have demonstrated that DNA methylation does not serve as a universal mechanism of repressing gene expression (Feng et al, 2010;Grow et al, 2015;Gutierrez-Arcelus et al, 2013;Jackson-Grusby et al, 2001;Jacob et al, 2015;Potok et al, 2013;Qi et al, 2015;Zemach et al, 2010;Zhang et al, 2016). In fact, most data indicate that relatively few genes are induced when methylation is lost: there is a lack of global gene induction in mammalian cells deficient in the proteins required for DNA methylation, namely ubiquitin-like with PHD and ring finger domains 1 (Uhrf1) (Qi et al, 2015) or DNA methyltransferase 1 (DNMT1) (Jackson-Grusby et al, 2001), or during the stage of early embryogenesis when the parental methylome is nearly completely erased (Grow et al, 2015;Potok et al, 2013).…”

Section: Introductionmentioning

confidence: 99%

Loss of DNA methylation in zebrafish embryos activates retrotransposons to trigger antiviral signaling

et al. 2017

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Complex cytoplasmic nucleotide-sensing mechanisms can recognize foreign DNA based on a lack of methylation and initiate an immune response to clear the infection. Zebrafish embryos with global DNA hypomethylation caused by mutations in the () or () genes exhibit a robust interferon induction characteristic of the first line of defense against viral infection. We found that this interferon induction occurred in non-immune cells and examined whether intracellular viral sensing pathways in these cells were the trigger. RNA-seq analysis of and mutants revealed widespread induction of Class I retrotransposons and activation of cytoplasmic DNA viral sensors. Attenuating Sting, phosphorylated Tbk1 and, importantly, blocking reverse transcriptase activity suppressed the expression of interferon genes in mutants. Thus, activation of transposons in cells with global DNA hypomethylation mimics a viral infection by activating cytoplasmic DNA sensors. This suggests that antiviral pathways serve as surveillance of cells that have derepressed intragenomic parasites due to DNA hypomethylation.

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Non-germ Line Restoration of Genomic Imprinting for a Small Subset of Imprinted Genes in Ubiquitin-like PHD and RING Finger Domain-Containing 1 (Uhrf1) Null Mouse Embryonic Stem Cells

Abstract: Background: Once erased, DNA methylation in imprinted genes was shown previously to be re-established only through germ line passage. Results: UHRF1 re-expression in Uhrf1Ϫ/Ϫ mouse embryonic stem (ES) cells restores DNA methylation for a few imprinted

Cited by 5 publications

References 46 publications

Widespread recovery of methylation at gametic imprints in hypomethylated mouse stem cells following rescue with DNMT3A2

Widespread recovery of methylation at gametic imprints in hypomethylated mouse stem cells following rescue with DNMT3A2

Target specificity of mammalian DNA methylation and demethylation machinery

Loss of DNA methylation in zebrafish embryos activates retrotransposons to trigger antiviral signaling

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