Identification of transcription termination defects at DNA hypomethylated transcription termination sites in DNA methyltransferase 3a-deficient vertebrates
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ABSTRACT:
ORGANISM(S): Danio rerio
PROVIDER: GSE178691 | GEO | 2022/06/14
REPOSITORIES: GEO
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Project description:CpG methylation in genomic DNA is well known as a repressive epigenetic marker in eukaryotic transcription, and hypermethylation of the promoter region is correlated with silencing of gene expression. In contrast to the promoter region, the function of DNA methylation at transcription termination remains to be elucidated. A recent study has revealed that mouse DNA methyltransferase 3a (Dnmt3a) mainly functions in de novo methylation in the promoter and gene body regions (including transcription termination sites (TTSs)) during development. To investigate the relationship between DNA methylation overlapping the TTSs and transcription termination, we employed two strategies: informatic analysis using already deposited datasets of Dnmt3a-/- mouse cells and the zebrafish model system. Bioinformatic analysis using methylome and transcriptome data showed that hypomethylated differentially methylated regions overlapping the TTSs were associated with increased transcript counts and chimeric transcripts downstream of TTSs in Dnmt3a-/- Agouti-related protein neurons, but not in Dnmt3a-/- ES cells and MEFs. We experimentally detected increased read-through and chimeric transcripts downstream of hypomethylated TTSs in zebrafish maternal-zygotic Dnmt3aa-/- mutants. This study is the first to identify transcription termination defects in DNA hypomethylated TTSs in Dnmt3a-/- vertebrates.
2022-06-14 | GSE178690 | GEO
Project description:CpG methylation in genomic DNA is well known as a repressive epigenetic marker in eukaryotic transcription, and hypermethylation of the promoter region is correlated with silencing of gene expression. In contrast to the promoter region, the function of DNA methylation at transcription termination remains to be elucidated. A recent study has revealed that mouse DNA methyltransferase 3a (Dnmt3a) mainly functions in de novo methylation in the promoter and gene body regions (including transcription termination sites (TTSs)) during development. To investigate the relationship between DNA methylation overlapping the TTSs and transcription termination, we employed two strategies: informatic analysis using already deposited datasets of Dnmt3a-/- mouse cells and the zebrafish model system. Bioinformatic analysis using methylome and transcriptome data showed that hypomethylated differentially methylated regions overlapping the TTSs were associated with increased transcript counts and chimeric transcripts downstream of TTSs in Dnmt3a-/- Agouti-related protein neurons, but not in Dnmt3a-/- ES cells and MEFs. We experimentally detected increased read-through and chimeric transcripts downstream of hypomethylated TTSs in zebrafish maternal-zygotic Dnmt3aa-/- mutants. This study is the first to identify transcription termination defects in DNA hypomethylated TTSs in Dnmt3a-/- vertebrates.
2022-06-14 | GSE178688 | GEO
Project description:Identification of transcription termination defects at DNA hypomethylated transcription termination sites in DNA methyltransferase 3a-deficient vertebrates
| PRJNA740013 | ENA
Project description:Identification of transcription termination defects at DNA hypomethylated transcription termination sites in DNA methyltransferase 3a-deficient vertebrates [WGBS]
| PRJNA740018 | ENA
Project description:Identification of transcription termination defects at DNA hypomethylated transcription termination sites in DNA methyltransferase 3a-deficient vertebrates [RNA-Seq]
| PRJNA740017 | ENA
Project description:BackgroundThermal plasticity is pivotal for evolution in changing climates and in mediating resilience to its potentially negative effects. The efficacy to respond to environmental change depends on underlying mechanisms. DNA methylation induced by DNA methyltransferase 3 enzymes in the germline or during early embryonic development may be correlated with responses to environmental change. This developmental plasticity can interact with reversible acclimation within adult organisms, which would increase the speed of response and could alleviate potential mismatches between parental or early embryonic environments and those experienced at later life stages. Our aim was to determine whether there is a causative relationship between DNMT3 enzyme and developmental thermal plasticity and whether either or both interact with short-term acclimation to alter fitness and thermal responses in zebrafish (Danio rerio).ResultsWe developed a novel DNMT3a knock-out model to show that sequential knock-out of DNA methyltransferase 3a isoforms (DNMT3aa-/- and DNMT3aa-/-ab-/-) additively decreased survival and increased deformities when cold developmental temperatures in zebrafish offspring mismatched warm temperatures experienced by parents. Interestingly, short-term cold acclimation of parents before breeding rescued DNMT3a knock-out offspring by restoring survival at cold temperatures. DNMT3a knock-out genotype interacted with developmental temperatures to modify thermal performance curves in offspring, where at least one DNMT3a isoform was necessary to buffer locomotion from increasing temperatures. The thermal sensitivity of citrate synthase activity, an indicator of mitochondrial density, was less severely affected by DNMT3a knock-out, but there was nonetheless a significant interaction between genotype and developmental temperatures.ConclusionsOur results show that DNMT3a regulates developmental thermal plasticity and that the phenotypic effects of different DNMT3a isoforms are additive. However, DNMT3a interacts with other mechanisms, such as histone (de)acetylation, induced during short-term acclimation to buffer phenotypes from environmental change. Interactions between these mechanisms make phenotypic compensation for climate change more efficient and make it less likely that thermal plasticity incurs a cost resulting from environmental mismatches.
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Project description:DNA methyltransferase 3A (DNMT3A) is one of two human de novo DNA methyltransferases essential for transcription regulation during cellular development and differentiation. There is increasing evidence that RNA plays a role in directing DNA methylation to specific genomic locations within mammalian cells. Here, we describe two modes of RNA regulation of DNMT3A in vitro. We show a single-stranded RNA molecule that is antisense to the E-cadherin promoter binds tightly to the catalytic domain in a structurally dependent fashion causing potent inhibition of DNMT3A activity. Two other RNA molecules bind DNMT3A at an allosteric site outside the catalytic domain, causing no change in catalysis. Our observation of the potent and specific in vitro modulation of DNMT3A activity by RNA supports in vivo data that RNA interacts with DNMT3A to regulate transcription.
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Project description:Gle1 is a nucleocytoplasmic shuttling protein with well-documented cytoplasmic roles as a modulator of ATP-dependent DEAD-box RNA helicases involved in messenger (m)RNA export, translation initiation and termination, and stress granule dynamics. Here, we identify a novel nuclear role for Gle1 during transcription termination. In HeLa cells treated with a peptide that disrupts Gle1 nucleocytoplasmic shuttling, we detected nuclear accumulation of specific mRNAs with elongated 3'-UTR (untranslated region). Enriched mRNAs were nascently transcribed and accumulated in the nucleus due to a change in transcription state and not due to altered nuclear export. Whereas Gle1 shuttling inhibition did not appear to perturb nuclear DDX19 functions, it did result in increased DDX1 nucleoplasmic localization and decreased DDX1 interactions with Gle1 and the pre-mRNA cleavage stimulation factor CstF-64. An increase in nuclear R-loop signal intensity was also observed with diminished Gle1 shuttling, as well as colocalization of Gle1 at R-loops. Taken together, these studies reveal a nuclear role for Gle1 in coordinating DDX1 function in transcription termination complexes.
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Project description:The DNA methyltransferases DNMT3A and DNMT3B are primarily responsible for de novo methylation of specific cytosine residues in CpG dinucleotides during mammalian development. While loss-of-function mutations in DNMT3A are highly recurrent in acute myeloid leukemia (AML), DNMT3A mutations are almost never found in AML patients with translocations that create oncogenic fusion genes such as PML-RARA, RUNX1-RUNX1T1, and MLL-AF9. Here, we explored how DNMT3A is involved in the function of these fusion genes. We used retroviral vectors to express PML-RARA, RUNX1-RUNX1T1, or MLL-AF9 in bone marrow cells derived from WT or DNMT3A-deficient mice. Additionally, we examined the phenotypes of hematopoietic cells from Ctsg-PML-RARA mice, which express PML-RARA in early hematopoietic progenitors and myeloid precursors, with or without DNMT3A. We determined that the methyltransferase activity of DNMT3A, but not DNMT3B, is required for aberrant PML-RARA-driven self-renewal ex vivo and that DNMT3A is dispensable for RUNX1-RUNX1T1- and MLL-AF9-driven self-renewal. Furthermore, both the PML-RARA-driven competitive transplantation advantage and development of acute promyelocytic leukemia (APL) required DNMT3A. Together, these findings suggest that PML-RARA requires DNMT3A to initiate APL in mice.
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Project description:Daily life stress markedly affects the response toward stressful stimuli. DNA methy-lation is one of the factors that regulate this response, and is a normal mechanism of somatic cell growth, but its regulatory gene variations may cause alterations in the stress response. The aim of the present study was to investigate genotypic variants of the DNA methyltransferase 3A (DNMT3A) gene in 129 healthy subjects and evaluate its association with daily life stress. Blood samples were collected, and genomic DNA was isolated. DNA was amplified using specific tetra primers for DNMT3A (C/T) rs11683424 and visualized following 2% agarose gel electrophoresis. The association of DNMT3A genetic variants with daily life stress was analyzed using the Kessler Psychological Distress Scale (K10). We observed that the distribution of subjects with genotype CC (wild type), CT (heteromutant), and TT (homomutant) was 13.95%, 81.4%, and 4.65%, respectively. Genetic variations significantly affected the daily life stress condition (p=0.04) in Indonesian healthy subjects, but most of the subjects with the CT phenotype were classified in a stress condition.
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