Similar Datasets
Project description:DNA methyltransferase 3A (DNMT3A) and DNA methyltransferase 3-Like (DNMT3L) form functional heterotetramers and ATRX-DNMT3-DNMT3L (ADD) domains, shared by both DNMT3A and DNMT3L, recognizes histone H3 tail unmethylated at lysine-4 (H3K4me0) to deposit DNA methylation properly in mammalian germ cells. However, the combinational and differential role of ADD domains of DNMT3A and DNMT3L in vivo has not been fully defined. Here we analyze both female and male germ cells derived from mouse with amino-acid substitutions in ADD domains of DNMT3A and/or DNMT3L that impair their domain function. Loss of either DNMT3A-ADD or DNMT3L-ADD domain function shows moderate reduction of global CG methylation level, but in different degree, in both germ cells. However, when both DNMT3A and DNMT3L lost their ADD domain functions, reduction of the global CG methylation level is much more severe and comparable with that of Dnmt3a/3L knockout germ cells. In contrast, such double mutant germ cells have thousands of genomic regions where non-CG methylation is aberrantly accumulated. These results highlight a critical role of the combinational function of ADD domains for the robust CG and non-CG methylation in germ cells.
2024-03-18 | GSE238227 | GEO
Project description:DNA methyltransferase 3A (DNMT3A) and its catalytically inactive cofactor DNA methyltransferase 3-Like (DNMT3L) proteins form functional heterotetramers to deposit DNA methylation in mammalian germ cells. While both proteins have an ATRX-DNMT3-DNMT3L (ADD) domain that recognizes histone H3 tail unmethylated at lysine-4 (H3K4me0), the combined and differential roles of the domains in the two proteins have not been fully defined in vivo. Here we investigate DNA methylation landscapes in female and male germ cells derived from mice with loss-of-function amino acid substitutions in the ADD domains of DNMT3A and/or DNMT3L. Mutations in either the DNMT3A-ADD or the DNMT3L-ADD domain moderately decrease global CG methylation levels, but to different degrees, in both germ cells. Furthermore, when the ADD domains of both DNMT3A and DNMT3L lose their functions, the CG methylation levels are much more reduced, especially in oocytes, comparable to the impact of the Dnmt3a/3L knockout. In contrast, aberrant accumulation of non-CG methylation occurs at thousands of genomic regions in the double mutant oocytes and spermatozoa. These results highlight the critical role of the ADD-H3K4me0 binding in proper CG and non-CG methylation in germ cells and the various impacts of the ADD domains of the two proteins.
| S-EPMC11021467 | biostudies-literature
Project description:DNA methyltransferase 3A (DNMT3A) and DNA methyltransferase 3-Like (DNMT3L) form functional heterotetramers and ATRX-DNMT3-DNMT3L (ADD) domains, shared by both DNMT3A and DNMT3L, recognizes histone H3 tail unmethylated at lysine-4 (H3K4me0) to deposit DNA methylation properly in mammalian germ cells. However, the combinational and differential role of ADD domains of DNMT3A and DNMT3L in vivo has not been fully defined. Here we analyze both female and male germ cells derived from mouse with amino-acid substitutions in ADD domains of DNMT3A and/or DNMT3L that impair their domain function. Loss of either DNMT3A-ADD or DNMT3L-ADD domain function shows moderate reduction of global CG methylation level, but in different degree, in both germ cells. However, when both DNMT3A and DNMT3L lost their ADD domain functions, reduction of the global CG methylation level is much more severe and comparable with that of Dnmt3a/3L knockout germ cells. In contrast, such double mutant germ cells have thousands of genomic regions where non-CG methylation is aberrantly accumulated. These results highlight a critical role of the combinational function of ADD domains for the robust CG and non-CG methylation in germ cells.
2024-03-18 | GSE238226 | GEO
Project description:Combinational effect of the ADD domains of DNMT3A and DNMT3L on DNA methylation landscapes in mouse germ cells
| PRJNA998390 | ENA
Project description:Combinational effect of the ADD domains of DNMT3A and DNMT3L on DNA methylation landscapes in mouse germ cells [WGBS]
| PRJNA998394 | ENA
Project description:Combinational effect of the ADD domains of DNMT3A and DNMT3L on DNA methylation landscapes in mouse germ cells [RNA-seq]
| PRJNA998391 | ENA
Project description:DNA methylation plays a critical role in the regulation and maintenance of cell-type specific transcriptional programs. Targeted epigenome editing is an emerging technology to specifically regulate cellular gene expression in order to modulate cell phenotypes or dissect the epigenetic mechanisms involved in their control. In this work, we employed a DNA methyltransferase Dnmt3a-Dnmt3L construct fused to the nuclease-inactivated dCas9 programmable targeting domain to introduce DNA methylation into the human genome specifically at the EpCAM, CXCR4 and TFRC gene promoters. We show that targeting of these loci with single gRNAs leads to efficient and widespread methylation of the promoters. Multiplexing of several guide RNAs does not increase the efficiency of methylation. Peaks of targeted methylation were observed around 25 bp upstream and 40 bp downstream of the PAM site, while 20-30 bp of the binding site itself are protected against methylation. Potent methylation is dependent on the multimerization of Dnmt3a/Dnmt3L complexes on the DNA. Furthermore, the introduced methylation causes transcriptional repression of the targeted genes. These new programmable epigenetic editors allow unprecedented control of the DNA methylation status in cells and will lead to further advances in the understanding of epigenetic signaling.
| S-EPMC5389507 | biostudies-literature
Project description:Dnmt3L is required for the establishment of maternal methylation imprints at imprinting centers (ICs). Dnmt3L, however, lacks the conserved catalytic domain common to DNA methyltransferases. In an attempt to define its function, we coexpressed DNMT3L with each of the two known de novo methyltransferases, Dnmt3a and DNMT3B, in human cells and monitored de novo methylation by using replicating minichromosomes carrying various ICs as targets. Coexpression of DNMT3L with DNMT3B led to little or no change in target methylation. However, coexpression of DNMT3L with Dnmt3a resulted in a striking stimulation of de novo methylation by Dnmt3a. Stimulation was observed at maternally methylated ICs such as small nuclear ribonucleoprotein polypeptide N (SNRPN), Snrpn, and Igf2rAir, as well as at various nonimprinted sequences present on the episomes. Stimulation of Dnmt3a by DNMT3L was also observed at endogenous sequences in the genome. Therefore, DNMT3L acts as a general stimulatory factor for de novo methylation by Dnmt3a. The implications of these findings for the function of DNMT3L and Dnmt3a in DNA methylation and genomic imprinting are discussed.
| S-EPMC139244 | biostudies-literature
Project description:DNMT3L (DNMT3-like), a member of the DNMT3 family, has no DNA methyltransferase activity but regulates de novo DNA methylation. While biochemical studies show that DNMT3L is capable of interacting with both DNMT3A and DNMT3B and stimulating their enzymatic activities, genetic evidence suggests that DNMT3L is essential for DNMT3A-mediated de novo methylation in germ cells but is dispensable for de novo methylation during embryogenesis, which is mainly mediated by DNMT3B. How DNMT3L regulates DNA methylation and what determines its functional specificity are not well understood. Here we show that DNMT3L-deficient mouse embryonic stem cells (mESCs) exhibit downregulation of DNMT3A, especially DNMT3A2, the predominant DNMT3A isoform in mESCs. DNA methylation analysis of DNMT3L-deficient mESCs reveals hypomethylation at many DNMT3A target regions. These results confirm that DNMT3L is a positive regulator of DNA methylation, contrary to a previous report that, in mESCs, DNMT3L regulates DNA methylation positively or negatively, depending on genomic regions. Mechanistically, DNMT3L forms a complex with DNMT3A2 and prevents DNMT3A2 from being degraded. Restoring the DNMT3A protein level in DNMT3L-deficient mESCs partially recovers DNA methylation. Thus, our work uncovers a role for DNMT3L in maintaining DNMT3A stability, which contributes to the effect of DNMT3L on DNMT3A-dependent DNA methylation.
| S-EPMC6326784 | biostudies-literature
Project description:Genetic imprinting, found in flowering plants and placental mammals, uses DNA methylation to yield gene expression that is dependent on the parent of origin. DNA methyltransferase 3a (Dnmt3a) and its regulatory factor, DNA methyltransferase 3-like protein (Dnmt3L), are both required for the de novo DNA methylation of imprinted genes in mammalian germ cells. Dnmt3L interacts specifically with unmethylated lysine 4 of histone H3 through its amino-terminal PHD (plant homeodomain)-like domain. Here we show, with the use of crystallography, that the carboxy-terminal domain of human Dnmt3L interacts with the catalytic domain of Dnmt3a, demonstrating that Dnmt3L has dual functions of binding the unmethylated histone tail and activating DNA methyltransferase. The complexed C-terminal domains of Dnmt3a and Dnmt3L showed further dimerization through Dnmt3a-Dnmt3a interaction, forming a tetrameric complex with two active sites. Substitution of key non-catalytic residues at the Dnmt3a-Dnmt3L interface or the Dnmt3a-Dnmt3a interface eliminated enzymatic activity. Molecular modelling of a DNA-Dnmt3a dimer indicated that the two active sites are separated by about one DNA helical turn. The C-terminal domain of Dnmt3a oligomerizes on DNA to form a nucleoprotein filament. A periodicity in the activity of Dnmt3a on long DNA revealed a correlation of methylated CpG sites at distances of eight to ten base pairs, indicating that oligomerization leads Dnmt3a to methylate DNA in a periodic pattern. A similar periodicity is observed for the frequency of CpG sites in the differentially methylated regions of 12 maternally imprinted mouse genes. These results suggest a basis for the recognition and methylation of differentially methylated regions in imprinted genes, involving the detection of both nucleosome modification and CpG spacing.
| S-EPMC2712830 | biostudies-literature