Your browser doesn't support javascript.
loading
Show: 20 | 50 | 100
Results 1 - 20 de 363
Filter
1.
Science ; 385(6705): eadl6173, 2024 Jul 12.
Article in English | MEDLINE | ID: mdl-38991060

ABSTRACT

Isocitrate dehydrogenase 1 (IDH1) is the most commonly mutated metabolic gene across human cancers. Mutant IDH1 (mIDH1) generates the oncometabolite (R)-2-hydroxyglutarate, disrupting enzymes involved in epigenetics and other processes. A hallmark of IDH1-mutant solid tumors is T cell exclusion, whereas mIDH1 inhibition in preclinical models restores antitumor immunity. Here, we define a cell-autonomous mechanism of mIDH1-driven immune evasion. IDH1-mutant solid tumors show selective hypermethylation and silencing of the cytoplasmic double-stranded DNA (dsDNA) sensor CGAS, compromising innate immune signaling. mIDH1 inhibition restores DNA demethylation, derepressing CGAS and transposable element (TE) subclasses. dsDNA produced by TE-reverse transcriptase (TE-RT) activates cGAS, triggering viral mimicry and stimulating antitumor immunity. In summary, we demonstrate that mIDH1 epigenetically suppresses innate immunity and link endogenous RT activity to the mechanism of action of a US Food and Drug Administration-approved oncology drug.


Subject(s)
Immune Evasion , Immunity, Innate , Isocitrate Dehydrogenase , Neoplasms , Animals , Humans , Mice , Cell Line, Tumor , DNA/metabolism , DNA Demethylation , DNA Methylation , DNA Transposable Elements , Epigenesis, Genetic , Glutarates/metabolism , Immunity, Innate/genetics , Isocitrate Dehydrogenase/genetics , Isocitrate Dehydrogenase/metabolism , Mutation , Neoplasms/immunology , Neoplasms/genetics , Nucleotidyltransferases/genetics , Tumor Escape , Immune Evasion/genetics
2.
Int J Mol Sci ; 25(11)2024 May 28.
Article in English | MEDLINE | ID: mdl-38892059

ABSTRACT

Global methylation levels differ in in vitro- and in vivo-developed embryos. Follicular fluid (FF) contains extracellular vesicles (EVs) containing miRNAs that affect embryonic development. Here, we examined our hypothesis that components in FF affect global DNA methylation and embryonic development. Oocytes and FF were collected from bovine ovaries. Treatment of zygotes with a low concentration of FF induced global DNA demethylation, improved embryonic development, and reduced DNMT1/3A levels. We show that embryos take up EVs containing labeled miRNA secreted from granulosa cells and the treatment of zygotes with EVs derived from FF reduces global DNA methylation in embryos. Furthermore, the methylation levels of in vitro-developed blastocysts were higher than those of in their vivo counterparts. Based on small RNA-sequencing and in silico analysis, we predicted miR-29b, -199a-3p, and -148a to target DNMTs and to induce DNA demethylation, thereby improving embryonic development. Moreover, among FF from 30 cows, FF with a high content of these miRNAs demethylated more DNA in the embryos than FF with a lower miRNA content. Thus, miRNAs in FF play a role in early embryonic development.


Subject(s)
Embryonic Development , Extracellular Vesicles , Follicular Fluid , MicroRNAs , Animals , Female , MicroRNAs/genetics , MicroRNAs/metabolism , Cattle , Follicular Fluid/metabolism , Extracellular Vesicles/metabolism , Embryonic Development/genetics , DNA Methylation , DNA Demethylation , Oocytes/metabolism , Blastocyst/metabolism , Embryo, Mammalian/metabolism , Gene Expression Regulation, Developmental , Zygote/metabolism
3.
Mol Biol Rep ; 51(1): 778, 2024 Jun 21.
Article in English | MEDLINE | ID: mdl-38904842

ABSTRACT

BACKGROUND: SETDB1 (SET domain bifurcated-1) is a histone H3-lysine 9 (H3K9)-specific methyltransferase that mediates heterochromatin formation and repression of target genes. Despite the assumed functional link between DNA methylation and SETDB1-mediated H3K9 trimethylations, several studies have shown that SETDB1 operates autonomously of DNA methylation in a region- and cell-specific manner. This study analyzes SETDB1-null HAP1 cells through a linked methylome and transcriptome analysis, intending to explore genes controlled by SETDB1-involved DNA methylation. METHODS AND RESULTS: We investigated SETDB1-mediated regulation of DNA methylation and gene transcription in human HAP1 cells using reduced-representation bisulfite sequencing (RRBS) and RNA sequencing. While two-thirds of differentially methylated CpGs (DMCs) in genic regions were hypomethylated in SETDB1-null cells, we detected a plethora of C2H2-type zinc-finger protein genes (C2H2-ZFP, 223 of 749) among the DMC-associated genes. Most C2H2-ZFPs with DMCs in their promoters were found hypomethylated in SETDB1-KO cells, while other non-ZFP genes with promoter DMCs were not. These C2H2-ZFPs with DMCs in their promoters were significantly upregulated in SETDB1-KO cells. Similarly, C2H2-ZFP genes were upregulated in SETDB1-null 293T cells, suggesting that SETDB1's function in ZFP gene repression is widespread. There are several C2H2-ZFP gene clusters on chromosome 19, which were selectively hypomethylated in SETDB1-KO cells. CONCLUSIONS: SETDB1 collectively and specifically represses a substantial fraction of the C2H2-ZFP gene family. Through the en-bloc silencing of a set of ZFP genes, SETDB1 may help establish a panel of ZFP proteins that are expressed cell-type specifically and thereby can serve as signature proteins for cellular identity.


Subject(s)
DNA Methylation , Histone-Lysine N-Methyltransferase , Zinc Fingers , Histone-Lysine N-Methyltransferase/genetics , Histone-Lysine N-Methyltransferase/metabolism , Humans , Zinc Fingers/genetics , DNA Methylation/genetics , Promoter Regions, Genetic/genetics , Up-Regulation/genetics , DNA Demethylation , Cell Line , CpG Islands/genetics , Gene Deletion , Histones/metabolism , Histones/genetics
4.
Nat Commun ; 15(1): 3734, 2024 May 03.
Article in English | MEDLINE | ID: mdl-38702312

ABSTRACT

Mutations in DNA damage response (DDR) factors are associated with human infertility, which affects up to 15% of the population. The DDR is required during germ cell development and meiosis. One pathway implicated in human fertility is DNA translesion synthesis (TLS), which allows replication impediments to be bypassed. We find that TLS is essential for pre-meiotic germ cell development in the embryo. Loss of the central TLS component, REV1, significantly inhibits the induction of human PGC-like cells (hPGCLCs). This is recapitulated in mice, where deficiencies in TLS initiation (Rev1-/- or PcnaK164R/K164R) or extension (Rev7 -/-) result in a > 150-fold reduction in the number of primordial germ cells (PGCs) and complete sterility. In contrast, the absence of TLS does not impact the growth, function, or homeostasis of somatic tissues. Surprisingly, we find a complete failure in both activation of the germ cell transcriptional program and in DNA demethylation, a critical step in germline epigenetic reprogramming. Our findings show that for normal fertility, DNA repair is required not only for meiotic recombination but for progression through the earliest stages of germ cell development in mammals.


Subject(s)
DNA Demethylation , DNA Repair , DNA-Directed DNA Polymerase , Germ Cells , Animals , Female , Humans , Male , Mice , DNA Damage , DNA-Directed DNA Polymerase/metabolism , DNA-Directed DNA Polymerase/genetics , Epigenesis, Genetic , Germ Cells/metabolism , Meiosis/genetics , Mice, Knockout , Nucleotidyltransferases/metabolism , Nucleotidyltransferases/genetics , Proliferating Cell Nuclear Antigen/metabolism , Translesion DNA Synthesis
5.
Proc Natl Acad Sci U S A ; 121(22): e2320468121, 2024 May 28.
Article in English | MEDLINE | ID: mdl-38768356

ABSTRACT

Spontaneous gain or loss of DNA methylation occurs in plant and animal genomes, and DNA methylation changes can lead to meiotically stable epialleles that generate heritable phenotypic diversity. However, it is unclear whether transgenerational epigenetic stability may be regulated by any cellular factors. Here, we examined spontaneously occurring variations in DNA methylation in wild-type and ros1 mutant Arabidopsis plants that were propagated for ten generations from single-seed descent. We found that the ros1 mutant, which is defective in active DNA demethylation, showed an increased transgenerational epimutation rate. The ros1 mutation led to more spontaneously gained methylation than lost methylation at individual cytosines, compared to the wild type which had similar numbers of spontaneously gained and lost methylation cytosines. Consistently, transgenerational differentially methylated regions were also biased toward hypermethylation in the ros1 mutant. Our results reveal a genetic contribution of the ROS1 DNA demethylase to transgenerational epigenetic stability and suggest that ROS1 may have an unexpected surveillance function in preventing transgenerational DNA methylation increases.


Subject(s)
Arabidopsis Proteins , Arabidopsis , DNA Demethylation , DNA Methylation , Epigenesis, Genetic , Mutation , Arabidopsis/genetics , Arabidopsis/metabolism , Arabidopsis Proteins/genetics , Arabidopsis Proteins/metabolism , Gene Expression Regulation, Plant , DNA, Plant/genetics , DNA, Plant/metabolism , Nuclear Proteins
6.
BMC Genomics ; 25(1): 344, 2024 Apr 05.
Article in English | MEDLINE | ID: mdl-38580899

ABSTRACT

BACKGROUND: Genome-wide DNA demethylation occurs in mammalian primordial germ cells (PGCs) as part of the epigenetic reprogramming important for gametogenesis and resetting the epigenetic information for totipotency. Dppa3 (also known as Stella or Pgc7) is highly expressed in mouse PGCs and oocytes and encodes a factor essential for female fertility. It prevents excessive DNA methylation in oocytes and ensures proper gene expression in preimplantation embryos: however, its role in PGCs is largely unexplored. In the present study, we investigated whether or not DPPA3 has an impact on CG methylation/demethylation in mouse PGCs. RESULTS: We show that DPPA3 plays a role in genome-wide demethylation in PGCs even before sex differentiation. Dppa3 knockout female PGCs show aberrant hypermethylation, most predominantly at H3K9me3-marked retrotransposons, which persists up to the fully-grown oocyte stage. DPPA3 works downstream of PRDM14, a master regulator of epigenetic reprogramming in embryonic stem cells and PGCs, and independently of TET1, an enzyme that hydroxylates 5-methylcytosine. CONCLUSIONS: The results suggest that DPPA3 facilitates DNA demethylation through a replication-coupled passive mechanism in PGCs. Our study identifies DPPA3 as a novel epigenetic reprogramming factor in mouse PGCs.


Subject(s)
Chromosomal Proteins, Non-Histone , DNA Demethylation , Epigenesis, Genetic , Animals , Female , Mice , Chromosomal Proteins, Non-Histone/metabolism , DNA Methylation , Genome , Germ Cells/metabolism , Mammals/genetics
7.
Sci Rep ; 14(1): 6481, 2024 03 18.
Article in English | MEDLINE | ID: mdl-38499584

ABSTRACT

The active DNA demethylation process, which involves TET proteins, can affect DNA methylation pattern. TET dependent demethylation results in DNA hypomethylation by oxidation 5-methylcytosine (5-mC) to 5-hydroxymethylcytosine (5-hmC) and its derivatives. Moreover, TETs' activity may be upregulated by ascorbate. Given that aberrant DNA methylation of genes implicated in breast carcinogenesis may be involved in tumor progression, we wanted to determine whether breast cancer patients exert changes in the active DNA demethylation process. The study included blood samples from breast cancer patients (n = 74) and healthy subjects (n = 71). We analyzed the expression of genes involved in the active demethylation process (qRT-PCR), and 5-mC and its derivatives level (2D-UPLC MS/MS). The ascorbate level was determined using UPLC-MS. Breast cancer patients had significantly higher TET3 expression level, lower 5-mC and 5-hmC DNA levels. TET3 was significantly increased in luminal B breast cancer patients with expression of hormone receptors. Moreover, the ascorbate level in the plasma of breast cancer patients was decreased with the accompanying increase of sodium-dependent vitamin C transporters (SLC23A1 and SLC23A2). The presented study indicates the role of TET3 in DNA demethylation in breast carcinogenesis.


Subject(s)
Breast Neoplasms , Dioxygenases , Humans , Female , DNA Demethylation , Breast Neoplasms/genetics , Chromatography, Liquid , Tandem Mass Spectrometry , 5-Methylcytosine/metabolism , DNA Methylation , Biomarkers/metabolism , DNA/metabolism , Epigenesis, Genetic , Leukocytes/metabolism , Carcinogenesis/genetics , Dioxygenases/genetics
8.
Sci Rep ; 14(1): 6155, 2024 03 14.
Article in English | MEDLINE | ID: mdl-38486042

ABSTRACT

As the most prevalent epitranscriptomic modification, N6-methyladenosine (m6A) shows important roles in a variety of diseases through regulating the processing, stability and translation of target RNAs. However, the potential contributions of m6A to RNA functions are unclear. Here, we identified a functional and prognosis-related m6A-modified RNA SREBF2-AS1 in hepatocellular carcinoma (HCC). The expression of SREBF2-AS1 and SREBF2 in HCC tissues and cells was measured by RT-qPCR. m6A modification level of SREBF2-AS1 was measured by methylated RNA immunoprecipitation assay. The roles of SREBF2-AS1 in HCC progression and sorafenib resistance were investigated by proliferation, apoptosis, migration, and cell viability assays. The regulatory mechanisms of SREBF2-AS1 on SREBF2 were investigated by Chromatin isolation by RNA purification, RNA immunoprecipitation, CUT&RUN, and bisulfite DNA sequencing assays. Our findings showed that the expression of SREBF2-AS1 was increased in HCC tissues and cells, and positively correlated with poor survival of HCC patients. m6A modification level of SREBF2-AS1 was also increased in HCC and positively correlated with poor prognosis of HCC patients. METTL3 and METTL14-induced m6A modification upregulated SREBF2-AS1 expression through increasing SREBF2-AS1 transcript stability. Functional assays showed that only m6A-modified, but not non-modified SREBF2-AS1 promoted HCC progression and sorafenib resistance. Mechanistic investigations revealed that m6A-modified SREBF2-AS1 bound and recruited m6A reader FXR1 and DNA 5-methylcytosine dioxygenase TET1 to SREBF2 promoter, leading to DNA demethylation at SREBF2 promoter and the upregulation of SREBF2 transcription. Functional rescue assays showed that SREBF2 was the critical mediator of the oncogenic roles of SREBF2-AS1 in HCC. Together, this study showed that m6A-modified SREBF2-AS1 exerted oncogenic roles in HCC through inducing DNA demethylation and transcriptional activation of SREBF2, and suggested m6A-modified SREBF2-AS1 as a prognostic biomarker and therapeutic target for HCC.


Subject(s)
Adenosine/analogs & derivatives , Carcinoma, Hepatocellular , Liver Neoplasms , MicroRNAs , Sterol Regulatory Element Binding Protein 2 , Humans , Carcinoma, Hepatocellular/drug therapy , Carcinoma, Hepatocellular/genetics , Carcinoma, Hepatocellular/metabolism , Sorafenib/pharmacology , Liver Neoplasms/drug therapy , Liver Neoplasms/genetics , Liver Neoplasms/metabolism , DNA Demethylation , Cell Line, Tumor , MicroRNAs/genetics , RNA-Binding Proteins/metabolism , Mixed Function Oxygenases/metabolism , Proto-Oncogene Proteins/metabolism , Methyltransferases/genetics , Methyltransferases/metabolism
9.
Sci Rep ; 14(1): 2683, 2024 02 01.
Article in English | MEDLINE | ID: mdl-38302503

ABSTRACT

TROP2 is a powerful cancer driver in colorectal cancer cells. Divergent epigenetic regulation mechanisms for the corresponding TACSTD2 gene exist such as miRNAs or DNA methylation. However, the role of TACSTD2 promoter hypermethylation in colorectal cancer has not been investigated yet. In this study, TROP2 expression strongly correlated with promoter methylation in different colorectal tumor cell lines. Treatment with 5-Azacytidine, a DNMT1 inhibitor, led to demethylation of the TACSTD2 promoter accompanied by an increase in TROP2 protein expression. TROP2 expression correlated with promoter methylation in vivo in human colon tumor tissue, thereby verifying promoter methylation as an important factor in the regulation of TROP2 expression in colorectal cancer. When performing a ChIP-Seq analysis in HCT116 and HT29 cells, we found that TACSTD2 promoter demethylation was accompanied by tri-methylation of H3K4. In silico analysis of GSE156613 data set confirmed that a higher binding of histone mark H3K4me3 around the TACSTD2 promoter was found in TACSTD2 high expressing tumors of colon cancer patients compared to the corresponding adjacent tumor tissue. Moreover, the link between TROP2 and the H3K4me3 code was even evident in tumors showing high intratumoral heterogeneity for TROP2 staining. Our data provide novel evidence for promoter demethylation and simultaneous gains of the active histone mark H3K4me3 across CpG-rich sequences, both being complementary mechanisms in the transcriptional regulation of TACSTD2 in colon cancer. The functional consequences of TROP2 loss in colorectal cancer needs to be further investigated.


Subject(s)
Colonic Neoplasms , Colorectal Neoplasms , Humans , Epigenesis, Genetic , DNA Demethylation , DNA Methylation , Cell Line, Tumor , Colonic Neoplasms/genetics , Gene Expression Regulation, Neoplastic , Colorectal Neoplasms/pathology , CpG Islands , Antigens, Neoplasm/genetics , Antigens, Neoplasm/metabolism , Cell Adhesion Molecules/genetics , Cell Adhesion Molecules/metabolism
10.
Nucleic Acids Res ; 52(7): 3886-3895, 2024 Apr 24.
Article in English | MEDLINE | ID: mdl-38324471

ABSTRACT

The eukaryotic epigenetic modifications 5-methyldeoxycytosine (5mC) and N6-methyldeoxyadenine (6mA) have indispensable regulatory roles in gene expression and embryonic development. We recently identified an atypical bifunctional dioxygenase CcTet from Coprinopsis cinerea that works on both 5mC and 6mA demethylation. The nonconserved residues Gly331 and Asp337 of CcTet facilitate 6mA accommodation, while D337F unexpectedly abolishes 5mC oxidation activity without interfering 6mA demethylation, indicating a prominent distinct but unclear 5mC oxidation mechanism to the conventional Tet enzymes. Here, we assessed the molecular mechanism of CcTet in catalyzing 5mC oxidation by representing the crystal structure of CcTet-5mC-dsDNA complex. We identified the distinct mechanism by which CcTet recognizes 5mC-dsDNA compared to 6mA-dsDNA substrate. Moreover, Asp337 was found to have a central role in compensating for the loss of a critical 5mC-stablizing H-bond observed in conventional Tet enzymes, and stabilizes 5mC and subsequent intermediates through an H-bond with the N4 atom of the substrates. These findings improve our understanding of Tet enzyme functions in the dsDNA 5mC and 6mA demethylation pathways, and provide useful information for future discovery of small molecular probes targeting Tet enzymes in DNA active demethylation processes.


Subject(s)
Agaricales , Dioxygenases , 5-Methylcytosine/metabolism , Crystallography, X-Ray , Dioxygenases/chemistry , Dioxygenases/genetics , Dioxygenases/metabolism , DNA Demethylation , DNA Methylation , Fungal Proteins/metabolism , Fungal Proteins/genetics , Fungal Proteins/chemistry , Hydrogen Bonding , Models, Molecular , Oxidation-Reduction , Substrate Specificity , Adenosine/analogs & derivatives , Agaricales/enzymology
11.
FASEB J ; 38(3): e23453, 2024 Feb 15.
Article in English | MEDLINE | ID: mdl-38318639

ABSTRACT

During early development, both genome-wide epigenetic reprogramming and metabolic remodeling are hallmark changes of normal embryogenesis. However, little is known about their relationship and developmental functions during the preimplantation window, which is essential for the acquisition of totipotency and pluripotency. Herein, we reported that glutathione (GSH), a ubiquitous intracellular protective antioxidant that maintains mitochondrial function and redox homeostasis, plays a critical role in safeguarding postfertilization DNA demethylation and is essential for establishing developmental potential in preimplantation embryos. By profiling mitochondria-related transcriptome that coupled with different pluripotency, we found GSH is a potential marker that is tightly correlated with full pluripotency, and its beneficial effect on prompting developmental potential was functionally conformed using in vitro fertilized mouse and bovine embryos as the model. Mechanistic study based on preimplantation embryos and embryonic stem cells further revealed that GSH prompts the acquisition of totipotency and pluripotency by facilitating ten-eleven-translocation (TET)-dependent DNA demethylation, and ascorbic acid (AsA)-GSH cycle is implicated in the process. In addition, we also reported that GSH serves as an oviductal paracrine factor that supports development potential of preimplantation embryos. Thus, our results not only advance the current knowledge of functional links between epigenetic reprogramming and metabolic remodeling during preimplantation development but also provided a promising approach for improving current in vitro culture system for assisted reproductive technology.


Subject(s)
DNA Demethylation , DNA Methylation , Animals , Cattle , Mice , Blastocyst/metabolism , Embryonic Stem Cells/metabolism , Glutathione/metabolism , Embryonic Development/genetics
12.
Annu Rev Immunol ; 42(1): 455-488, 2024 Jun.
Article in English | MEDLINE | ID: mdl-38360546

ABSTRACT

Ten-eleven translocation (TET) proteins are iron-dependent and α-ketoglutarate-dependent dioxygenases that sequentially oxidize the methyl group of 5-methylcytosine (5mC) to 5-hydroxymethylcytosine (5hmC), 5-formylcytosine (5fC) and 5-carboxylcytosine (5caC). All three epigenetic modifications are intermediates in DNA demethylation. TET proteins are recruited by transcription factors and by RNA polymerase II to modify 5mC at enhancers and gene bodies, thereby regulating gene expression during development, cell lineage specification, and cell activation. It is not yet clear, however, how the established biochemical activities of TET enzymes in oxidizing 5mC and mediating DNA demethylation relate to the known association of TET deficiency with inflammation, clonal hematopoiesis, and cancer. There are hints that the ability of TET deficiency to promote cell proliferation in a signal-dependent manner may be harnessed for cancer immunotherapy. In this review, we draw upon recent findings in cells of the immune system to illustrate established as well as emerging ideas of how TET proteins influence cellular function.


Subject(s)
DNA Demethylation , Dioxygenases , Immunotherapy , Inflammation , Neoplasms , Humans , Neoplasms/therapy , Neoplasms/immunology , Neoplasms/etiology , Neoplasms/metabolism , Animals , Inflammation/metabolism , Inflammation/immunology , Immunotherapy/methods , Dioxygenases/metabolism , Immune System/metabolism , Immune System/immunology , Epigenesis, Genetic , Proto-Oncogene Proteins/metabolism , Proto-Oncogene Proteins/genetics , DNA Methylation , DNA-Binding Proteins/metabolism , DNA-Binding Proteins/genetics , Mixed Function Oxygenases/metabolism , Mixed Function Oxygenases/genetics
13.
Genes (Basel) ; 15(1)2024 01 08.
Article in English | MEDLINE | ID: mdl-38254969

ABSTRACT

DNA methylation is critically involved in the regulation of chromatin states and cell-type-specific gene expression. The exclusive expression of imprinted genes from either the maternal or the paternal allele is regulated by allele-specific DNA methylation at imprinting control regions (ICRs). Aberrant DNA hyper- or hypomethylation at the ICR1 of the H19/IGF2 imprinting locus is characteristic for the imprinting disorders Beckwith-Wiedemann syndrome (BWS) and Silver-Russell syndrome (SRS), respectively. In this paper, we performed epigenome editing to induce targeted DNA demethylation at ICR1 in HEK293 cells using dCas9-SunTag and the catalytic domain of TET1. 5-methylcytosine (5mC) levels at the target locus were reduced up to 90% and, 27 days after transient transfection, >60% demethylation was still observed. Consistent with the stable demethylation of CTCF-binding sites within the ICR1, the occupancy of the DNA methylation-sensitive insulator CTCF protein increased by >2-fold throughout the 27 days. Additionally, the H19 expression was increased by 2-fold stably, while IGF2 was repressed though only transiently. Our data illustrate the ability of epigenome editing to implement long-term changes in DNA methylation at imprinting control regions after a single transient treatment, potentially paving the way for therapeutic epigenome editing approaches in the treatment of imprinting disorders.


Subject(s)
DNA Demethylation , Imprinting Disorders , Humans , Catalytic Domain , Epigenome , HEK293 Cells , Alleles , Mixed Function Oxygenases/genetics , Proto-Oncogene Proteins , Insulin-Like Growth Factor II/genetics
14.
J Virol ; 98(2): e0172123, 2024 Feb 20.
Article in English | MEDLINE | ID: mdl-38179947

ABSTRACT

Liver-specific ten-eleven translocation (Tet) methylcytosine dioxygenases 2 and 3 (Tet2 plus Tet3)-deficient hepatitis B virus (HBV) transgenic mice fail to support viral biosynthesis. The levels of viral transcription and replication intermediates are dramatically reduced. Hepatitis B core antigen is only observed in a very limited number of pericentral hepatocytes in a pattern that is similar to glutamate-ammonia ligase (Glul), a ß-catenin target gene. HBV transcript abundance in adult Tet-deficient mice resembles that observed in wild-type neonatal mice. Furthermore, the RNA levels of several ß-catenin target genes including Glul, Lhpp, Notun, Oat, Slc1a2, and Tbx3 in Tet-deficient mice were also similar to that observed in wild-type neonatal mice. As HBV transcription is regulated by ß-catenin, these findings support the suggestion that neonatal Tet deficiency might limit ß-catenin target gene expression, limiting viral biosynthesis. Additionally, HBV transgene DNA displays increased 5-methylcytosine (5mC) frequency at CpG sequences consistent with neonatal Tet deficiency being responsible for decreased developmental viral DNA demethylation mediated by 5mC oxidation to 5-hydroxymethylcytosine, a process that might be responsible for the reduction in cellular ß-catenin target gene expression and viral transcription and replication.IMPORTANCEChronic hepatitis B virus (HBV) infection causes significant worldwide morbidity and mortality. There are no curative therapies available to resolve chronic HBV infections, and the small viral genome limits molecular targets for drug development. An alternative approach to drug development is to target cellular genes essential for HBV biosynthesis. In the liver, ten-eleven translocation (Tet) genes encode cellular enzymes that are not essential for postnatal mouse development but represent essential activities for viral DNA demethylation and transcription. Consequently, Tet inhibitors may potentially be developed into therapeutic agents capable of inducing and/or maintaining HBV covalently closed circular DNA methylation, resulting in transcriptional silencing and the resolution of chronic viral infection.


Subject(s)
DNA-Binding Proteins , Dioxygenases , Hepatitis B virus , Animals , Mice , beta Catenin/genetics , Dioxygenases/genetics , Dioxygenases/metabolism , DNA Demethylation , DNA Methylation , DNA, Viral/genetics , DNA, Viral/metabolism , DNA-Binding Proteins/genetics , DNA-Binding Proteins/metabolism , Hepatitis B virus/metabolism , Mice, Transgenic
15.
Nat Commun ; 15(1): 184, 2024 Jan 02.
Article in English | MEDLINE | ID: mdl-38167803

ABSTRACT

The intracellular ATP-ribosyltransferases PARP1 and PARP2, contribute to DNA base excision repair (BER) and DNA demethylation and have been implicated in epigenetic programming in early mammalian development. Recently, proteomic analyses identified BER proteins to be covalently poly-ADP-ribosylated by PARPs. The role of this posttranslational modification in the BER process is unknown. Here, we show that PARP1 senses AP-sites and SSBs generated during TET-TDG mediated active DNA demethylation and covalently attaches PAR to each BER protein engaged. Covalent PARylation dissociates BER proteins from DNA, which accelerates the completion of the repair process. Consistently, inhibition of PARylation in mESC resulted both in reduced locus-specific TET-TDG-targeted DNA demethylation, and in reduced general repair of random DNA damage. Our findings establish a critical function of covalent protein PARylation in coordinating molecular processes associated with dynamic DNA methylation.


Subject(s)
DNA Repair , Excision Repair , Animals , Poly ADP Ribosylation , DNA Demethylation , Proteomics , Poly (ADP-Ribose) Polymerase-1/metabolism , DNA Damage , DNA/genetics , DNA/metabolism , Mammals/genetics
16.
Biochem Biophys Res Commun ; 695: 149463, 2024 Feb 05.
Article in English | MEDLINE | ID: mdl-38176172

ABSTRACT

Cisplatin-induced acute kidney injury (AKI) restricts the use of cisplatin as a first-line chemotherapeutic agent. Our previous study showed that prophylactic vitamin C supplementation may act as an epigenetic modulator in alleviating cisplatin-induced AKI in mice. However, the targets of vitamin C and the mechanisms underlying the epigenetics changes remain largely unknown. Herein, whole-genome bisulfite sequencing and bulk RNA sequencing were performed on the kidney tissues of mice treated with cisplatin with prophylactic vitamin C supplementation (treatment mice) or phosphate-buffered saline (control mice) at 24 h after cisplatin treatment. Ascorbyl phosphate magnesium (APM), an oxidation-resistant vitamin C derivative, was found that led to global hypomethylation in the kidney tissue and regulated different functional genes in the promoter region and gene body region. Integrated evidence suggested that APM enhanced renal ion transport and metabolism, and reduced apoptosis and inflammation in the kidney tissues. Strikingly, Mapk15, Slc22a6, Cxcl5, and Cd44 were the potential targets of APM that conferred protection against cisplatin-induced AKI. Moreover, APM was found to be difficult to rescue cell proliferation and apoptosis caused by cisplatin in the Slc22a6 knockdown cell line. These results elucidate the mechanism by which vitamin C as an epigenetic regulator to protects against cisplatin-induced AKI and provides a new perspective and evidence support for controlling the disease process through regulating DNA methylation.


Subject(s)
Acute Kidney Injury , Antineoplastic Agents , Mice , Animals , Cisplatin/adverse effects , Antineoplastic Agents/pharmacology , DNA Demethylation , Acute Kidney Injury/chemically induced , Acute Kidney Injury/genetics , Acute Kidney Injury/prevention & control , Kidney/metabolism , Apoptosis , Magnesium/metabolism , Vitamins/pharmacology , Dietary Supplements , Ascorbic Acid/metabolism , Phosphates/metabolism , Mice, Inbred C57BL
17.
Biochem Pharmacol ; 219: 115913, 2024 01.
Article in English | MEDLINE | ID: mdl-37995981

ABSTRACT

The role of cancer stem cells in metastasis, recurrence, and resistance to conventional therapies is significant. Addressing these cells could potentially decrease cancer reoccurrences and mortality rates. TET1, a crucial gene involved in stem cell self-renewal and potency, may also play a part in cancer stem cells, which warrants further research. To explore the role of TET1 in cancer stem cells, we conducted experiments involving loss and gain. We then analyzed factors such as migration, invasion, cell cycle, cell viability, mammosphere formation, and the CD44+/CD24- subpopulation of cancer cells. We also investigate the influence of TET1 on CCNB1, CDK1, and OCT4. Our study reveals that TET1 can regulate the phenotype of cancer stem cells via OCT4. Additionally, it can control the cell cycle by increasing CDK1 and CCNB1 levels. These findings suggest that targeting DNA methylation and TET1 could be an effective strategy to overcome obstacles posed by Cancer stem cells. Our research also indicates that TET1 can influence the phenotype of cancer stem cells and the cell cycle of breast cancer cells potentially through OCT4, CCNB1, and CDK1. This highlights the importance of TET1 in breast cancer cases and suggests a potential therapeutic approach through DNA methylation and modulation of TET1.


Subject(s)
Mixed Function Oxygenases , Proto-Oncogene Proteins , Triple Negative Breast Neoplasms , Female , Humans , Cell Cycle , Cell Line, Tumor , DNA Demethylation , DNA Methylation , Mixed Function Oxygenases/genetics , Neoplastic Stem Cells/pathology , Proto-Oncogene Proteins/genetics , Triple Negative Breast Neoplasms/genetics
18.
J Biol Chem ; 300(2): 105597, 2024 Feb.
Article in English | MEDLINE | ID: mdl-38160798

ABSTRACT

Increased expression of angiotensin II AT1A receptor (encoded by Agtr1a) and Na+-K+-Cl- cotransporter-1 (NKCC1, encoded by Slc12a2) in the hypothalamic paraventricular nucleus (PVN) contributes to hypertension development. However, little is known about their transcriptional control in the PVN in hypertension. DNA methylation is a critical epigenetic mechanism that regulates gene expression. Here, we determined whether transcriptional activation of Agtr1a and Slc12a2 results from altered DNA methylation in spontaneously hypertensive rats (SHR). Methylated DNA immunoprecipitation and bisulfite sequencing-PCR showed that CpG methylation at Agtr1a and Slc12a2 promoters in the PVN was progressively diminished in SHR compared with normotensive Wistar-Kyoto rats (WKY). Chromatin immunoprecipitation-quantitative PCR revealed that enrichment of DNA methyltransferases (DNMT1 and DNMT3A) and methyl-CpG binding protein 2, a DNA methylation reader protein, at Agtr1a and Slc12a2 promoters in the PVN was profoundly reduced in SHR compared with WKY. By contrast, the abundance of ten-eleven translocation enzymes (TET1-3) at Agtr1a and Slc12a2 promoters in the PVN was much greater in SHR than in WKY. Furthermore, microinjecting of RG108, a selective DNMT inhibitor, into the PVN of WKY increased arterial blood pressure and correspondingly potentiated Agtr1a and Slc12a2 mRNA levels in the PVN. Conversely, microinjection of C35, a specific TET inhibitor, into the PVN of SHR markedly reduced arterial blood pressure, accompanied by a decrease in Agtr1a and Slc12a2 mRNA levels in the PVN. Collectively, our findings suggest that DNA hypomethylation resulting from the DNMT/TET switch at gene promoters in the PVN promotes transcription of Agtr1a and Slc12a2 and hypertension development.


Subject(s)
DNA Demethylation , Hypothalamus , Receptor, Angiotensin, Type 1 , Solute Carrier Family 12, Member 2 , Animals , Rats , Blood Pressure , DNA/metabolism , Hypertension/metabolism , Hypothalamus/metabolism , Paraventricular Hypothalamic Nucleus/metabolism , Rats, Inbred SHR , Rats, Inbred WKY , Receptor, Angiotensin, Type 1/metabolism , RNA, Messenger/genetics , Sympathetic Nervous System/metabolism , Solute Carrier Family 12, Member 2/metabolism
19.
J Cell Physiol ; 239(2): e31170, 2024 Feb.
Article in English | MEDLINE | ID: mdl-38149721

ABSTRACT

Osteoarthritis (OA) is one of the most prevalent joint diseases in aged people and characterized by articular cartilage degeneration, synovial inflammation, and abnormal bone remodeling. Recent advances in OA research have clearly shown that OA development is associated with aberrant DNA methylation status of many OA-related genes. As one of most important cartilage degrading proteases in OA, a disintegrin and metalloproteinase with thrombospondin motifs subtype 5 (ADAMTS-5) is activated to mediate cartilage degradation in human OA and experimental murine OA models. The pathological factors and signaling pathways mediating ADAMTS-5 activation during OA development are not well defined and have been a focus of intense research. ADAMTS-5 promoter is featured by CpG islands. So far there have been no reports concerning the DNA methylation status in ADAMTS-5 promoter during OA development. In this study, we sought to investigate DNA methylation status in ADAMTS-5 promoter, the role of DNA methylation in ADAMTS-5 activation in OA, and the underlying mechanisms. The potential for anti-OA intervention therapy which is based on modulating DNA methylation is also explored. Our results showed that DNA methyltransferases 1 (Dnmt1) downregulation-associated ADAMTS-5 promoter demethylation played an important role in ADAMTS-5 activation in OA, which facilitated SPI-1 binding on ADAMTS-5 promoter to activate ADAMTS-5 expression. More importantly, OA pathological phenotype of mice was alleviated in response to Dnmt1-induced DNA methylation of ADAMTS-5 promoter. Our study will benefit not only for deeper insights into the functional role and regulation mechanisms of ADAMTS-5 in OA, but also for the discovery of disease-modifying OA drugs on the basis of ADAMTS-5 via modulating DNA methylation status.


Subject(s)
Cartilage, Articular , Intercellular Signaling Peptides and Proteins , Osteoarthritis , Aged , Animals , Humans , Male , Mice , Cartilage, Articular/metabolism , Chondrocytes/metabolism , DNA Demethylation , HEK293 Cells , Mice, Inbred C57BL , Osteoarthritis/pathology , Promoter Regions, Genetic/genetics
20.
Cell Rep Methods ; 3(12): 100663, 2023 Dec 18.
Article in English | MEDLINE | ID: mdl-38070507

ABSTRACT

Small molecules have enabled expansion of hematopoietic stem and progenitor cells (HSPCs), but limited knowledge is available on whether these agonists can act synergistically. In this work, we identify a stem cell agonist in AA2P and optimize a series of stem cell agonist cocktails (SCACs) to help promote robust expansion of human HSPCs. We find that SCACs provide strong growth-promoting activities while promoting retention and function of immature HSPC. We show that AA2P-mediated HSPC expansion is driven through DNA demethylation leading to enhanced expression of AXL and GAS6. Further, we demonstrate that GAS6 enhances the serial engraftment activity of HSPCs and show that the GAS6/AXL pathway is critical for robust HSPC expansion.


Subject(s)
DNA Demethylation , Hematopoietic Stem Cell Transplantation , Humans , Cells, Cultured , Hematopoietic Stem Cells/metabolism
SELECTION OF CITATIONS
SEARCH DETAIL