Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 368
Filtrar
1.
Folia Biol (Praha) ; 70(2): 113-122, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-39231319

RESUMO

Recent studies have highlighted the significant role of 5-hydroxymethylcytosine (5hmC) in carcinogenesis. However, the specific role of 5hmC in osteosarcoma (OS) remains largely unexplored. The-re-fore, this study aimed to investigate the function of 5hmC and TET3 in OS. In this study, we found a decreased total level of 5hmC in OS tissues. The expression of the TET3 protein was also decreased in OS. Importantly, the decreased levels of TET3 were associated with a decreased disease-free survival (DFS) rate in patients. To investigate the role of TET3 and 5hmC in OS, we manipulated the levels of TET3 in MG-63 cells. Silencing TET3 in these cells resulted in a twofold increase in proliferation. Additio-nally, the level of 5hmC decreased in these cells. Con-versely, over-expression of TET3 in MG-63 cells led to the expected inhibition of proliferation and invasion, accompanied by an increase in 5hmC levels. In conclusion, both 5hmC and TET3 protein levels were decreased in OS. Additionally, the over-expression of TET3 inhibited the proliferation of MG-63 cells, while the suppression of TET3 had the opposite effect. These findings suggest that decreased levels of 5hmC and TET3 may serve as potential markers for OS.


Assuntos
5-Metilcitosina , Proliferação de Células , Desmetilação do DNA , Dioxigenases , Epigênese Genética , Feminino , Humanos , Masculino , 5-Metilcitosina/análogos & derivados , 5-Metilcitosina/metabolismo , Neoplasias Ósseas/genética , Neoplasias Ósseas/metabolismo , Neoplasias Ósseas/patologia , Linhagem Celular Tumoral , Dioxigenases/metabolismo , Regulação Neoplásica da Expressão Gênica , Osteossarcoma/genética , Osteossarcoma/metabolismo , Osteossarcoma/patologia , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Proto-Oncogênicas/genética
2.
Clin Epigenetics ; 16(1): 125, 2024 Sep 11.
Artigo em Inglês | MEDLINE | ID: mdl-39261937

RESUMO

BACKGROUND: Breast tumorigenesis is a complex and multistep process accompanied by both genetic and epigenetic dysregulation. In contrast to the extensive studies on DNA epigenetic modifications 5-hydroxymethylcytosine (5hmC) and 5-methylcytosine (5mC) in malignant breast tumors, their roles in the early phases of breast tumorigenesis remain ambiguous. RESULTS: DNA 5hmC and 5mC exhibited a consistent and significant decrease from usual ductal hyperplasia to atypical ductal hyperplasia and subsequently to ductal carcinoma in situ (DCIS). However, 5hmC showed a modest increase in invasive ductal breast cancer compared to DCIS. Genomic analyses showed that the changes in 5hmC and 5mC levels occurred around the transcription start sites (TSSs), and the modification levels were strongly correlated with gene expression levels. Meanwhile, it was found that differentially hydroxymethylated regions (DhMRs) and differentially methylated regions (DMRs) were overlapped in the early phases and accompanied by the enrichment of active histone marks. In addition, TET2-related DNA demethylation was found to be involved in breast tumorigenesis, and four transcription factor binding sites (TFs: ESR1, FOXA1, GATA3, FOS) were enriched in TET2-related DhMRs/DMRs. Intriguingly, we also identified a certain number of common DhMRs between tumor samples and cell-free DNA (cfDNA). CONCLUSIONS: Our study reveals that dynamic changes in DNA 5hmC and 5mC play a vital role in propelling breast tumorigenesis. Both TFs and active histone marks are involved in TET2-related DNA demethylation. Concurrent changes in 5hmC signals in primary breast tumors and cfDNA may play a promising role in breast cancer screening.


Assuntos
5-Metilcitosina , Neoplasias da Mama , Proteínas de Ligação a DNA , Dioxigenases , Proteínas Proto-Oncogênicas , Humanos , 5-Metilcitosina/análogos & derivados , 5-Metilcitosina/metabolismo , Feminino , Neoplasias da Mama/genética , Proteínas de Ligação a DNA/genética , Dioxigenases/genética , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas/metabolismo , Carcinogênese/genética , Metilação de DNA/genética , Epigênese Genética/genética , Regulação Neoplásica da Expressão Gênica , Desmetilação do DNA
3.
Sci Adv ; 10(35): eado5424, 2024 Aug 30.
Artigo em Inglês | MEDLINE | ID: mdl-39196941

RESUMO

DNA methylation is extensively reconfigured during development, but the functional significance and cell type-specific dependencies of DNA demethylation in lineage specification remain poorly understood. Here, we demonstrate that developmental DNA demethylation, driven by ten-eleven translocation 1/2/3 (TET1/2/3) enzymes, is essential for establishment of neural stem cell (NSC) identity and gliogenic potential. We find that loss of all three TETs during NSC specification is dispensable for neural induction and neuronal differentiation but critical for astrocyte and oligodendrocyte formation, demonstrating a selective loss of glial competence. Mechanistically, TET-mediated demethylation was essential for commissioning neural-specific enhancers in proximity to master neurodevelopmental and glial transcription factor genes and for induction of these genes. Consistently, loss of all three TETs in embryonic NSCs in mice compromised glial gene expression and corticogenesis. Thus, TET-dependent developmental demethylation is an essential regulatory mechanism for neural enhancer commissioning during NSC specification and is a cell-intrinsic determinant of NSC identity and gliogenic potential.


Assuntos
Diferenciação Celular , Desmetilação do DNA , Células-Tronco Neurais , Animais , Células-Tronco Neurais/metabolismo , Células-Tronco Neurais/citologia , Camundongos , Proteínas de Ligação a DNA/metabolismo , Proteínas de Ligação a DNA/genética , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Proto-Oncogênicas/genética , Neuroglia/metabolismo , Neuroglia/citologia , Neurogênese , Regulação da Expressão Gênica no Desenvolvimento , Metilação de DNA , Elementos Facilitadores Genéticos , Dioxigenases/metabolismo , Neurônios/metabolismo , Neurônios/citologia
4.
Nat Commun ; 15(1): 7310, 2024 Aug 25.
Artigo em Inglês | MEDLINE | ID: mdl-39181896

RESUMO

In mammals, global passive demethylation contributes to epigenetic reprogramming during early embryonic development. At this stage, the majority of DNA-methyltransferase 1 (DNMT1) protein is excluded from nucleus, which is considered the primary cause. However, whether the remaining nuclear activity of DNMT1 is regulated by additional mechanisms is unclear. Here, we report that nuclear DNMT1 abundance is finetuned through proteasomal degradation in mouse zygotes. We identify a maternal factor, Pramel15, which targets DNMT1 for degradation via Cullin-RING E3 ligases. Loss of Pramel15 elevates DNMT1 levels in the zygote pronuclei, impairs zygotic DNA demethylation, and causes a stochastic gain of DNA methylation in early embryos. Thus, Pramel15 can modulate the residual level of DNMT1 in the nucleus during zygotic DNA replication, thereby ensuring efficient DNA methylation reprogramming in early embryos.


Assuntos
Núcleo Celular , DNA (Citosina-5-)-Metiltransferase 1 , Desmetilação do DNA , Zigoto , Animais , DNA (Citosina-5-)-Metiltransferase 1/metabolismo , DNA (Citosina-5-)-Metiltransferase 1/genética , Zigoto/metabolismo , Camundongos , Núcleo Celular/metabolismo , Feminino , Metilação de DNA , Proteólise , Desenvolvimento Embrionário/genética , Masculino , Embrião de Mamíferos/metabolismo , Camundongos Knockout , Regulação da Expressão Gênica no Desenvolvimento , Replicação do DNA
5.
Science ; 385(6705): eadl6173, 2024 Jul 12.
Artigo em Inglês | MEDLINE | ID: mdl-38991060

RESUMO

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.


Assuntos
Evasão da Resposta Imune , Imunidade Inata , Isocitrato Desidrogenase , Neoplasias , Animais , Humanos , Camundongos , Linhagem Celular Tumoral , DNA/metabolismo , Desmetilação do DNA , Metilação de DNA , Elementos de DNA Transponíveis , Epigênese Genética , Glutaratos/metabolismo , Imunidade Inata/genética , Isocitrato Desidrogenase/genética , Isocitrato Desidrogenase/metabolismo , Mutação , Neoplasias/imunologia , Neoplasias/genética , Nucleotidiltransferases/genética , Evasão Tumoral , Evasão da Resposta Imune/genética
6.
Cryobiology ; 116: 104947, 2024 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-39084504

RESUMO

Grapevine (Vitis vinifera L.) crops are continuously exposed to biotic and abiotic stresses, which can cause genetic and epigenetic alterations. To determine the possible effects of grapevine cryopreservation on the regulation of DNA demethylase genes, this work studied the expression of DNA demethylase genes in cryopreserved and post-cryopreserved grapevine tissues. V. vinifera DNA demethylases were characterized by in silico analysis, and gene expression quantification was conducted by RT‒qPCR. Three DNA demethylase sequences were found: VIT_13s0074g00450 (VvDMT), VIT_08s0007g03920 (VvROS1), and VIT_06s0061g01270 (VvDML3). Phylogenetic analysis revealed that the sequences from V. vinifera and A. thaliana had a common ancestry. In the promoters of responsive elements to transcription factors such as AP-2, Myb, bZIP, TBP, and GATA, the conserved domains RRM DME and Perm CXXC were detected. These responsive elements play roles in the response to abiotic stress and the regulation of cell growth. These data helped us characterize the V. vinifera DNA demethylase genes. Gene expression analysis indicated that plant vitrification solution 2 (PVS2) treatment does not alter the expression of DNA demethylase genes. The expression levels of VvDMT and VvROS1 increased in response to cryopreservation by vitrification. Furthermore, in post-cryopreservation, VvROS1 was highly induced, and VvDML3 was repressed in all the treatment groups. Gene expression differences between different treatments and tissues may play roles in controlling methylation patterns during gene regulation in tissues stressed by cryopreservation procedures and in the post-cryopreservation period during plant growth and development.


Assuntos
Criopreservação , Regulação da Expressão Gênica de Plantas , Proteínas de Plantas , Vitis , Vitis/genética , Vitis/crescimento & desenvolvimento , Criopreservação/métodos , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Filogenia , Sementes/genética , Sementes/crescimento & desenvolvimento , Desmetilação do DNA , Zigoto/metabolismo , Metilação de DNA , Crioprotetores/farmacologia
7.
Int J Mol Sci ; 25(11)2024 May 28.
Artigo em Inglês | MEDLINE | ID: mdl-38892059

RESUMO

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.


Assuntos
Desenvolvimento Embrionário , Vesículas Extracelulares , Líquido Folicular , MicroRNAs , Animais , Feminino , MicroRNAs/genética , MicroRNAs/metabolismo , Bovinos , Líquido Folicular/metabolismo , Vesículas Extracelulares/metabolismo , Desenvolvimento Embrionário/genética , Metilação de DNA , Desmetilação do DNA , Oócitos/metabolismo , Blastocisto/metabolismo , Embrião de Mamíferos/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Zigoto/metabolismo
8.
Mol Biol Rep ; 51(1): 778, 2024 Jun 21.
Artigo em Inglês | MEDLINE | ID: mdl-38904842

RESUMO

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.


Assuntos
Metilação de DNA , Histona-Lisina N-Metiltransferase , Dedos de Zinco , Histona-Lisina N-Metiltransferase/genética , Histona-Lisina N-Metiltransferase/metabolismo , Humanos , Dedos de Zinco/genética , Metilação de DNA/genética , Regiões Promotoras Genéticas/genética , Regulação para Cima/genética , Desmetilação do DNA , Linhagem Celular , Ilhas de CpG/genética , Deleção de Genes , Histonas/metabolismo , Histonas/genética
9.
Proc Natl Acad Sci U S A ; 121(22): e2320468121, 2024 May 28.
Artigo em Inglês | MEDLINE | ID: mdl-38768356

RESUMO

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.


Assuntos
Proteínas de Arabidopsis , Arabidopsis , Desmetilação do DNA , Metilação de DNA , Epigênese Genética , Mutação , Arabidopsis/genética , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Regulação da Expressão Gênica de Plantas , DNA de Plantas/genética , DNA de Plantas/metabolismo , Proteínas Nucleares
10.
Nat Commun ; 15(1): 3734, 2024 May 03.
Artigo em Inglês | MEDLINE | ID: mdl-38702312

RESUMO

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.


Assuntos
Desmetilação do DNA , Reparo do DNA , DNA Polimerase Dirigida por DNA , Células Germinativas , Animais , Feminino , Humanos , Masculino , Camundongos , Dano ao DNA , DNA Polimerase Dirigida por DNA/metabolismo , DNA Polimerase Dirigida por DNA/genética , Epigênese Genética , Células Germinativas/metabolismo , Meiose/genética , Camundongos Knockout , Nucleotidiltransferases/metabolismo , Nucleotidiltransferases/genética , Antígeno Nuclear de Célula em Proliferação/metabolismo , Síntese de DNA Translesão
11.
BMC Genomics ; 25(1): 344, 2024 Apr 05.
Artigo em Inglês | MEDLINE | ID: mdl-38580899

RESUMO

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.


Assuntos
Proteínas Cromossômicas não Histona , Desmetilação do DNA , Epigênese Genética , Animais , Feminino , Camundongos , Proteínas Cromossômicas não Histona/metabolismo , Metilação de DNA , Genoma , Células Germinativas/metabolismo , Mamíferos/genética
12.
J Gynecol Oncol ; 35(5): e64, 2024 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-38456588

RESUMO

OBJECTIVE: Src homology phosphotyrosin phosphatase 2 (SHP2) has been implicated in the progression of several cancer types. However, its function in endometrial cancer (EC) remains unclear. Here, we report that the ten-eleven translocation 3 (TET3)-mediated DNA demethylation modification is responsible for the oncogenic role of SHP2 in EC and explore the detailed mechanism. METHODS: The transcriptomic differences between EC tissues and control tissues were analyzed using bioinformatics tools, followed by protein-protein interaction network establishment. EC cells were treated with shRNA targeting SHP2 alone or in combination with isoprocurcumenol, an epidermal growth factor receptor (EGFR) signaling activator. The cell biological behavior was examined using cell counting kit-8, colony formation, flow cytometry, scratch assay, and transwell assays, and the median inhibition concentration values to medroxyprogesterone acetate/gefitinib were calculated. The binding of TET3 to the SHP2 promoter was verified. EC cells with TET3 knockdown and combined with SHP2 overexpression were selected to construct tumor xenografts in mice. RESULTS: TET3 and SHP2 were overexpressed in EC cells. TET3 bound to the SHP2 promoter, thereby increasing the DNA hydroxymethylation modification and activating SHP2 to induce the EGFR/extracellular signal-regulated kinase (ERK) pathway. Knockdown of TET3 or SHP2 inhibited EC cell malignant aggressiveness and impaired the EGFR/ERK pathway. Silencing of TET3 inhibited the tumorigenic capacity of EC cells, and ectopic expression of SHP2 or isoprocurcumenol reversed the inhibitory effect of TET3 knockdown on the biological activity of EC cells. CONCLUSION: TET3 promoted the DNA demethylation modification in the SHP2 promoter and activated SHP2, thus activating the EGFR/ERK pathway and leading to EC progression.


Assuntos
Desmetilação do DNA , Dioxigenases , Progressão da Doença , Neoplasias do Endométrio , Receptores ErbB , Sistema de Sinalização das MAP Quinases , Proteína Tirosina Fosfatase não Receptora Tipo 11 , Feminino , Humanos , Proteína Tirosina Fosfatase não Receptora Tipo 11/metabolismo , Proteína Tirosina Fosfatase não Receptora Tipo 11/genética , Receptores ErbB/metabolismo , Receptores ErbB/genética , Animais , Neoplasias do Endométrio/patologia , Neoplasias do Endométrio/genética , Neoplasias do Endométrio/metabolismo , Linhagem Celular Tumoral , Dioxigenases/metabolismo , Camundongos , Sistema de Sinalização das MAP Quinases/fisiologia , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Proto-Oncogênicas/genética , Camundongos Nus , Regulação Neoplásica da Expressão Gênica , Regiões Promotoras Genéticas , Camundongos Endogâmicos BALB C
13.
Sci Rep ; 14(1): 6481, 2024 03 18.
Artigo em Inglês | MEDLINE | ID: mdl-38499584

RESUMO

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.


Assuntos
Neoplasias da Mama , Dioxigenases , Humanos , Feminino , Desmetilação do DNA , Neoplasias da Mama/genética , Cromatografia Líquida , Espectrometria de Massas em Tandem , 5-Metilcitosina/metabolismo , Metilação de DNA , Biomarcadores/metabolismo , DNA/metabolismo , Epigênese Genética , Leucócitos/metabolismo , Carcinogênese/genética , Dioxigenases/genética
14.
Sci Rep ; 14(1): 6155, 2024 03 14.
Artigo em Inglês | MEDLINE | ID: mdl-38486042

RESUMO

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.


Assuntos
Adenosina/análogos & derivados , Carcinoma Hepatocelular , Neoplasias Hepáticas , MicroRNAs , Proteína de Ligação a Elemento Regulador de Esterol 2 , Humanos , Carcinoma Hepatocelular/tratamento farmacológico , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/metabolismo , Sorafenibe/farmacologia , Neoplasias Hepáticas/tratamento farmacológico , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/metabolismo , Desmetilação do DNA , Linhagem Celular Tumoral , MicroRNAs/genética , Proteínas de Ligação a RNA/metabolismo , Oxigenases de Função Mista/metabolismo , Proteínas Proto-Oncogênicas/metabolismo , Metiltransferases/genética , Metiltransferases/metabolismo
15.
Annu Rev Immunol ; 42(1): 455-488, 2024 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-38360546

RESUMO

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.


Assuntos
Desmetilação do DNA , Dioxigenases , Imunoterapia , Inflamação , Neoplasias , Humanos , Neoplasias/terapia , Neoplasias/imunologia , Neoplasias/etiologia , Neoplasias/metabolismo , Animais , Inflamação/metabolismo , Inflamação/imunologia , Imunoterapia/métodos , Dioxigenases/metabolismo , Sistema Imunitário/metabolismo , Sistema Imunitário/imunologia , Epigênese Genética , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Proto-Oncogênicas/genética , Metilação de DNA , Proteínas de Ligação a DNA/metabolismo , Proteínas de Ligação a DNA/genética , Oxigenases de Função Mista/metabolismo , Oxigenases de Função Mista/genética
16.
FASEB J ; 38(3): e23453, 2024 Feb 15.
Artigo em Inglês | MEDLINE | ID: mdl-38318639

RESUMO

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.


Assuntos
Desmetilação do DNA , Metilação de DNA , Animais , Bovinos , Camundongos , Blastocisto/metabolismo , Células-Tronco Embrionárias/metabolismo , Glutationa/metabolismo , Desenvolvimento Embrionário/genética
17.
Sci Rep ; 14(1): 2683, 2024 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-38302503

RESUMO

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.


Assuntos
Neoplasias do Colo , Neoplasias Colorretais , Humanos , Epigênese Genética , Desmetilação do DNA , Metilação de DNA , Linhagem Celular Tumoral , Neoplasias do Colo/genética , Regulação Neoplásica da Expressão Gênica , Neoplasias Colorretais/patologia , Ilhas de CpG , Antígenos de Neoplasias/genética , Antígenos de Neoplasias/metabolismo , Moléculas de Adesão Celular/genética , Moléculas de Adesão Celular/metabolismo
18.
Nucleic Acids Res ; 52(7): 3886-3895, 2024 Apr 24.
Artigo em Inglês | MEDLINE | ID: mdl-38324471

RESUMO

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.


Assuntos
Agaricales , Dioxigenases , 5-Metilcitosina/metabolismo , Cristalografia por Raios X , Dioxigenases/química , Dioxigenases/genética , Dioxigenases/metabolismo , Desmetilação do DNA , Metilação de DNA , Proteínas Fúngicas/metabolismo , Proteínas Fúngicas/genética , Proteínas Fúngicas/química , Ligação de Hidrogênio , Modelos Moleculares , Oxirredução , Especificidade por Substrato , Adenosina/análogos & derivados , Agaricales/enzimologia
19.
Genes (Basel) ; 15(1)2024 01 08.
Artigo em Inglês | MEDLINE | ID: mdl-38254969

RESUMO

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.


Assuntos
Desmetilação do DNA , Transtornos da Impressão Genômica , Humanos , Domínio Catalítico , Epigenoma , Células HEK293 , Alelos , Oxigenases de Função Mista/genética , Proteínas Proto-Oncogênicas , Fator de Crescimento Insulin-Like II/genética
20.
Nat Commun ; 15(1): 184, 2024 Jan 02.
Artigo em Inglês | MEDLINE | ID: mdl-38167803

RESUMO

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.


Assuntos
Reparo do DNA , Reparo por Excisão , Animais , Poli ADP Ribosilação , Desmetilação do DNA , Proteômica , Poli(ADP-Ribose) Polimerase-1/metabolismo , Dano ao DNA , DNA/genética , DNA/metabolismo , Mamíferos/genética
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA