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Science (New York, N.Y.) May 2022-methyladenosine (mA) is the most abundant internal modification on mammalian messenger RNA. It is installed by a writer complex and can be reversed by erasers such as...
-methyladenosine (mA) is the most abundant internal modification on mammalian messenger RNA. It is installed by a writer complex and can be reversed by erasers such as the fat mass and obesity-associated protein FTO. Despite extensive research, the primary physiological substrates of FTO in mammalian tissues and development remain elusive. Here, we show that FTO mediates mA demethylation of long-interspersed element-1 (LINE1) RNA in mouse embryonic stem cells (mESCs), regulating LINE1 RNA abundance and the local chromatin state, which in turn modulates the transcription of LINE1-containing genes. FTO-mediated LINE1 RNA mA demethylation also plays regulatory roles in shaping chromatin state and gene expression during mouse oocyte and embryonic development. Our results suggest broad effects of LINE1 RNA mA demethylation by FTO in mammals.
Topics: Adenosine; Alpha-Ketoglutarate-Dependent Dioxygenase FTO; Animals; Chromatin; Demethylation; Gene Expression Regulation, Developmental; Long Interspersed Nucleotide Elements; Mice; Mouse Embryonic Stem Cells; Oocytes; RNA, Messenger
PubMed: 35511947
DOI: 10.1126/science.abe9582 -
Cancer Cell Jun 2023Cyclic GMP-AMP synthase (cGAS) is the major sensor for cytosolic DNA and activates type I interferon signaling and plays an essential role in antitumor immunity....
Cyclic GMP-AMP synthase (cGAS) is the major sensor for cytosolic DNA and activates type I interferon signaling and plays an essential role in antitumor immunity. However, it remains unclear whether the cGAS-mediated antitumor activity is affected by nutrient status. Here, our study reports that methionine deprivation enhances cGAS activity by blocking its methylation, which is catalyzed by methyltransferase SUV39H1. We further show that methylation enhances the chromatin sequestration of cGAS in a UHRF1-dependent manner. Blocking cGAS methylation enhances cGAS-mediated antitumor immunity and suppresses colorectal tumorigenesis. Clinically, cGAS methylation in human cancers correlates with poor prognosis. Thus, our results indicate that nutrient stress promotes cGAS activation via reversible methylation, and suggest a potential therapeutic strategy for targeting cGAS methylation in cancer treatment.
Topics: Humans; Chromatin; Methionine; Nucleotidyltransferases; DNA; Immunity, Innate; Demethylation; CCAAT-Enhancer-Binding Proteins; Ubiquitin-Protein Ligases
PubMed: 37267951
DOI: 10.1016/j.ccell.2023.05.005 -
Science (New York, N.Y.) Dec 2022Neurons harbor high levels of single-strand DNA breaks (SSBs) that are targeted to neuronal enhancers, but the source of this endogenous damage remains unclear. Using...
Neurons harbor high levels of single-strand DNA breaks (SSBs) that are targeted to neuronal enhancers, but the source of this endogenous damage remains unclear. Using two systems of postmitotic lineage specification-induced pluripotent stem cell-derived neurons and transdifferentiated macrophages-we show that thymidine DNA glycosylase (TDG)-driven excision of methylcytosines oxidized with ten-eleven translocation enzymes (TET) is a source of SSBs. Although macrophage differentiation favors short-patch base excision repair to fill in single-nucleotide gaps, neurons also frequently use the long-patch subpathway. Disrupting this gap-filling process using anti-neoplastic cytosine analogs triggers a DNA damage response and neuronal cell death, which is dependent on TDG. Thus, TET-mediated active DNA demethylation promotes endogenous DNA damage, a process that normally safeguards cell identity but can also provoke neurotoxicity after anticancer treatments.
Topics: Cell Differentiation; DNA Demethylation; Induced Pluripotent Stem Cells; Neurons; DNA Breaks, Single-Stranded; Enhancer Elements, Genetic; Thymine DNA Glycosylase; DNA Repair; 5-Methylcytosine; Humans; Cell Transdifferentiation
PubMed: 36454826
DOI: 10.1126/science.add9838 -
Molecular Cell Sep 2018FTO, the first RNA demethylase discovered, mediates the demethylation of internal N-methyladenosine (mA) and N, 2-O-dimethyladenosine (mA) at theĀ +1 position from the...
FTO, the first RNA demethylase discovered, mediates the demethylation of internal N-methyladenosine (mA) and N, 2-O-dimethyladenosine (mA) at theĀ +1 position from the 5' cap in mRNA. Here we demonstrate that the cellular distribution of FTO is distinct among different cell lines, affecting the access of FTO to different RNA substrates. We find that FTO binds multiple RNA species, including mRNA, snRNA, and tRNA, and can demethylate internal mA and cap mA in mRNA, internal mA in U6 RNA, internal and cap mA in snRNAs, and N-methyladenosine (mA) in tRNA. FTO-mediated demethylation has a greater effect on the transcript levels of mRNAs possessing internal mA than the ones with cap mA in the tested cells. We also show that FTO can directly repress translation by catalyzing mA tRNA demethylation. Collectively, FTO-mediated RNA demethylation occurs to mA and mA in mRNA and snRNA as well as mA in tRNA.
Topics: 3T3-L1 Cells; Adenosine; Alpha-Ketoglutarate-Dependent Dioxygenase FTO; Animals; Cell Nucleus; Cytoplasm; Demethylation; Gene Expression; HEK293 Cells; HeLa Cells; Humans; Methylation; Mice; RNA Processing, Post-Transcriptional; RNA, Messenger; RNA, Small Nuclear; RNA, Transfer
PubMed: 30197295
DOI: 10.1016/j.molcel.2018.08.011 -
Cell Metabolism May 2021Poor maternal diet increases the risk of obesity and type 2 diabetes in offspring, adding to the ever-increasing prevalence of these diseases. In contrast, we find that...
Poor maternal diet increases the risk of obesity and type 2 diabetes in offspring, adding to the ever-increasing prevalence of these diseases. In contrast, we find that maternal exercise improves the metabolic health of offspring, and here, we demonstrate that this occurs through a vitamin D receptor-mediated increase in placental superoxide dismutase 3 (SOD3) expression and secretion. SOD3 activates an AMPK/TET signaling axis in fetal offspring liver, resulting in DNA demethylation at the promoters of glucose metabolic genes, enhancing liver function, and improving glucose tolerance. In humans, SOD3 is upregulated in serum and placenta from physically active pregnant women. The discovery of maternal exercise-induced cross talk between placenta-derived SOD3 and offspring liver provides a central mechanism for improved offspring metabolic health. These findings may lead to novel therapeutic approaches to limit the transmission of metabolic disease to the next generation.
Topics: AMP-Activated Protein Kinases; Animals; Cells, Cultured; DNA Demethylation; Diet, High-Fat; Exercise; Female; Hepatocytes; Humans; Liver; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Mixed Function Oxygenases; Placenta; Pregnancy; Proto-Oncogene Proteins; Receptors, Calcitriol; Signal Transduction; Superoxide Dismutase
PubMed: 33770509
DOI: 10.1016/j.cmet.2021.03.004 -
The New England Journal of Medicine May 2022Although hypomethylating agents are currently used to treat patients with cancer, whether they can also reactivate and up-regulate oncogenes is not well elucidated.
BACKGROUND
Although hypomethylating agents are currently used to treat patients with cancer, whether they can also reactivate and up-regulate oncogenes is not well elucidated.
METHODS
We examined the effect of hypomethylating agents on , a known oncogene that plays an important role in myelodysplastic syndrome and other cancers. Paired bone marrow samples that were obtained from two cohorts of patients with myelodysplastic syndrome before and after treatment with a hypomethylating agent were used to explore the relationships among changes in expression, treatment response, and clinical outcome. Leukemic cell lines with low or undetectable expression were used to study the relationship between methylation and expression. A locus-specific demethylation technology, CRISPR-DNMT1-interacting RNA (CRISPR-DiR), was used to identify the CpG island that is critical for expression.
RESULTS
up-regulation after treatment with hypomethylating agents was observed in 10 of 25 patients (40%) in cohort 1 and in 13 of 43 patients (30%) in cohort 2 and was associated with a worse outcome. Using CRISPR-DiR, we discovered that demethylation of a CpG island within the 5' untranslated region was critical for expression. In cell lines and patients, we confirmed that treatment with a hypomethylating agent led to demethylation of the same CpG region and up-regulation of expression.
CONCLUSIONS
By combining analysis of patient samples with CRISPR-DiR technology, we found that demethylation and up-regulation of an oncogene after treatment with a hypomethylating agent can indeed occur and should be further studied. (Funded by Associazione Italiana per la Ricerca sul Cancro and others.).
Topics: Antineoplastic Agents; Clustered Regularly Interspaced Short Palindromic Repeats; Demethylation; Humans; Myelodysplastic Syndromes; Neoplasms; Oncogenes; Transcription Factors; Up-Regulation
PubMed: 35613022
DOI: 10.1056/NEJMoa2119771 -
Journal of Experimental & Clinical... Sep 2021Chemotherapy resistance remains a barrier to improving the prognosis of epithelial ovarian cancer (EOC). ALKBH5 has recently been shown to be one of the RNA...
BACKGROUND
Chemotherapy resistance remains a barrier to improving the prognosis of epithelial ovarian cancer (EOC). ALKBH5 has recently been shown to be one of the RNA N6-methyladenosine (m6A) demethyltransferases associated with various cancers, but its role in cancer therapeutic resistance remains unclear. This study aimed to investigate the role of AlkB homolog 5 (ALKBH5) in cisplatin-resistant EOC.
METHODS
Functional assays were performed both in vitro and in vivo. RNA sequencing (RNA-seq), m6A-modified RNA immunoprecipitation sequencing (MeRIP-seq), chromatin immunoprecipitation, RNA immunoprecipitation, and luciferase reporter and actinomycin-D assays were performed to investigate RNA/RNA interaction and m6A modification of the ALKBH5-HOXA10 loop.
RESULTS
ALKBH5 was upregulated in cisplatin-resistant EOC and promoted cancer cell cisplatin resistance both in vivo and in vitro. Notably, HOXA10 formed a loop with ALKBH5 and was found to be the upstream transcription factor of ALKBH5. HOXA10 overexpression also facilitated EOC cell chemoresistance both in vivo and in vitro. Collective results of MeRIP-seq and RNA-seq showed that JAK2 is the m6A-modified gene targeted by ALKBH5. The JAK2/STAT3 signaling pathway was activated by overexpression of the ALKBH5-HOXA10 loop, resulting in EOC chemoresistance. Cell sensitivity to cisplatin was rescued by ALKBH5 and HOXA10 knockdown or inhibition of the JAK2/STAT3 signaling pathway in EOC cells overexpressing ALKBH5-HOXA10.
CONCLUSIONS
The ALKBH5-HOXA10 loop jointly activates the JAK2/STAT3 signaling pathway by mediating JAK2 m6A demethylation, promoting EOC resistance to cisplatin. Thus, inhibition of the expression of the ALKBH5-HOXA10 loop may be a potential strategy to overcome cisplatin resistance in EOC.
Topics: Animals; Carcinoma, Ovarian Epithelial; Cell Line, Tumor; Demethylation; Drug Resistance, Neoplasm; Female; Homeobox A10 Proteins; Humans; Janus Kinase 2; Mice; Mice, Nude; Transfection
PubMed: 34496932
DOI: 10.1186/s13046-021-02088-1 -
Experimental & Molecular Medicine Apr 2017
Topics: Animals; Chromatin; Cytosine; DNA Demethylation; DNA Methylation; Epigenesis, Genetic; Genetic Markers; Histones; Humans; Neoplasms
PubMed: 28450735
DOI: 10.1038/emm.2017.38 -
The Plant Cell Dec 2017-methyladenosine (mA) is the most abundant, internal, posttranscriptional modification in mRNA among all higher eukaryotes. In mammals, this modification is reversible...
-methyladenosine (mA) is the most abundant, internal, posttranscriptional modification in mRNA among all higher eukaryotes. In mammals, this modification is reversible and plays broad roles in the regulation of mRNA metabolism and processing. Despite its importance, previous studies on the role and mechanism of mA methylation in have been limited. Here, we report that ALKBH10B is a demethylase that oxidatively reverses mA methylation in mRNA in vitro and in vivo. Depletion of ALKBH10B in the mutant delays flowering and represses vegetative growth. Complementation with wild-type , but not a catalytically inactive mutant (), rescues these effects in mutant plants, suggesting the observed phenotypes are controlled by the catalytic action of We show that -mediated mRNA demethylation affects the stability of target transcripts, thereby influencing floral transition. We identified 1190 mA hypermethylated transcripts in the mutant involved in plant development. The discovery and characterization of the archetypical RNA demethylase in Arabidopsis sheds light on the occurrence and functional role(s) of reversible mRNA methylation in plants and defines the role of mA RNA modification in Arabidopsis floral transition.
Topics: Adenosine; Arabidopsis; Arabidopsis Proteins; Demethylation; Flowers; Gene Expression Regulation, Plant; Genes, Plant; Methylation; Mutation; Oxidoreductases, N-Demethylating; Protein Stability; RNA, Messenger; RNA, Plant; RNA-Binding Proteins; Substrate Specificity; Up-Regulation
PubMed: 29180595
DOI: 10.1105/tpc.16.00912 -
Nature Aug 2023Cells undergo a major epigenome reconfiguration when reprogrammed to human induced pluripotent stem cells (hiPS cells). However, the epigenomes of hiPS cells and human...
Cells undergo a major epigenome reconfiguration when reprogrammed to human induced pluripotent stem cells (hiPS cells). However, the epigenomes of hiPS cells and human embryonic stem (hES) cells differ significantly, which affects hiPS cell function. These differences include epigenetic memory and aberrations that emerge during reprogramming, for which the mechanisms remain unknown. Here we characterized the persistence and emergence of these epigenetic differences by performing genome-wide DNA methylation profiling throughout primed and naive reprogramming of human somatic cells to hiPS cells. We found that reprogramming-induced epigenetic aberrations emerge midway through primed reprogramming, whereas DNA demethylation begins early in naive reprogramming. Using this knowledge, we developed a transient-naive-treatment (TNT) reprogramming strategy that emulates the embryonic epigenetic reset. We show that the epigenetic memory in hiPS cells is concentrated in cell of origin-dependent repressive chromatin marked by H3K9me3, lamin-B1 and aberrant CpH methylation. TNT reprogramming reconfigures these domains to a hES cell-like state and does not disrupt genomic imprinting. Using an isogenic system, we demonstrate that TNT reprogramming can correct the transposable element overexpression and differential gene expression seen in conventional hiPS cells, and that TNT-reprogrammed hiPS and hES cells show similar differentiation efficiencies. Moreover, TNT reprogramming enhances the differentiation of hiPS cells derived from multiple cell types. Thus, TNT reprogramming corrects epigenetic memory and aberrations, producing hiPS cells that are molecularly and functionally more similar to hES cells than conventional hiPS cells. We foresee TNT reprogramming becoming a new standard for biomedical and therapeutic applications and providing a novel system for studying epigenetic memory.
Topics: Humans; Cellular Reprogramming; Chromatin; DNA Demethylation; DNA Methylation; DNA Transposable Elements; Epigenesis, Genetic; Induced Pluripotent Stem Cells; Human Embryonic Stem Cells; Lamin Type B
PubMed: 37587336
DOI: 10.1038/s41586-023-06424-7