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Molecular Cell Feb 2022N-methyladenosine (mA) is an abundant RNA modification that plays critical roles in RNA regulation and cellular function. Global mA profiling has revealed important...
N-methyladenosine (mA) is an abundant RNA modification that plays critical roles in RNA regulation and cellular function. Global mA profiling has revealed important aspects of mA distribution and function, but to date such studies have been restricted to large populations of cells. Here, we develop a method to identify mA sites transcriptome-wide in single cells. We uncover surprising heterogeneity in the presence and abundance of mA sites across individual cells and identify differentially methylated mRNAs across the cell cycle. Additionally, we show that cellular subpopulations can be distinguished based on their RNA methylation signatures, independent from gene expression. These studies reveal fundamental features of mA that have been missed by mA profiling of bulk cells and suggest the presence of cell-intrinsic mechanisms for mA deposition.
Topics: Adenosine; Gene Expression Profiling; HEK293 Cells; Humans; Methylation; RNA Processing, Post-Transcriptional; RNA, Messenger; Sequence Analysis, RNA; Single-Cell Analysis; Transcriptome
PubMed: 35081365
DOI: 10.1016/j.molcel.2021.12.038 -
Nature Genetics Aug 2021The most prevalent post-transcriptional mRNA modification, N-methyladenosine (mA), plays diverse RNA-regulatory roles, but its genetic control in human tissues remains...
The most prevalent post-transcriptional mRNA modification, N-methyladenosine (mA), plays diverse RNA-regulatory roles, but its genetic control in human tissues remains uncharted. Here we report 129 transcriptome-wide mA profiles, covering 91 individuals and 4 tissues (brain, lung, muscle and heart) from GTEx/eGTEx. We integrate these with interindividual genetic and expression variation, revealing 8,843 tissue-specific and 469 tissue-shared mA quantitative trait loci (QTLs), which are modestly enriched in, but mostly orthogonal to, expression QTLs. We integrate mA QTLs with disease genetics, identifying 184 GWAS-colocalized mA QTL, including brain mA QTLs underlying neuroticism, depression, schizophrenia and anxiety; lung mA QTLs underlying expiratory flow and asthma; and muscle/heart mA QTLs underlying coronary artery disease. Last, we predict novel mA regulators that show preferential binding in mA QTLs, protein interactions with known mA regulators and expression correlation with the mA levels of their targets. Our results provide important insights and resources for understanding both cis and trans regulation of epitranscriptomic modifications, their interindividual variation and their roles in human disease.
Topics: Adenosine; Brain; Genome-Wide Association Study; Heart; Humans; Lung; Methylation; Muscle, Skeletal; Organ Specificity; Polymorphism, Single Nucleotide; Quantitative Trait Loci; RNA Processing, Post-Transcriptional; RNA-Binding Proteins; Reproducibility of Results
PubMed: 34211177
DOI: 10.1038/s41588-021-00890-3 -
Journal of Drug Targeting 2015Adenosine is a naturally occurring purine nucleoside in every cell. Many critical treatments such as modulating irregular heartbeat (arrhythmias), regulation of central... (Review)
Review
Adenosine is a naturally occurring purine nucleoside in every cell. Many critical treatments such as modulating irregular heartbeat (arrhythmias), regulation of central nervous system (CNS) activity and inhibiting seizural episodes can be carried out using adenosine. Despite the significant potential therapeutic impact of adenosine and its derivatives, the severe side effects caused by their systemic administration have significantly limited their clinical use. In addition, due to adenosine's extremely short half-life in human blood (<10 s), there is an unmet need for sustained delivery systems to enhance efficacy and reduce side effects. In this article, various adenosine delivery techniques, including encapsulation into biodegradable polymers, cell-based delivery, implantable biomaterials and mechanical-based delivery systems, are critically reviewed and the existing challenges are highlighted.
Topics: Adenosine; Animals; Drug Delivery Systems; Drug Design; Half-Life; Humans; Polymers
PubMed: 26453156
DOI: 10.3109/1061186X.2015.1058803 -
Nature Methods Dec 2019N-methyladenosine (mA) is a widespread RNA modification that influences nearly every aspect of the messenger RNA lifecycle. Our understanding of mA has been facilitated...
N-methyladenosine (mA) is a widespread RNA modification that influences nearly every aspect of the messenger RNA lifecycle. Our understanding of mA has been facilitated by the development of global mA mapping methods, which use antibodies to immunoprecipitate methylated RNA. However, these methods have several limitations, including high input RNA requirements and cross-reactivity to other RNA modifications. Here, we present DART-seq (deamination adjacent to RNA modification targets), an antibody-free method for detecting mA sites. In DART-seq, the cytidine deaminase APOBEC1 is fused to the mA-binding YTH domain. APOBEC1-YTH expression in cells induces C-to-U deamination at sites adjacent to mA residues, which are detected using standard RNA-seq. DART-seq identifies thousands of mA sites in cells from as little as 10 ng of total RNA and can detect mA accumulation in cells over time. Additionally, we use long-read DART-seq to gain insights into mA distribution along the length of individual transcripts.
Topics: APOBEC-1 Deaminase; Adenosine; Base Sequence; Deamination; HEK293 Cells; Humans; Transcriptome
PubMed: 31548708
DOI: 10.1038/s41592-019-0570-0 -
Molecular Cell Jul 2016N(6)-methyladenosine (m(6)A) is a prevalent, reversible chemical modification of functional RNAs and is important for central events in biology. The core m(6)A writers...
N(6)-methyladenosine (m(6)A) is a prevalent, reversible chemical modification of functional RNAs and is important for central events in biology. The core m(6)A writers are Mettl3 and Mettl14, which both contain methyltransferase domains. How Mettl3 and Mettl14 cooperate to catalyze methylation of adenosines has remained elusive. We present crystal structures of the complex of Mettl3/Mettl14 methyltransferase domains in apo form as well as with bound S-adenosylmethionine (SAM) or S-adenosylhomocysteine (SAH) in the catalytic site. We determine that the heterodimeric complex of methyltransferase domains, combined with CCCH motifs, constitutes the minimally required regions for creating m(6)A modifications in vitro. We also show that Mettl3 is the catalytically active subunit, while Mettl14 plays a structural role critical for substrate recognition. Our model provides a molecular explanation for why certain mutations of Mettl3 and Mettl14 lead to impaired function of the methyltransferase complex.
Topics: Adenosine; Allosteric Regulation; Binding Sites; Catalytic Domain; HEK293 Cells; Humans; Methylation; Methyltransferases; Models, Molecular; Mutation; Protein Binding; Protein Conformation; RNA; S-Adenosylhomocysteine; S-Adenosylmethionine; Structure-Activity Relationship
PubMed: 27373337
DOI: 10.1016/j.molcel.2016.05.041 -
Nature Medicine Nov 2017N-methyladenosine (mA) is an abundant nucleotide modification in mRNA that is required for the differentiation of mouse embryonic stem cells. However, it remains unknown...
N-methyladenosine (mA) is an abundant nucleotide modification in mRNA that is required for the differentiation of mouse embryonic stem cells. However, it remains unknown whether the mA modification controls the differentiation of normal and/or malignant myeloid hematopoietic cells. Here we show that shRNA-mediated depletion of the mA-forming enzyme METTL3 in human hematopoietic stem/progenitor cells (HSPCs) promotes cell differentiation, coupled with reduced cell proliferation. Conversely, overexpression of wild-type METTL3, but not of a catalytically inactive form of METTL3, inhibits cell differentiation and increases cell growth. METTL3 mRNA and protein are expressed more abundantly in acute myeloid leukemia (AML) cells than in healthy HSPCs or other types of tumor cells. Furthermore, METTL3 depletion in human myeloid leukemia cell lines induces cell differentiation and apoptosis and delays leukemia progression in recipient mice in vivo. Single-nucleotide-resolution mapping of mA coupled with ribosome profiling reveals that mA promotes the translation of c-MYC, BCL2 and PTEN mRNAs in the human acute myeloid leukemia MOLM-13 cell line. Moreover, loss of METTL3 leads to increased levels of phosphorylated AKT, which contributes to the differentiation-promoting effects of METTL3 depletion. Overall, these results provide a rationale for the therapeutic targeting of METTL3 in myeloid leukemia.
Topics: Adenosine; Bone Marrow Cells; Cell Differentiation; Cells, Cultured; Clustered Regularly Interspaced Short Palindromic Repeats; Humans; Leukemia, Myeloid, Acute; Methyltransferases; Tumor Cells, Cultured
PubMed: 28920958
DOI: 10.1038/nm.4416 -
Genes Dec 2022Chronic lung diseases are highly prevalent worldwide and cause significant mortality. Lung cancer is the end stage of many chronic lung diseases. RNA epigenetics can... (Review)
Review
Chronic lung diseases are highly prevalent worldwide and cause significant mortality. Lung cancer is the end stage of many chronic lung diseases. RNA epigenetics can dynamically modulate gene expression and decide cell fate. Recently, studies have confirmed that RNA epigenetics plays a crucial role in the developing of chronic lung diseases. Further exploration of the underlying mechanisms of RNA epigenetics in chronic lung diseases, including lung cancer, may lead to a better understanding of the diseases and promote the development of new biomarkers and therapeutic strategies. This article reviews basic information on RNA modifications, including methylation of adenosine (mA), methylation of adenosine (mA), -methylguanosine (mG), 5-methylcytosine (mC), 2'O-methylation (2'-O-Me or Nm), pseudouridine (5-ribosyl uracil or Ψ), and adenosine to inosine RNA editing (A-to-I editing). We then show how they relate to different types of lung disease. This paper hopes to summarize the mechanisms of RNA modification in chronic lung disease and finds a new way to develop early diagnosis and treatment of chronic lung disease.
Topics: Humans; RNA; Methylation; Epigenesis, Genetic; Adenosine; Lung Neoplasms
PubMed: 36553648
DOI: 10.3390/genes13122381 -
Clinical and Translational Medicine Jun 2024Dysregulated RNA modifications, stemming from the aberrant expression and/or malfunction of RNA modification regulators operating through various pathways, play pivotal... (Review)
Review
Dysregulated RNA modifications, stemming from the aberrant expression and/or malfunction of RNA modification regulators operating through various pathways, play pivotal roles in driving the progression of haematological malignancies. Among RNA modifications, N-methyladenosine (mA) RNA modification, the most abundant internal mRNA modification, stands out as the most extensively studied modification. This prominence underscores the crucial role of the layer of epitranscriptomic regulation in controlling haematopoietic cell fate and therefore the development of haematological malignancies. Additionally, other RNA modifications (non-mA RNA modifications) have gained increasing attention for their essential roles in haematological malignancies. Although the roles of the mA modification machinery in haematopoietic malignancies have been well reviewed thus far, such reviews are lacking for non-mA RNA modifications. In this review, we mainly focus on the roles and implications of non-mA RNA modifications, including N-acetylcytidine, pseudouridylation, 5-methylcytosine, adenosine to inosine editing, 2'-O-methylation, N-methyladenosine and N-methylguanosine in haematopoietic malignancies. We summarise the regulatory enzymes and cellular functions of non-mA RNA modifications, followed by the discussions of the recent studies on the biological roles and underlying mechanisms of non-mA RNA modifications in haematological malignancies. We also highlight the potential of therapeutically targeting dysregulated non-mA modifiers in blood cancer.
Topics: Humans; Hematologic Neoplasms; RNA Processing, Post-Transcriptional; RNA; Adenosine
PubMed: 38880983
DOI: 10.1002/ctm2.1666 -
American Journal of Physiology. Cell... Jun 2022Fibroblasts play an important role in the pathogenic mechanisms of several socially significant diseases, including pulmonary and cardiovascular fibrosis, liver... (Review)
Review
Fibroblasts play an important role in the pathogenic mechanisms of several socially significant diseases, including pulmonary and cardiovascular fibrosis, liver cirrhosis, systemic sclerosis, progressive kidney disease. The alterations of the epitranscriptome, including more than 170 distinct posttranscriptional RNA modifications or editing events, justified their investigation as an important modulator of fibrosis. Recent development of high-throughput methods allows the identification of RNA modification sites and their mechanistic aspect in the fibrosis development. The most common RNA modification is methylation of N-adenosine deposited by the mA methyltransferase complex (METTL3/14/16, WTAP, KIAA1429, and RBM15/15B), erased by demethylases (FTO and ALKBH5), and recognized by binding proteins (e.g., YTHDF1/2/3, YTHDC1/2, IGF2BP1/2/3, etc.). Adenosine to inosine (A-to-I) RNA editing is another abundant editing event converting adenosine to inosine in double-stranded RNA regions through the action of the adenosine deaminase (ADAR) proteins. Last but not least, 5-methylcytosine (mC) regulates the stability and translation of mRNAs. All those RNA modifications have been observed in mRNA as well as the noncoding regions of pre-mRNA and noncoding RNAs (ncRNAs) and demonstrated to be involved in fibrosis in different cellular and animal models. This Mini-Review focuses on the latest research on epitranscriptomic marks related to fibroblast biology and fibrosis as well as elucidates the future research directions in this context.
Topics: Adenosine; Animals; Fibroblasts; Fibrosis; Inosine; RNA; RNA, Messenger
PubMed: 35508185
DOI: 10.1152/ajpcell.00121.2022 -
Current Opinion in Neurobiology Jun 2017Slow wave activity (SWA) during slow wave sleep (SWS) is the best indicator of the sleep homeostasis. The intensity of the SWA observed during SWS that follows prolonged... (Review)
Review
Slow wave activity (SWA) during slow wave sleep (SWS) is the best indicator of the sleep homeostasis. The intensity of the SWA observed during SWS that follows prolonged waking is directly correlated with the duration of prior waking and its intensity decays during SWS suggesting a buildup and a resolution of sleep need. This sleep-homeostasis related SWA results from a buildup and decay of extracellular adenosine that acts at neuronal adenosine A1 receptors to facilitate SWA and is metabolized by adenosine kinase found in glia. This local neuronal-glial circuit for homeostatic SWA is primarily under the requisite control of two genes, the Adora1 and Adk, encoding the responsible adenosine receptor and adenosine's highest affinity metabolizing enzyme.
Topics: Adenosine; Homeostasis; Humans; Neuroglia; Neurons; Sleep
PubMed: 28633050
DOI: 10.1016/j.conb.2017.05.015