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Transfer RNAs (tRNAs) contain dozens of chemical modifications. These modifications are critical for maintaining tRNA tertiary structure and optimizing protein synthesis. Here we advance the use of Nanopore direct RNA-sequencing (DRS) to investigate the synergy between modifications that are known to stabilize tRNA structure. We sequenced the 42 cytosolic tRNA isoacceptors from wild-type yeast and five tRNA-modifying enzyme knockout mutants. These data permitted comprehensive analysis of three neighboring and conserved modifications in T-loops: 5-methyluridine (m5U54), pseudouridine (Ψ55), and 1-methyladenosine (m1A58). Our results were validated using direct measurements of chemical modifications by mass spectrometry. We observed concerted T-loop modification circuits-the potent influence of Ψ55 for subsequent m1A58 modification on more tRNA isoacceptors than previously observed. Growing cells under nutrient depleted conditions also revealed a novel condition-specific increase in m1A58 modification on some tRNAs. A global and isoacceptor-specific classification strategy was developed to predict the status of T-loop modifications from a user-input tRNA DRS dataset, applicable to other conditions and tRNAs in other organisms. These advancements demonstrate how orthogonal technologies combined with genetics enable precise detection of modification landscapes of individual, full-length tRNAs, at transcriptome-scale., (© The Author(s) 2024. Published by Oxford University Press on behalf of Nucleic Acids Research.)

RNA editing is a highly conserved process. Adenosine deaminase acting on RNA (ADAR) mediated deamination of adenosine (A-to-I editing) is associated with human disease and immune checkpoint control. Functional implications of A-to-I editing are currently of broad interest to academic and industrial research as underscored by the fast-growing number of clinical studies applying base editors as therapeutic tools. Analyzing the dynamics of A-to-I editing, in a biological or therapeutic context, requires the sensitive detection of differential A-to-I editing, a currently unmet need. We introduce the local differential editing index (LoDEI) to detect differential A-to-I editing in RNA-seq datasets using a sliding-window approach coupled with an empirical q value calculation that detects more A-to-I editing sites at the same false-discovery rate compared to existing methods. LoDEI is validated on known and novel datasets revealing that the oncogene MYCN increases and that a specific small non-coding RNA reduces A-to-I editing., (© 2024. The Author(s).)

Objective: This study aims to identify m6A methylation-related and immune cell-related key genes with diagnostic potential for heart failure (HF) by leveraging various bioinformatics techniques., Methods: The GSE116250 and GSE141910 datasets were sourced from the Gene Expression Omnibus (GEO) database. Correlation analysis was conducted between differentially expressed genes (DEGs) in HF and control groups, alongside differential m6A regulatory factors, to identify m6A-related DEGs (m6A-DEGs). Subsequently, candidate genes were narrowed down by intersecting key module genes derived from weighted gene co-expression network analysis (WGCNA) with m6A-DEGs. Key genes were then identified through the Least Absolute Shrinkage and Selection Operator (LASSO) analysis. Correlation analyses between key genes and differentially expressed immune cells were performed, followed by the validation of key gene expression levels in public datasets. To ensure clinical applicability, five pairs of blood samples were collected for quantitative real-time fluorescence PCR (qRT-PCR) validation., Results: A total of 93 m6A-DEGs were identified (|COR| > 0.6, P < 0.05), and five key genes (LACTB2, NAMPT, SCAMP5, HBA1, and PRKAR2A) were selected for further analysis. Correlation analysis revealed that differential immune cells were negatively associated with the expression of LACTB2, NAMPT, and PRKAR2A (P < 0.05), while positively correlated with SCAMP5 and HBA1 (P < 0.05). Subsequent expression validation confirmed significant differences in key gene expression between the HF and control groups, with consistent expression trends observed across both training and validation sets. The expression trends of LACTB2, PRKAR2A, and HBA1 in blood samples from the qRT-PCR assay aligned with the results derived from public databases., Conclusion: This study successfully identified five m6A methylation-related key genes with diagnostic significance, providing a theoretical foundation for further exploration of m6A methylation's molecular mechanisms in HF., (© 2024. The Author(s).)

The most common epigenetic modification of messenger RNAs (mRNAs) is N6-methyladenosine (m6A), which is mainly located near the 3' untranslated region of mRNAs, near the stop codons, and within internal exons. The biological effect of m6A is dynamically modulated by methyltransferases (writers), demethylases (erasers), and m6A-binding proteins (readers). By controlling post-transcriptional gene expression, m6A has a significant impact on numerous biological functions, including RNA transcription, translation, splicing, transport, and degradation. Hence, m6A influences various physiological and pathological processes, such as spermatogenesis, oogenesis, embryogenesis, placental function, and human reproductive system diseases. During gametogenesis and embryogenesis, genetic material undergoes significant changes, including epigenomic modifications such as m6A. From spermatogenesis and oogenesis to the formation of an oosperm and early embryogenesis, m6A changes occur at every step. m6A abnormalities can lead to gamete abnormalities, developmental delays, impaired fertilization, and maternal-to-zygotic transition blockage. Both mice and humans with abnormal m6A modifications exhibit impaired fertility. In this review, we discuss the dynamic biological effects of m6A and its regulators on gamete and embryonic development and review the possible mechanisms of infertility caused by m6A changes. We also discuss the drugs currently used to manipulate m6A and provide prospects for the prevention and treatment of infertility at the epigenetic level., (© The Author(s) 2024. Published by Oxford University Press and Science Press on behalf of the Beijing Institute of Genomics, Chinese Academy of Sciences / China National Center for Bioinformation and Genetics Society of China.)

As the most abundant messenger RNA (mRNA) modification, N6-methyladenosine (m6A) plays a crucial role in RNA fate, impacting cellular and physiological processes in various tumor types. However, our understanding of the role of the m6A methylome in tumor heterogeneity remains limited. Herein, we collected and analyzed m6A methylomes across nine human tissues from 97 m6A sequencing (m6A-seq) and RNA sequencing (RNA-seq) samples. Our findings demonstrate that m6A exhibits different heterogeneity in most tumor tissues compared to normal tissues, which contributes to the diverse clinical outcomes in different cancer types. We also found that the cancer type-specific m6A level regulated the expression of different cancer-related genes in distinct cancer types. Utilizing a novel and reliable method called "m6A-express", we predicted m6A-regulated genes and revealed that cancer type-specific m6A-regulated genes contributed to the prognosis, tumor origin, and infiltration level of immune cells in diverse patient populations. Furthermore, we identified cell-specific m6A regulators that regulate cancer-specific m6A and constructed a regulatory network. Experimental validation was performed, confirming that the cell-specific m6A regulator CAPRIN1 controls the m6A level of TP53. Overall, our work reveals the clinical relevance of m6A in various tumor tissues and explains how such heterogeneity is established. These results further suggest the potential of m6A in cancer precision medicine for patients with different cancer types., (© The Author(s) 2024. Published by Oxford University Press and Science Press on behalf of the Beijing Institute of Genomics, Chinese Academy of Sciences / China National Center for Bioinformation and Genetics Society of China.)

Adenosine-to-inosine (A-to-I) RNA editing plays an important role in the post-transcriptional regulation of eukaryotic cell physiology. However, our understanding of the occurrence, function and regulation of A-to-I editing in bacteria remains limited. Bacterial mRNA editing is catalysed by the deaminase TadA, which was originally described to modify a single tRNA in Escherichia coli. Intriguingly, several bacterial species appear to perform A-to-I editing on more than one tRNA. Here, we provide evidence that in the human pathogen Streptococcus pyogenes, tRNA editing has expanded to an additional tRNA substrate. Using RNA sequencing, we identified more than 27 editing sites in the transcriptome of S. pyogenes SF370 and demonstrate that the adaptation of S. pyogenes TadA to a second tRNA substrate has also diversified the sequence context and recoding scope of mRNA editing. Based on the observation that editing is dynamically regulated in response to several infection-relevant stimuli, such as oxidative stress, we further investigated the underlying determinants of editing dynamics and identified mRNA stability as a key modulator of A-to-I editing. Overall, our findings reveal the presence and diversification of A-to-I editing in S. pyogenes and provide novel insights into the plasticity of the editome and its regulation in bacteria., (© The Author(s) 2024. Published by Oxford University Press on behalf of Nucleic Acids Research.)

N 6 -methyladenosine (m 6 A) is an abundant and pervasive post-transcriptional modification in eukaryotic mRNAs. AlkB homolog (ALKBH) proteins play crucial roles in RNA metabolism and translation, participating in m 6 A methylation modification to regulate plant development. However, no comprehensive investigations have been conducted on ALKBH in potato. Here, 11 StALKBH family genes were identified in potato and renamed according to BLASTP and phylogenetic analyses following the Arabidopsis genome. The characteristics, sequence structures, motif compositions, phylogenetics, chromosomal locations, synteny, and promoter cis -acting element predictions were analyzed, revealing distinct evolutionary relationships between potato and other species (tomato and Arabidopsis ). Homologous proteins were classified into seven groups depending on similar conserved domains, which implies that they possess a potentially comparable function. Moreover, the StALKBH s were ubiquitous, and their expression was examined in the various tissues of a whole potato, in which the StALKBH genes, except for StALKBH2, were most highly expressed in the stolon and flower. Multiple hormone and stress-response elements were found to be located in the promoters of the StALKBH genes. Further qRT-PCR results suggest that they may be significantly upregulated in response to phytohormones and abiotic stress (except for cold), and the expression of most of the StALKBH genes exhibited positively modulated trends. Overall, this study is the first to report a genome-wide assessment of the ALKBH family in potato, providing valuable insights into candidate gene selection and facilitating in-depth functional analyses of ALKBH-mediated m 6 A methylation mechanisms in potato.

N 6 -methyladenosine (m 6 A) is involved in most biological processes and actively participates in the regulation of reproduction. According to recent research, long non-coding RNAs (lncRNAs) and their m 6 A modifications are involved in reproductive diseases. In the present study, using m 6 A-modified RNA immunoprecipitation sequencing (m 6 A-seq), we established the m 6 A methylation transcription profiles in patients with recurrent implantation failure (RIF) for the first time. There were 1443 significantly upregulated m 6 A peaks and 425 significantly downregulated m 6 A peaks in RIF. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes pathway analyses revealed that genes associated with differentially methylated lncRNAs are involved in the p53 signalling pathway and amino acid metabolism. The competing endogenous RNA network revealed a regulatory relationship between lncRNAs, microRNAs and messenger RNAs. We verified the m 6 A methylation abundances of lncRNAs by using m 6 A-RNA immunoprecipitation (MeRIP)-real-time polymerase chain reaction. This study lays a foundation for further exploration of the potential role of m 6 A modification in the pathogenesis of RIF., (© 2024. The Author(s).)

N 6 -Methyladenosine (m 6 A) is the most prevalent mRNA modification. Its biological function primarily relies on its "Reader" protein, such as YTHDC2. Previous studies have shown that YTHDC2 downregulation is a procarcinogenic phenomenon in lung adenocarcinoma (LUAD). However, further investigation is needed to understand the molecular mechanisms of downstream genes and the associated biological phenomena following YTHDC2 downregulation. Here, we found that YTHDC2 knockout upregulated exosome content in LUAD. Following YTHDC2 knockout, the mRNA levels of OAS family members (OASs) and IFIT family members (IFITs) also decreased; and inhibition of OASs and IFITs could promote exosome content. Several m 6 A modification sites on the NT domain of OASs and the TPR12 domain of IFITs were found to increase the stability of OASs and IFITs in a YTHDC2-dependent manner. OASs and IFITs affected exosome content through target genes including RAB5A, RAB7, and RAB11A, and three arginine (R) amino acids on IFITs were critical for combination IFITs with targeted RAB mRNAs and subsequent degradation. Simultaneously, OASs degraded targeted RABs through RNAseL. Additionally, mutual bindings between OASs and IFITs were critical for their target gene degradation. Collectively, the above findings might provide a theoretical basis for the treatment of LUAD patients with low YTHDC2 expression., Competing Interests: Conflict of interest The authors declare that they have no conflicts of interest with the contents of this article., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

N 6 -methyladenosine (m 6 A) is the most prevalent internal modification of mRNA and plays an important role in regulating plant growth. However, there is still a lack of effective tools to precisely modify m 6 A sites of individual transcripts in plants. Here, programmable m 6 A editing tools are developed by combining CRISPR/dCas13(Rx) with the methyltransferase GhMTA (Targeted RNA Methylation Editor, TME) or the demethyltransferase GhALKBH10 (Targeted RNA Demethylation Editor, TDE). These editors enable efficient deposition or removal of m 6 A modifications at targeted sites of endo-transcripts GhECA1 and GhDi19 within a broad editing window ranging from 0 to 46 nt. TDE editor significantly decreases m 6 A levels by 24%-76%, while the TME editor increases m 6 A enrichment, ranging from 1.37- to 2.51-fold. Furthermore, installation and removal of m 6 A modifications play opposing roles in regulating GhECA1 and GhDi19 mRNA transcripts, which may be attributed to the fact that their m 6 A sites are located in different regions of the genes. Most importantly, targeting the GhDi19 transcript with TME editor plants results in a significant increase in root length and enhanced drought resistance. Collectively, these m 6 A editors can be applied to study the function of specific m 6 A modifications and have the potential for future applications in crop improvement., (© 2024 The Author(s). Advanced Science published by Wiley‐VCH GmbH.)

N 6 -methyladenosine (m 6 A) is the most abundant internal modification of eukaryotic mRNAs. m 6 A affects the fate of its targets in all aspects of the mRNA life cycle and has important roles in various physiological and pathophysiological processes. Aberrant m 6 A patterns have been observed in numerous cancers and appear closely linked to oncogenic phenotypes. However, most studies relied on antibody-dependent modification detection, which is known to suffer from important limitations. Novel, antibody-independent, quantitative approaches will be critical to investigate changes in the m 6 A landscape of cancers. Furthermore, pharmaceutical targeting of the m 6 A writer Methyltransferase-like 3 (METTL3) has demonstrated the potential to modulate cancer cell phenotypes. However, the enzyme also appears to be essential for the viability of healthy cells. Further refinement of therapeutic strategies is therefore needed to fully realize the potential of m 6 A-related cancer therapies., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Background: Circular RNAs (circRNAs) can influence a variety of biological functions and act as a significant role in the progression and recurrence of glioblastoma (GBM). However, few coding circRNAs have been discovered in cancer, and their role in GBM is still unknown. The aim of this study was to identify coding circRNAs and explore their potential roles in the progression and recurrence of GBM., Methods: CircSPECC1 was screened via circRNAs microarray of primary and recurrent GBM samples. To ascertain the characteristics and coding ability of circSPECC1, we conducted a number of experiments. Afterward, through in vivo and in vitro experiments, we investigated the biological functions of circSPECC1 and its encoded novel protein (SPECC1-415aa) in GBM, as well as their effects on TMZ sensitivity., Results: By analyzing primary and recurrent GBM samples via circRNAs microarray, circSPECC1 was found to be a downregulated circRNA with coding potential in recurrent GBM compared with primary GBM. CircSPECC1 suppressed the proliferation, migration, invasion, and colony formation abilities of GBM cells by encoding a new protein known as SPECC1-415aa. CircSPECC1 restored TMZ sensitivity in TMZ-resistant GBM cells by encoding the new protein SPECC1-415aa. The m 6 A reader protein IGF2BP1 can bind to circSPECC1 to promote its expression and stability. Mechanistically, SPECC1-415aa can bind to ANXA2 and competitively inhibit the binding of ANXA2 to EGFR, thus resulting in the inhibition of the phosphorylation of EGFR (Tyr845) and its downstream pathway protein AKT (Ser473). In vivo experiments showed that the overexpression of circSPECC1 could combine with TMZ to treat TMZ-resistant GBM, thereby restoring the sensitivity of TMZ-resistant GBM to TMZ., Conclusions: CircSPECC1 was downregulated in recurrent GBM compared with primary GBM. The m6A reader protein IGF2BP1 could promote the expression and stability of circSPECC1. The sequence of SPECC1-415aa, which is encoded by circSPECC1, can inhibit the binding of ANXA2 to EGFR by competitively binding to ANXA2 and inhibiting the phosphorylation of EGFR and AKT, thereby restoring the sensitivity of TMZ-resistant GBM cells to TMZ., (© 2024. The Author(s).)

N6-methyladenosine (m6A) is one of the most abundant and well-known modifications in messenger RNAs since its discovery in the 1970s. Recent studies have demonstrated that m6A is involved in various biological processes, such as alternative splicing and RNA degradation, playing an important role in a variety of diseases. To better understand the role of m6A, transcriptome-wide m6A profiling data are indispensable. In recent years, the Oxford Nanopore Technology Direct RNA Sequencing (DRS) platform has shown promise for RNA modification detection based on current disruptions measured in transcripts. However, decoding current intensity data into modification profiles remains a challenging task. Here, we introduce the m6A Transcriptome-wide Mapper (m6ATM), a novel Python-based computational pipeline that applies deep neural networks to predict m6A sites at a single-base resolution using DRS data. The m6ATM model architecture incorporates a WaveNet encoder and a dual-stream multiple-instance learning model to extract features from specific target sites and characterize the m6A epitranscriptome. For validation, m6ATM achieved an accuracy of 80% to 98% across in vitro transcription datasets containing varying m6A modification ratios and outperformed other tools in benchmarking with human cell line data. Moreover, we demonstrated the versatility of m6ATM in providing reliable stoichiometric information and used it to pinpoint PEG10 as a potential m6A target transcript in liver cancer cells. In conclusion, m6ATM is a high-performance m6A detection tool, and our results pave the way for future advancements in epitranscriptomic research., (© The Author(s) 2024. Published by Oxford University Press.)

RNA methylation adjacent to the 5' cap plays a critical role in controlling mRNA stability and protein synthesis. In trypanosomes the 5'-terminus of mRNA is protected by hypermethylated cap 4. Trypanosomes encode a cytoplasmic recapping enzyme TbCe1 which possesses an RNA kinase and guanylyltransferase activities that can convert decapped 5'-monophosphate-terminated pRNA into GpppRNA. Here, we demonstrated that the RNA kinase activity is stimulated by two orders of magnitude on a hypermethylated pRNA derived from cap 4. The N6, N6-2'-O trimethyladenosine modification on the first nucleotide was primarily accountable for enhancing both the RNA kinase and the guanylyltransferase activity of TbCe1. In contrast, N6 methyladenosine severely inhibits the guanylyltransferase activity of the mammalian capping enzyme. Furthermore, we showed that TbCmt1 cap (guanine N7) methyltransferase was localized in the cytoplasm, and its activity was also stimulated by hypermethylation at 2'-O ribose, suggesting that TbCe1 and TbCmt1 act together as a recapping enzyme to regenerate translatable mRNA from decapped mRNA. Our result establishes the functional role of cap 4 hypermethylation in recruitment and activation of mRNA recapping pathway. Methylation status at the 5'-end of transcripts could serve as a chemical landmark to selectively regulate the level of functional mRNA by recapping enzymes., (© The Author(s) 2024. Published by Oxford University Press on behalf of Nucleic Acids Research.)

Background: N6-methyladenosine (m 6 A) and 5-methylcytosine (m 5 C) play a role in modifying long non-coding RNAs (lncRNAs) implicated in tumorigenesis and progression. This study was performed to evaluate prognostic value of m 6 A- and m 5 C-related lncRNAs and develop an efficient model for prognosis prediction in cervical cancer (CC)., Methods: Using gene expression data of TCGA set, we identified m 6 A- and m 5 C-related lncRNAs. Consensus Clustering Analysis was performed for samples subtyping based on survival-related lncRNAs, followed by analyzing tumor infiltrating immune cells (TIICs). Optimal signature lncRNAs were obtained using lasso Cox regression analysis for constructing a prognostic model and a nomogram to predict prognosis., Results: We built a co-expression network of 23 m 6 A-related genes, 15 m 5 C-related genes, and 62 lncRNAs. Based on 9 m 6 A- and m 5 C-related lncRNAs significantly associated with overall survival (OS) time, two molecular subtypes were obtained, which had significantly different OS time and fractions of TIICs. A prognostic model based on six m 6 A- and m 5 C-related signature lncRNAs was constructed, which could dichotomize patients into two risk subgroups with significantly different OS time. Prognostic power of the model was successfully validated in an independent dataset. We subsequently constructed a nomogram which could accurately predict survival probabilities. Drug sensitivity analysis found preferred chemotherapeutic agents for high and low-risk patients, respectively., Conclusion: Our study reveals that m 6 A- and m 5 C-related lncRNAs are associated with prognosis and immune microenvironment of CC. The m 6 A- and m 5 C-related six-lncRNA signature may be a useful tool for survival stratification in CC and open new avenues for individualized therapies., (© 2024. The Author(s).)

Extracellular adenosine is extensively involved in regulating the tumor microenvironment. Given the disappointing results of adenosine-targeted therapy trials, personalized treatment might be necessary, tailored to the microenvironment status of individual patients. Here, we introduce the adenosine signaling score (ADO-score) model using non-negative matrix fraction identified patient subtypes using publicly available melanoma dataset, which aimed to profile adenosine signaling-related genes and construct a model to predict prognosis. We analyzed 580 malignant melanoma samples and demonstrated its robust value for prognosis. Further investigation in immune checkpoint inhibitor dataset suggests its potential as a stratified factor of immune checkpoint inhibitor efficacy. We validated the power of the ADO-score at the protein level immunofluorescence in a melanoma cohort from Xiangya Hospital. More importantly, single-cell and spatial transcriptomic data highlighted the cell-specific expression patterns of adenosine signaling-related genes and the existence of adenosine signaling-mediated crosstalk between tumor cells and immune cells in melanoma. Our study reveals a robust connection between adenosine signaling and clinical benefits in melanoma patients and proposes a universally applicable adenosine signaling model, the ADO-score, in gene expression profiles and histological sections. This model enables us to more precisely and conveniently select patients who are likely to benefit from immunotherapy., (© 2024. The Author(s).)

Fibrosis is an abnormal tissue healing process characterized by the excessive accumulation of ECM components, such as COL I and COL III, in response to tissue injury or chronic inflammation. Recent advances in epitranscriptomics have underscored the importance of m6A modification in fibrosis. m6A, the most prevalent modification in eukaryotic RNA, is catalyzed by methyltransferases (e.g., METTL3), removed by demethylases (e.g., FTO), and recognized by reader proteins (e.g., YTHDF1/2). These modifications are crucial in regulating collagen metabolism and associated diseases. Understanding the role of m6A modification in fibrosis and other collagen-related conditions holds promise for developing targeted therapies. This review highlights the latest progress in this area., (© 2024. The Author(s).)

Purpose: The current study aims to investigate the significance of N6-methyladenosine (m 6 A) methylationrelated genes in the clinical prognosis of childhood relapsed B-cell acute lymphoblastic leukemia (B-ALLL) patient., Methods: Transcriptome data and corresponding clinical data on m 6 A methylation-related genes (including 20 genes) were obtained from the Therapeutically Applicable Research To Generate Effective Treatments (TARGET) database., Results: The bone marrow (BM) samples of 134 newly diagnosed (naive) and 116 relapsed B-ALL from TARGET were enrolled in the current study. Three genes (FTO, HNRNPC, RBM15B) showed significant up-regulation in relapsed B-ALL compared with that in naive B-ALL.The three genes had a significantly worse survival (P < 0.05). The LASSO Cox regression model was used to select the most predictive genes as prognostic indicators, and YTHDC1 and FTO were identified as prognostic factors for relapsed B-ALL. Finally, the results of multivariate regression analysis showed that the risk score of m 6 A methylation-related genes was an independent prognostic factor in relapsed B-ALL (P < 0.05)., Conclusion: We found that the expression levels of m 6 A methylation-related genes were different in naive and relapsed patients with B-ALL and correlated with survival and prognosis.This implies that m 6 A methylation-related genes may be promising prognostic indicators or therapeutic targets for relapsed B-ALL., (© 2024. The Author(s).)

The progression of kidney disease varies among individuals, but a general methodology to quantify disease timelines is lacking. Particularly challenging is the task of determining the potential for recovery from acute kidney injury following various insults. Here, we report that quantitation of post-transcriptional adenosine-to-inosine (A-to-I) RNA editing offers a distinct genome-wide signature, enabling the delineation of disease trajectories in the kidney. A well-defined murine model of endotoxemia permitted the identification of the origin and extent of A-to-I editing, along with temporally discrete signatures of double-stranded RNA stress and adenosine deaminase isoform switching. We found that A-to-I editing of antizyme inhibitor 1 (AZIN1), a positive regulator of polyamine biosynthesis, serves as a particularly useful temporal landmark during endotoxemia. Our data indicate that AZIN1 A-to-I editing, triggered by preceding inflammation, primes the kidney and activates endogenous recovery mechanisms. By comparing genetically modified human cell lines and mice locked in either A-to-I-edited or uneditable states, we uncovered that AZIN1 A-to-I editing not only enhances polyamine biosynthesis but also engages glycolysis and nicotinamide biosynthesis to drive the recovery phenotype. Our findings implicate that quantifying AZIN1 A-to-I editing could potentially identify individuals who have transitioned to an endogenous recovery phase. This phase would reflect their past inflammation and indicate their potential for future recovery.

Background: Elevated extracellular matrix (ECM) accumulation is a major contributing factor to the pathogenesis of fibrotic diseases. Recent studies have indicated that N6-methyladenosine (m 6 A) RNA modification plays a pivotal role in modulating RNA stability and contribute to the initiation of various pathological conditions. Howbeit, the precise mechanism by which m 6 A influences ECM deposition remains unclear., Methods: In this study, we used hypertrophic scars (HTSs) as a paradigm to investigate ECM-related diseases. We focused on the role of ALKBH5-mediated m 6 A demethylation within the pathological progression of HTSs and examined its correlation with clinical stages. The effects of ALKBH5 ablation on ECM components were studied both in vivo and in vitro. Downstream targets of ALKBH5, along with their underlying mechanisms, were identified using integrated high-throughput analysis, RNA-binding protein immunoprecipitation and RNA pull-down assays. Furthermore, the therapeutic potential of exogenous ALKBH5 overexpression was evaluated in fibrotic scar models., Results: ALKBH5 was decreased in fibroblasts derived from HTS lesions and was negatively correlated with their clinical stages. Importantly, ablation of ALKBH5 promoted the expression of COL3A1, COL1A1, and ELN, leading to pathological deposition and reconstruction of the ECM both in vivo and in vitro. From a therapeutic perspective, the exogenous overexpression of ALKBH5 significantly inhibited abnormal collagen deposition in fibrotic scar models. As determined by integrated high-throughput analysis, key ECM components including COL3A1, COL1A1, and ELN are direct downstream targets of ALKBH5. By means of its mechanism, ALKBH5 inhibits the expression of COL3A1, COL1A1, and ELN by removing m 6 A from mRNAs, thereby decreasing their stability in a YTHDF1-dependent manner., Conclusions: Our study identified ALKBH5 as an endogenous suppressor of pathological ECM deposition, contributing to the development of a reprogrammed m6A-targeted therapy for HTSs., (© 2024 The Author(s). Clinical and Translational Medicine published by John Wiley & Sons Australia, Ltd on behalf of Shanghai Institute of Clinical Bioinformatics.)

Genetic and epigenetic alterations occur in many physiological and pathological processes. The existing knowledge regarding the association of PIWI-interacting RNAs (piRNAs) and their genetic variants on risk and progression of prostate cancer (PCa) is limited. In this study, three genome-wide association study datasets are combined, including 85,707 PCa cases and 166,247 controls, to uncover genetic variants in piRNAs. Functional investigations involved manipulating piRNA expression in cellular and mouse models to study its oncogenetic role in PCa. A specific genetic variant, rs17201241 is identified, associated with increased expression of PROPER (piRNA overexpressed in prostate cancer) in tumors and are located within the gene, conferring an increased risk and malignant progression of PCa. Mechanistically, PROPER coupled with YTHDF2 to recognize N 6 -methyladenosine (m 6 A) and facilitated RNA-binding protein interactions between EIF2S3 at 5'-untranslated region (UTR) and YTHDF2/YBX3 at 3'-UTR to promote DUSP1 circularization. This m 6 A-dependent mRNA-looping pattern enhanced DUSP1 degradation and inhibited DUSP1 translation, ultimately reducing DUSP1 expression and promoting PCa metastasis via the p38 mitogen-activated protein kinase (MAPK) signaling pathway. Inhibition of PROPER expression using antagoPROPER effectively suppressed xenograft growth, suggesting its potential as a therapeutic target. Thus, targeting piRNA PROPER-mediated genetic and epigenetic fine control is a promising strategy for the concurrent prevention and treatment of PCa., (© 2024 The Author(s). Advanced Science published by Wiley‐VCH GmbH.)

The N 6 -methyladenosine (m 6 A) modification is found in thousands of cellular mRNAs and is a critical regulator of gene expression and cellular physiology. m 6 A dysregulation contributes to several human diseases, and the m 6 A methyltransferase machinery has emerged as a promising therapeutic target. However, current methods for studying m 6 A require RNA isolation and do not provide a real-time readout of mRNA methylation in living cells. Here we present a genetically encoded m 6 A sensor (GEMS) technology, which couples a fluorescent signal with cellular mRNA methylation. GEMS detects changes in m 6 A caused by pharmacological inhibition of the m 6 A methyltransferase, giving it potential utility for drug discovery efforts. Additionally, GEMS can be programmed to achieve m 6 A-dependent delivery of custom protein payloads in cells. Thus, GEMS is a versatile platform for m 6 A sensing that provides both a simple readout for m 6 A methylation and a system for m 6 A-coupled protein expression., (© 2024. The Author(s), under exclusive licence to Springer Nature America, Inc.)

To explore the role of lncRNA m 6 A methylation modification in aqueous humour (AH) of patients with pseudoexfoliation glaucoma (PXG). Patients with open-angle PXG under surgery from June 2021 to December 2021 were selected. Age- and gender-matched patients with age-related cataract (ARC) were chosen as control. Patients underwent detailed ophthalmic examinations. 0.05-0.1 ml AH were extracted during surgery for MeRIP-Seq and RNA-Seq. Joint analysis was used to screen lncRNAs with differential m 6 A methylation modification and expression. Online software tools were used to draw lncRNA-miRNA-mRNA network (ceRNA). Expression of lncRNAs and mRNAs was confirmed using quantitative real-time PCR. A total of 4151 lncRNAs and 4386 associated m 6 A methylation modified peaks were identified in the PXG group. Similarly, 2490 lncRNAs and 2595 associated m 6 A methylation modified peaks were detected in the control. Compared to the ARC group, the PXG group had 234 hypermethylated and 402 hypomethylated m 6 A peaks, with statistically significant differences (| Fold Change (FC) |≥2, p  < 0.05). Bioinformatic analysis revealed that these differentially methylated lncRNA enriched in extracellular matrix formation, tight adhesion, TGF- β signalling pathway, AMPK signalling pathway, and MAPK signalling pathway. Joint analysis identified 10 lncRNAs with differential m 6 A methylation and expression simultaneously. Among them, the expression of ENST000000485383 and ROCK1 were confirmed downregulated in the PXG group by RT-qPCR. m 6 A methylation modification may affect the expression of lncRNA and participate in the pathogenesis of PXG through the ceRNA network. ENST000000485383-hsa miR592-ROCK1 May be a potential target pathway for further investigation in PXG m 6 A methylation.

Adenosine-to-inosine (A-to-I) RNA editing recodes the genome and confers flexibility for the organisms to adapt to the environment. It is believed that RNA recoding sites are well suited for facilitating adaptive evolution by increasing the proteomic diversity in a temporal-spatial manner. The function and essentiality of a few conserved recoding sites are recognized. However, the experimentally discovered functional sites only make up a small corner of the total sites, and there is still the need to expand the repertoire of such functional sites with bioinformatic approaches. In this study, we define a new category of RNA editing sites termed 'conserved editing with non-conserved recoding' and systematically identify such sites in Drosophila editomes, figuring out their selection pressure and signals of adaptation at inter-species and intra-species levels. Surprisingly, conserved editing sites with non-conserved recoding are not suppressed and are even slightly overrepresented in Drosophila . DNA mutations leading to such cases are also favoured during evolution, suggesting that the function of those recoding events in different species might be diverged, specialized, and maintained. Finally, structural prediction suggests that such recoding in potassium channel Shab might increase ion permeability and compensate the effect of low temperature. In conclusion, conserved editing with non-conserved recoding might be functional as well. Our study provides novel aspects in considering the adaptive evolution of RNA editing sites and meanwhile expands the candidates of functional recoding sites for future validation.

Background: Psoriasis is a common, chronic, and multifactorial inflammatory cutaneous disorder that involves genetic and epigenetic factors. N6-methyladenosine methylation (m 6 A) is the most prevalent RNA modification implicated in various diseases; however, its role in psoriasis still needs to be further explored. We aimed to explore the mechanisms underlying the effects of m 6 A in psoriasis pathogenesis, prompting new therapeutic targets., Methods: Three psoriasis-related datasets, including GSE155702, GSE109248, and GSE142582, were collected. Differentially m 6 A methylated genes (DMGs) between psoriasis lesions of psoriasis patients and healthy skin controls were identified from the GSE155702 dataset, and corresponding Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses were performed. Differentially expressed genes (DEGs) and the common DEGs between the two groups were screened from the GSE109248 and GSE142582 datasets; the expression and interactions of the m 6 A regulators were analyzed. The m 6 A levels of total RNAs and the protein expression levels of METTL3, WTAP, ALKBH5, FTO, and METTL14 in imiquimod (IMQ)-induced psoriasiform lesions were evaluated., Results: 66 significantly upregulated and 381 significantly downregulated m 6 A peaks were identified, corresponding to 414 genes which were particularly associated with cell and tissue development processes and cell cycle related items. 271 common DEGs were identified, associating with keratinocyte differentiation, epidermis development, cytokine-cytokine receptor interaction, and fatty acid metabolic processes. 15 crucial m 6 A related differentially expressed genes were obtained after the intersection of the DMGs and common DEGs, including NEU2, GALNT6, MTCL1, DOC2B, CAMK2N1, SNTB1, RNF150, CGNL1, CCDC102A, MEOX2, EEF2K, OBSCN, SLC46A2, CCDC85A, and DACH1 . In addition, we found that m 6 A methylation and these five m 6 A regulators were both upregulated in psoriatic lesions., Conclusions: It revealed that psoriasis pathophysiological processes encompass m 6 A epigenetic alterations, and that m 6 A alterations may specifically influence cell proliferation and neural regulation, and closely associated with osteoarticular involvement and metabolic syndrome in psoriasis., (© 2024 The Author(s). Published by IMR Press.)

pre-mRNA splicing is a critical feature of eukaryotic gene expression. Both cis- and trans-splicing rely on accurately recognising splice site sequences by spliceosomal U snRNAs and associated proteins. Spliceosomal snRNAs carry multiple RNA modifications with the potential to affect different stages of pre-mRNA splicing. Here, we show that the conserved U6 snRNA m6A methyltransferase METT-10 is required for accurate and efficient cis- and trans-splicing of C. elegans pre-mRNAs. The absence of METT-10 in C. elegans and METTL16 in humans primarily leads to alternative splicing at 5' splice sites with an adenosine at +4 position. In addition, METT-10 is required for splicing of weak 3' cis- and trans-splice sites. We identified a significant overlap between METT-10 and the conserved splicing factor SNRNP27K in regulating 5' splice sites with +4A. Finally, we show that editing endogenous 5' splice site +4A positions to +4U restores splicing to wild-type positions in a mett-10 mutant background, supporting a direct role for U6 snRNA m6A modification in 5' splice site recognition. We conclude that the U6 snRNA m6A modification is important for accurate and efficient pre-mRNA splicing., (© The Author(s) 2024. Published by Oxford University Press on behalf of Nucleic Acids Research.)

Bacterial gene expression is a complex process involving extensive regulatory mechanisms. Along with growing interests in this field, Nanopore Direct RNA Sequencing (DRS) provides a promising platform for rapid and comprehensive characterization of bacterial RNA biology. However, the DRS of bacterial RNA is currently deficient in the yield of mRNA-mapping reads and has yet to be exploited for transcriptome-wide RNA modification mapping. Here, we showed that pre-processing of bacterial total RNA (size selection followed by ribosomal RNA depletion and polyadenylation) guaranteed high throughputs of sequencing data and considerably increased the amount of mRNA reads. This way, complex transcriptome architectures were reconstructed for Escherichia coli and Staphylococcus aureus and extended the boundaries of 225 known E. coli operons and 89 defined S. aureus operons. Utilizing unmodified in vitro-transcribed (IVT) RNA libraries as a negative control, several Nanopore-based computational tools globally detected putative modification sites in the E. coli and S. aureus transcriptomes. Combined with Next-Generation Sequencing-based N6-methyladenosine (m6A) detection methods, 75 high-confidence m6A candidates were identified in the E. coli protein-coding transcripts, while none were detected in S. aureus. Altogether, we demonstrated the potential of Nanopore DRS in systematic and convenient transcriptome and epitranscriptome analysis., (© The Author(s) 2024. Published by Oxford University Press on behalf of Nucleic Acids Research.)

Background: Adenosine-to-inosine (A-to-I) RNA editing is a co-/post-transcriptional modification introducing A-to-G variations in RNAs. There is extensive discussion on whether the flexibility of RNA editing exerts a proteomic diversification role, or it just acts like hardwired mutations to correct the genomic allele. Eusocial insects evolved the ability to generate phenotypically differentiated individuals with the same genome, indicating the involvement of epigenetic/transcriptomic regulation., Methods: We obtained the genomes of 104 Hymenoptera insects and the transcriptomes of representative species. Comparative genomic analysis was performed to parse the evolutionary trajectory of a regulatory Ile > Met auto-recoding site in Adar gene., Results: At genome level, the pre-editing Ile codon is conserved across a node containing all eusocial hymenopterans. At RNA level, the editing events are confirmed in representative species and shows considerable condition-specificity. Compared to random expectation, the editable Ile codon avoids genomic substitutions to Met or to uneditable Ile codons, but does not avoid mutations to other unrelated amino acids., Conclusions: The flexibility of Adar auto-recoding site in Hymenoptera is selectively maintained, supporting the flexible RNA editing hypothesis. We proposed a new angle to view the adaptation of RNA editing, providing another layer to explain the great phenotypical plasticity of eusocial insects., (© 2024. The Author(s).)

Crosstalk between N 6 -methyladenosine (m 6 A) and epigenomes is crucial for gene regulation, but its regulatory directionality and disease significance remain unclear. Here, we utilize quantitative trait loci (QTLs) as genetic instruments to delineate directional maps of crosstalk between m 6 A and two epigenomic traits, DNA methylation (DNAme) and H3K27ac. We identify 47 m 6 A-to-H3K27ac and 4,733 m 6 A-to-DNAme and, in the reverse direction, 106 H3K27ac-to-m 6 A and 61,775 DNAme-to-m 6 A regulatory loci, with differential genomic location preference observed for different regulatory directions. Integrating these maps with complex diseases, we prioritize 20 genome-wide association study (GWAS) loci for neuroticism, depression, and narcolepsy in brain; 1,767 variants for asthma and expiratory flow traits in lung; and 249 for coronary artery disease, blood pressure, and pulse rate in muscle. This study establishes disease regulatory paths, such as rs3768410-DNAme-m 6 A-asthma and rs56104944-m 6 A-DNAme-hypertension, uncovering locus-specific crosstalk between m 6 A and epigenomic layers and offering insights into regulatory circuits underlying human diseases., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

Background: Long noncoding RNAs (lncRNAs) and N6-methyladenosine (m6A) modification of RNA play pivotal roles in tumorigenesis and cancer progression. However, knowledge regarding the expression patterns of m6A-related lncRNAs and their corresponding m6A regulators in prostate cancer (PCa) is limited. This study aimed to delineate the landscape of m6A-related lncRNAs, develop a predictive model, and identify the critical m6A regulators of prognostic lncRNAs in PCa., Methods: Clinical and transcriptome data of PCa patients were downloaded from The Cancer Genome Atlas (TCGA) database. Prognostic m6A-related lncRNAs were subsequently identified through Pearson correlation and univariate Cox regression analyses. The prognostic lncRNAs were clustered into two groups by consensus clustering analysis, and a risk signature model was constructed using least absolute shrinkage and selection operator (LASSO) regression analysis of the lncRNAs. This model was evaluated using survival, clinicopathological, and immunological analyses. Furthermore, based on the constructed lncRNA-m6A regulatory network and RT-qPCR results, RBM15 was identified as a critical regulator of m6A-related lncRNAs. The biological roles of RBM15 in PCa were explored through bioinformatics analysis and biological experiments., Results: Thirty-four prognostic m6A-related lncRNAs were identified and categorized into two clusters with different expression patterns and survival outcomes in PCa patients. Seven m6A lncRNAs (AC105345.1, AL354989.1, AC138028.4, AC022211.1, AC020558.2, AC004076.2, and LINC02666) were selected to construct a risk signature with robust predictive ability for overall survival and were correlated with clinicopathological characteristics and the immune microenvironment of PCa patients. Among them, LINC02666 and AC022211.1 were regulated by RBM15. In addition, RBM15 expression correlated with PCa progression, survival, and the immune response. Patients with elevated RBM15 expression were more susceptible to the drug AMG-232. Moreover, silencing RBM15 decreased the viability of PCa cells and promoted apoptosis., Conclusion: RBM15 is involved in the regulation of prognostic lncRNAs in the risk signature and has a robust predictive ability for PCa, making it a promising biomarker in PCa., (© 2024. The Author(s).)

The resistance of cancer cells to treatment significantly impedes the success of therapy, leading to the recurrence of various types of cancers. Understanding the specific mechanisms of therapy resistance may offer novel approaches for alleviating drug resistance in cancer. Recent research has shown a reciprocal relationship between circular RNAs (circRNAs) and N6-methyladenosine (m6A) modification, and their interaction can affect the resistance and sensitivity of cancer therapy. This review aims to summarize the latest developments in the m6A modification of circRNAs and their importance in regulating therapy resistance in cancer. Furthermore, we explore their mutual interaction and exact mechanisms and provide insights into potential future approaches for reversing cancer resistance., (© 2024. The Author(s).)

Adenosine-to-inosine (A-to-I) RNA editing recodes the genetic information. Apart from diversifying the proteome, another tempting advantage of RNA recoding is to correct deleterious DNA mutation and restore ancestral allele. Solid evidences for beneficial restorative editing are very rare in animals. By searching for "convergent recoding" under a phylogenetic context, we proposed this term for judging the potential restorative functions of particular editing site. For the well-known mammalian Gln>Arg (Q>R) recoding site, its ancestral state in vertebrate genomes was the pre-editing Gln, and all 470 available mammalian genomes strictly avoid other three equivalent ways to achieve Arg in protein. The absence of convergent recoding from His>Arg, or synonymous mutations on Gln codons, could be attributed to the strong maintenance on editing motif and structure, but the absence of direct A-to-G mutation is extremely unexpected. With similar ideas, we found cases of convergent recoding in Drosophila genus, reducing the possibility of their restorative function. In summary, we defined an interesting scenario of convergent recoding, the occurrence of which could be used as preliminary judgements for whether a recoding site has a sole restorative role. Our work provides novel insights to the natural selection and evolution of RNA editing., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

N 6 -methyladenosine (m 6 A) is the most abundant internal modification of mRNAs in eukaryotes. Numerous studies have shown that m 6 A plays key roles in many biological and pathophysiological processes, including fertility. The factors involved in m 6 A-dependent mRNA regulation include writers, which deposit the m 6 A mark, erasers, which remove it, and readers, which bind to m 6 A-modified transcripts and mediate the regulation of mRNA fate. Many of these proteins are highly expressed in the germ cells of mammals, and some have been linked to fertility disorders in human patients. In this review, we summarise recent findings on the important roles played by proteins involved in m 6 A biology in mammalian gametogenesis and fertility. Continued study of the m 6 A pathway in the mammalian germline will shed further light on the importance of epitranscriptomics in reproduction and may lead to effective treatment of human fertility disorders., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Helicobacter pylori (H. pylori) infection is the primary risk factor for the pathogenesis of gastric cancer (GC). N6-methyladenosine (m 6 A) plays pivotal roles in mRNA metabolism and hnRNPA2B1 as an m 6 A reader is shown to exert m 6 A-dependent mRNA stabilization in cancer. This study aims to explore the role of hnRNPA2B1 in H. pylori-associated GC and its novel molecular mechanism. Multiple datasets and tissue microarray are utilized for assessing hnRNPA2B1 expression in response to H. pylori infection and its clinical prognosis in patients with GC. The roles of hnRNPA2B1 are investigated through a variety of techniques including glucose metabolism analysis, m 6 A-epitranscriptomic microarray, Ribo-seq, polysome profiling, RIP-seq. In addition, hnRNPA2B1 interaction with poly(A) binding protein cytoplasmic 1 (PABPC1) is validated using mass spectrometry and co-IP. These results show that hnRNPA2B1 is upregulated in GC and correlated with poor prognosis. H. pylori infection induces hnRNPA2B1 upregulation through recruiting NF-κB to its promoter. Intriguingly, cytoplasm-anchored hnRNPA2B1 coordinated PABPC1 to stabilize its relationship with cap-binding eIF4F complex, which facilitated the translation of CIP2A, DLAT and GPX1 independent of m 6 A modification. In summary, hnRNPA2B1 facilitates the non-m 6 A translation of epigenetic mRNAs in GC progression by interacting with PABPC1-eIF4F complex and predicts poor prognosis for patients with GC., (© 2024 The Author(s). Advanced Science published by Wiley‐VCH GmbH.)

The RNA modification of N 6 -methyladenosine (m 6 A) controls many aspects of RNA function that impact biological processes, including viral infection. In this review, we highlight recent work that shapes our current understanding of the diverse mechanisms by which m 6 A can regulate viral infection by acting on viral or cellular mRNA molecules. We focus on emerging concepts and understanding, including how viral infection alters the localization and function of m 6 A machinery proteins, how m 6 A regulates antiviral innate immunity, and the multiple roles of m 6 A in regulating specific viral infections. We also summarize the recent studies on m 6 A during SARS-CoV-2 infection, focusing on points of convergence and divergence. Ultimately, this review provides a snapshot of the latest research on m 6 A during viral infection., Competing Interests: Declaration of Competing Interest None., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

The conversion of Adenosine (A) to Inosine (I), by Adenosine Deaminases Acting on RNA or ADARs, is an essential post-transcriptional modification that contributes to proteome diversity and regulation in metazoans including humans. In addition to its transcriptome-regulating role, ADARs also play a major part in immune response to viral infection, where an interferon response activates interferon-stimulated genes, such as ADARp150, in turn dynamically regulating host-virus interactions. A previous report has shown that infection from reoviruses, despite strong activation of ADARp150, does not influence the editing of some of the major known editing targets, while likely editing others, suggesting a potentially nuanced editing pattern that may depend on different factors. However, the results were based on a handful of selected editing sites and did not cover the entire transcriptome. Thus, to determine whether and how reovirus infection specifically affects host ADAR editing patterns, we analyzed a publicly available deep-sequenced RNA-seq dataset, from murine fibroblasts infected with wild-type and mutant reovirus strains that allowed us to examine changes in editing patterns on a transcriptome-wide scale. To the best of our knowledge, this is the first transcriptome-wide report on host editing changes after reovirus infection. Our results demonstrate that reovirus infection induces unique nuanced editing changes in the host, including introducing sites uniquely edited in infected samples. Genes with edited sites are overrepresented in pathways related to immune regulation, cellular signaling, metabolism, and growth. Moreover, a shift in editing targets has also been observed, where the same genes are edited in infection and control conditions but at different sites, or where the editing rate is increased for some and decreased for other differential targets, supporting the hypothesis of dynamic and condition-specific editing by ADARs., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Altered RNA modification patterns and dysregulated expression of epitranscriptomic machinery proteins (EMPs) have been causatively correlated with several diseases. Modulation of EMP gene expression has shown promise in reversing disease-associated phenotypes, making EMPs attractive therapeutic targets. Various therapeutic strategies, including small-molecule modulators, proteolysis-targeting chimeras, and molecular tools for site-specific engineering of RNA modifications, have been introduced to modulate EMPs and RNA modifications themselves and are currently being investigated to enrich the physician's armamentarium. At the forefront of research are small-molecule inhibitors of the key players involved in the N 6 -methyladenosine RNA modification, with an inhibitor of methyltransferase 3 in clinical trials. Preclinical studies have also demonstrated proof-of-concept for the other approaches, raising expectations for this exciting new frontier of therapy., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Ltd.. All rights reserved.)

N 6 -methyladenosine (m 6 A) has been established as the most prevalent chemical modification in message RNA (mRNA), playing an essential role in determining the fate of RNA molecules. Dysregulation of m 6 A has been revealed to lead to abnormal physiological conditions and cause various types of human diseases. Recent studies have delineated the genetic regulatory maps for m 6 A methylation by mapping the quantitative trait loci of m 6 A (m 6 A-QTLs), thereby building up the regulatory circuits linking genetic variants, m 6 A, and human complex traits. Here, we review the recent discoveries concerning the genetic regulatory maps of m 6 A, describing the methodological and technical details of m 6 A-QTL identification, and introducing the key findings of the cis- and trans-acting drivers of m 6 A. We further delve into the tissue- and ethnicity-specificity of m 6 A-QTL, the association with other molecular phenotypes in light of genetic regulation, the regulators underlying m 6 A genetics, and importantly, the functional roles of m 6 A in mediating human complex diseases. Lastly, we discuss potential research avenues that can accelerate the translation of m 6 A genetics studies toward the development of therapies for human genetic diseases., (© 2024. Science China Press.)

Background: This study aims to clarify the N6-methyladenosine (m6A) modification of LINC01006, which is involved in migration, invasion and proliferation of non-small cell lung cancer (NSCLC)., Materials and Methods: LINC01006 and METTL3 expressions were analyzed in TCGA-LUAD cohort. Colony formation assay, wound-healing assay and transwell assay were performed to evaluate the ability of colony formation, migration and invasion. Q-PCR and western blot analysis determined gene expressions. M6A-RNA immunoprecipitation and m6A quantification assay were used to evaluate m6A modification. qChIP assay was used to validate transcriptional target. Luciferase assay validated the miRNA targets and transcriptional targets. In-situ xenograft model were included to evaluate tumor proliferation in vivo., Results: LINC01006 and METTL3 expressions were elevated in NSCLC cells and tissues. LINC01006 promoted the migration and invasion of NSCLC via epithelial - mesenchymal transition (EMT). The expression of LINC01006 was positively correlated to the expression of METTL3. METTL3 promoted tumor formation and proliferation in the in-situ xenograft model of NSCLC. The expression of LINC01006 was increased by METTL3 via m6A modification. c-MYC directly induced METTL3. Both c-MYC and LINC01006 were commonly targeted by miR-34a/b/c and miR-2682, and thereby c-MYC/METTL3/LINC01006 formed a positive feedback loop through miRNA targets in NSCLC., Conclusions: LINC01006 is an oncogenic lncRNA, which induces migration, invasion and proliferation of NSCLC. METTL3 increases LINC01006 expression through stabilizing LINC01006 mRNA. c-MYC, as a transcription factor, activates METTL3, which results in an elevated level of LINC01006. c-MYC, METTL3 and LINC01006 form a positive feedback loop through multiple miRNA targets in NSCLC., Competing Interests: Conflicts of interest The authors have no relevant financial or non-financial interests to disclose., (© 2023 The Authors. Published by Elsevier B.V. on behalf of Chang Gung University. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).)

N 6 -methyladenosine (m 6 A) is the most abundant modification to mRNAs. Loss-of-function studies of main m 6 A regulators have indicated the role of m 6 A in pre-mRNA splicing. Recent studies have reported the role of splicing in preventing m 6 A deposition. Understanding the interplay between m 6 A and mRNA splicing holds the potential to clarify the significance of these fundamental molecular mechanisms in cell development and function, thereby shedding light on their involvement in the pathogenesis of myriad diseases., Competing Interests: Declaration of Competing Interest There is no competing interest related to the submitted manuscript., (Copyright © 2024 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Metastasis is the primary cause of cancer-related death, with the dissemination and colonization of primary tumor cells at the metastatic site facilitated by various molecules and complex pathways. Understanding the biological mechanisms underlying the metastatic process is critical for the development of effective interventions. Several epigenetic modifications have been identified that play critical roles in regulating cancer metastasis. This review aims to provide a comprehensive summary of recent advances in understanding the role of epigenetic modifiers, including histone modifications, DNA methylation, non-coding RNAs, enhancer reprogramming, chromatin accessibility, and N6-methyladenosine, in metastasis-associated processes, such as epithelial-mesenchymal transition (EMT), cancer cell migration, and invasion. In particular, this review provides a detailed and in-depth description of the role of crosstalk between epigenetic regulators in tumor metastasis. Additionally, we explored the potential and limitations of epigenetics-related target molecules in the diagnosis, treatment, and prognosis of cancer metastasis.

N6-methyladenosine (m$^{6}$A) is a widely-studied methylation to messenger RNAs, which has been linked to diverse cellular processes and human diseases. Numerous databases that collate m$^{6}$A profiles of distinct cell types have been created to facilitate quick and easy mining of m$^{6}$A signatures associated with cell-specific phenotypes. However, these databases contain inherent complexities that have not been explicitly reported, which may lead to inaccurate identification and interpretation of m$^{6}$A-associated biology by end-users who are unaware of them. Here, we review various m$^{6}$A-related databases, and highlight several critical matters. In particular, differences in peak-calling pipelines across databases drive substantial variability in both peak number and coordinates with only moderate reproducibility, and the inclusion of peak calls from early m$^{6}$A sequencing protocols may lead to the reporting of false positives or negatives. The awareness of these matters will help end-users avoid the inclusion of potentially unreliable data in their studies and better utilize m$^{6}$A databases to derive biologically meaningful results., (© The Author(s) 2024. Published by Oxford University Press.)

Breast cancer (BC) is the most prevalent cancer in women globally. The tumor microenvironment (TME), comprising epithelial tumor cells and stromal elements, is vital for breast tumor development. N6-methyladenosine (m6A) modification plays a key role in RNA metabolism, influencing its various aspects such as stability and translation. There is a notable link between m6A methylation and immune cells in the TME, although this relationship is complex and not fully deciphered. In this research, BC expression and clinicopathological data from TCGA were scrutinized to assess expression profiles, mutations, and CNVs of 31 m6A genes and immune microenvironment-related genes, examining their correlations, functions, and prognostic impacts. Lasso and Cox regression identified prognostic genes for constructing a nomogram. Single-cell analyses mapped the distribution and patterns of these genes in BC cell development. We investigated associations between gene-derived risk scores and factors like immune infiltration, TME, checkpoints, TMB, CSC indices, and drug response. As a complement to computational analyses, in vitro experiments were conducted to confirm these expression patterns. We included 31 m6A regulatory genes and discovered a correlation between these genes and the extent of immune cell infiltration. Subsequently, a 7-gene risk score was generated, encompassing HSPA2, TAP1, ULBP2, CXCL1, RBP1, STC2, and FLT3. It was observed that the low-risk group exhibited better overall survival (OS) in BC, with higher immune scores but lower tumor mutational burden (TMB) and cancer stem cell (CSC) indices, as well as lower IC50 values for commonly used drugs. To enhance clinical applicability, age and stage were incorporated into the risk score, and a more comprehensive nomogram was constructed to predict OS. This nomogram was validated and demonstrated good predictive performance, with area under the curve (AUC) values for 1-year, 3-year, and 5-year OS being 0.848, 0.807, and 0.759, respectively. Our findings highlight the profound impact of prognostic-related genes on BC immune response and prognostic outcomes, suggesting that modulation of the m6A-immune pathway could offer new avenues for personalized BC treatment and potentially improve clinical outcomes., (© 2024. The Author(s).)

Adenosine-to-inosine (A-to-I) RNA editing is an important post-transcriptional modification mediated by the adenosine deaminases acting on RNA (ADAR) family of enzymes, expanding the transcriptome by altering selected nucleotides A to I in RNA molecules. Recently, A-to-I editing has been explored for correcting disease-causing mutations in RNA using therapeutic guide oligonucleotides to direct ADAR editing at specific sites. Humans have two active ADARs whose preferences and specificities are not well understood. To investigate their substrate specificity, we introduced hADAR1 and hADAR2, respectively, into Schizosaccharomyces pombe ( S. pombe ), which lacks endogenous ADARs, and evaluated their editing activities in vivo. Using transcriptome sequencing of S. pombe cultured at optimal growth temperature (30 °C), we identified 483 A-to-I high-confident editing sites for hADAR1 and 404 for hADAR2, compared with the non-editing wild-type control strain. However, these sites were mostly divergent between hADAR1 and hADAR2-expressing strains, sharing 33 common sites that are less than 9% for each strain. Their differential specificity for substrates was attributed to their differential preference for neighboring sequences of editing sites. We found that at the -3-position relative to the editing site, hADAR1 exhibits a tendency toward T, whereas hADAR2 leans toward A. Additionally, when varying the growth temperature for hADAR1- and hADAR2-expressing strains, we observed increased editing sites for them at both 20 and 35 °C, compared with them growing at 30 °C. However, we did not observe a significant shift in hADAR1 and hADAR2's preference for neighboring sequences across three temperatures. The vast changes in RNA editing sites at lower and higher temperatures were also observed for hADAR2 previously in budding yeast, which was likely due to the influence of RNA folding at these different temperatures, among many other factors. We noticed examples of longer lengths of dsRNA around the editing sites that induced editing at 20 or 35 °C but were absent at the other two temperature conditions. We found genes' functions can be greatly affected by editing of their transcripts, for which over 50% of RNA editing sites for both hADAR1 and hADAR2 in S. pombe were in coding sequences (CDS), with more than 60% of them resulting in amino acid changes in protein products. This study revealed the extensive differences in substrate selectivity between the two active human ADARS, i.e., ADAR1 and ADAR2, and provided novel insight when utilizing the two different enzymes for in vivo treatment of human genetic diseases using the RNA editing approach.

Background: The aim of this study was to investigate the correlation between m 6 A methylation regulators and cell infiltration characteristics in tumor immune microenvironment (TIME), so as to help understand the immune mechanism of early-stage lung adenocarcinoma (LUAD)., Methods: The expression and consensus cluster analyses of m 6 A methylation regulators in early-stage LUAD were performed. The clinicopathological features, immune cell infiltration, survival and functional enrichment in different subtypes were analyzed. We also constructed a prognostic model. Clinical tissue samples were used to validate the expression of model genes through real-time polymerase chain reaction (RT-PCR). In addition, cell scratch assay and Transwell assay were also performed., Results: Expression of m 6 A methylation regulators was abnormal in early-stage LUAD. According to the consensus clustering of m 6 A methylation regulators, patients with early-stage LUAD were divided into two subtypes. Two subtypes showed different infiltration levels of immune cell and survival time. A prognostic model consisting of HNRNPC, IGF2BP1 and IGF2BP3 could be used to predict the survival of early-stage LUAD. RT-PCR results showed that HNRNPC, IGF2BP1 and IGF2BP3 were significantly up-regulated in early-stage LUAD tissues. The results of cell scratch assay and Transwell assay showed that overexpression of HNRNPC promotes the migration and invasion of NCI-H1299 cells, while knockdown HNRNPC inhibits the migration and invasion of NCI-H1299 cells., Conclusions: This work reveals that m 6 A methylation regulators may be potential biomarkers for prognosis in patients with early-stage LUAD. Our prognostic model may be of great value in predicting the prognosis of early-stage LUAD.

Background: N 6 -methyladenosine (m 6 A) modification is essential for modulating RNA processing as well as expression, particularly in the context of malignant tumour progression. However, the exploration of m 6 A modification in nasopharyngeal carcinoma (NPC) remains very limited., Methods: RNA m 6 A levels were analysed in NPC using m 6 A dot blot assay. The expression level of methyltransferase-like 14 (METTL14) within NPC tissues was analysed from public databases as well as RT-qPCR and immunohistochemistry. The influences on METTL14 expression on NPC proliferation and metastasis were explored via in vitro as well as in vivo functional assays. Targeted genes of METTL14 were screened using the m 6 A and gene expression profiling microarray data. Actinomycin D treatment and polysome analysis were used to detect the half-life and translational efficiency of ANKRD22. Flow cytometry, immunofluorescence and immunoprecipitation were used to validate the role of ANKRD22 on lipid metabolism in NPC cells. ChIP-qPCR analysis of H3K27AC signalling near the promoters of METTL14, GINS3, POLE2, PLEK2 and FERMT1 genes., Results: We revealed METTL14, in NPC, correlating with poor patient prognosis. In vitro and in vivo assays indicated METTL14 actively promoted NPC cells proliferation and metastasis. METTL14 catalysed m 6 A modification on ANKRD22 messenger ribonucleic acid (mRNA), recognized by the reader IGF2BP2, leading to increased mRNA stability and higher translational efficiency. Moreover, ANKRD22, a metabolism-related protein on mitochondria, interacted with SLC25A1 to enhance citrate transport, elevating intracellular acetyl-CoA content. This dual impact of ANKRD22 promoted lipid metabolism reprogramming and cellular lipid synthesis while upregulating the expression of genes associated with the cell cycle (GINS3 and POLE2) and the cytoskeleton (PLEK2 and FERMT1) through heightened epigenetic histone acetylation levels in the nucleus. Intriguingly, our findings highlighted elevated ANKRD22-mediated histone H3 lysine 27 acetylation (H3K27AC) signals near the METTL14 promoter, which contributes to a positive feedback loop perpetuating malignant progression in NPC., Conclusions: The identified METTL14-ANKRD22-SLC25A1 axis emerges as a promising therapeutic target for NPC, and also these molecules may serve as novel diagnostic biomarkers., (© 2024 The Author(s). Clinical and Translational Medicine published by John Wiley & Sons Australia, Ltd on behalf of Shanghai Institute of Clinical Bioinformatics.)

Nonmutational epigenetic reprogramming is a crucial mechanism contributing to the pronounced heterogeneity of prostate cancer (PCa). Among these mechanisms, N6-methyladenosine (m6A)-modified long non-coding RNAs (lncRNAs) have emerged as key players. However, the precise roles of m6A-modified lncRNAs in PCa remain to be elucidated. In this study, methylated RNA immunoprecipitation sequencing (MeRIP-seq) was conducted on primary and metastatic PCa samples, leading to the identification of 21 lncRNAs exhibiting differential methylation and expression patterns. We further established a PCa prognostic signature, named m6A-modified lncRNA score (mLs), based on 9 differential methylated lncRNAs in 4 multicenter cohorts. The high mLs score cohort exhibited a tendency for earlier biochemical recurrence (BCR) compared to the low mLs score cohort. Remarkably, the predictive performance of the mLs score surpassed that of five previously reported lncRNA-based signatures. Functional enrichment analysis underscored a negative correlation between the mLs score and lipid metabolism. Additionally, through MeRIP-qPCR, we pinpointed a hub gene, MIR210HG, which was validated through in vitro and in vivo experiments. These findings collectively illuminate the landscape of m6A-methylated lncRNAs in PCa tissue via MeRIP-seq and harness this information to prognosticate PCa outcomes using the mLs score. Furthermore, our study validates, both experimentally and mechanistically, the facilitative role of MIR210HG in driving PCa progression., (© 2023. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

N-methyladenosine (m6A) represents a prevalent RNA modification observed in colorectal cancer. Despite its abundance, the biological implications of m6A methylation on the lncRNA CARMN remain elusive in colorectal cancer, especially for mutant p53 gain-of-function. Here, we elucidate that CARMN exhibits diminished expression levels in colorectal cancer patients with mutant p53, attributed to its rich m6A methylation, which promotes cancer proliferation, invasion and metastasis in vitro and in vivo. Further investigation illustrates that ALKBH5 acts as a direct demethylase of CARMN, targeting 477 methylation sites, thereby preserving CARMN expression. However, the interaction of mutant p53 with the ALKBH5 promoter impedes its transcription, enhancing m6A methylation levels on CARMN. Subsequently, YTHDF2/YTHDF3 recognise and degrade m6A-modified CARMN. Concurrently, overexpressing CARMN significantly suppressed colorectal cancer progression in vitro and in vivo. Additionally, miR-5683 was identified as a direct downstream target of lncRNA CARMN, exerting an antitumour effect by cooperatively downregulating FGF2 expression. Our findings revealed the regulator and functional mechanism of CARMN in colorectal cancer with mutant p53, potentially offering insights into demethylation-based strategies for cancer diagnosis and therapy. The m6A methylation of CARMN that is prime for mutant p53 gain-of-function-induced malignant progression of colorectal cancer, identifying a promising approach for cancer therapy., (© 2024 The Author(s). Clinical and Translational Medicine published by John Wiley & Sons Australia, Ltd on behalf of Shanghai Institute of Clinical Bioinformatics.)

In eukaryotic RNA, N 6 -methyladenosine (m 6 A) is a prevalent form of methylation modification. The m 6 A modification process is reversible and dynamic, written by m 6 A methyltransferase complex, erased by m 6 A demethylase, and recognized by m 6 A binding proteins. Through mediating RNA stability, decay, alternative splicing, and translation processes, m 6 A modification regulates gene expression at the post-transcriptional level. Erythropoiesis is the process of hematopoietic stem cells undergoing proliferation, a series of differentiation and maturation to form red blood cells (RBCs). Thalassemia is a common monogenic disease characterized by excessive production of ineffective RBCs in the peripheral circulation, resulting in hemolytic anemia. Increasing evidence suggests that m 6 A modification plays a crucial role in erythropoiesis. In this review, we comprehensively summarize the function of m 6 A modification in erythropoiesis and further generalize the mechanism of m 6 A modification regulating ineffective erythropoiesis and fetal hemoglobin expression. The purpose is to improve the understanding of the pathogenesis of erythroid dysplasia and offer new perspectives for the diagnosis and treatment of thalassemia., (© 2024 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

N 6 -methyladenonsine (m 6 A) is the most prevalent internal modification of messenger RNA (mRNA) and plays critical roles in mRNA processing and metabolism. However, perturbation of individual m 6 A modification to reveal its function and the phenotypic effects is still lacking in plants. Here, we describe the construction and characterization of programmable m 6 A editing tools by fusing the m 6 A writers, the core catalytic domain of the MTA and MTB complex, and the AlkB homologue 5 (ALKBH5) eraser, to catalytically dead Cas13a (dCas13a) to edit individual m 6 A sites on mRNAs. We demonstrated that our m 6 A editors could efficiently and specifically deposit and remove m 6 A modifications on specific RNA transcripts in both Nicotiana benthamiana and Arabidopsis thaliana. Moreover, we found that targeting SHORT-ROOT (SHR) transcripts with a methylation editor could significantly increase its m 6 A levels with limited off-target effects and promote its degradation. This leads to a boost in plant growth with enlarged leaves and roots, increased plant height, plant biomass, and total grain weight in Arabidopsis. Collectively, these findings suggest that our programmable m 6 A editing tools can be applied to study the functions of individual m 6 A modifications in plants, and may also have potential applications for future crop improvement., (© 2024 The Authors. Plant Biotechnology Journal published by Society for Experimental Biology and The Association of Applied Biologists and John Wiley & Sons Ltd.)