An association study of m6A methylation with major depressive disorder.

OBJECTIVE: To find the relationship between N6-methyladenosine (m6A) genes and Major Depressive Disorder (MDD). METHODS: Differential expression of m6A associated genes between normal and MDD samples was initially identified. Subsequent analysis was conducted on the functions of these genes and the pathways they may affect. A diagnostic model was constructed using the expression matrix of these differential genes, and visualized using a nomogram. Simultaneously, an unsupervised classification method was employed to classify all patients based on the expression of these m6A associated genes. Following this, common differential genes among different clusters were computed. By analyzing the functions of the common differential expressed genes among clusters, the role of m6A-related genes in the pathogenesis of MDD patients was elucidated. RESULTS: Differential expression was observed in ELAVL1 and YTHDC2 between the MDD group and the control group. ELAVL1 was associated with comorbid anxiety in MDD patients. A linear regression model based on these two genes could accurately predict whether patients in the GSE98793 dataset had MDD and could provide a net benefit for clinical decision-making. Based on the expression matrix of ELAVL1 and YTHDC2, MDD patients were classified into three clusters. Among these clusters, there were 937 common differential genes. Enrichment analysis was also performed on these genes. The ssGSEA method was applied to predict the content of 23 immune cells in the GSE98793 dataset samples. The relationship between these immune cells and ELAVL1, YTHDC2, and different clusters was analyzed. CONCLUSION: Among all the m6A genes, ELAVL1 and YTHDC2 are closely associated with MDD, ELAVL1 is related to comorbid anxiety in MDD. ELAVL1 and YTHDC2 have opposite associations with immune cells in MDD.

Role of N6-methyladenosine-related lncRnas in pseudoexfoliation glaucoma.

To explore the role of lncRNA m6A 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 m6A 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 m6A methylation modified peaks were identified in the PXG group. Similarly, 2490 lncRNAs and 2595 associated m6A methylation modified peaks were detected in the control. Compared to the ARC group, the PXG group had 234 hypermethylated and 402 hypomethylated m6A 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- ß signalling pathway, AMPK signalling pathway, and MAPK signalling pathway. Joint analysis identified 10 lncRNAs with differential m6A methylation and expression simultaneously. Among them, the expression of ENST000000485383 and ROCK1 were confirmed downregulated in the PXG group by RT-qPCR. m6A 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 m6A methylation.

Direct Analysis of HIV mRNA m6A Methylation by Nanopore Sequencing.

The post-transcriptional processing and chemical modification of HIV RNA are understudied aspects of HIV virology, primarily due to the limited ability to accurately map and quantify RNA modifications. Modification-specific antibodies or modification-sensitive endonucleases coupled with short-read RNA sequencing technologies have allowed for low-resolution or limited mapping of important regulatory modifications of HIV RNA such as N6-methyladenosine (m6A). However, a high-resolution map of where these sites occur on HIV transcripts is needed for detailed mechanistic understanding. This has recently become possible with new sequencing technologies. Here, we describe the direct RNA sequencing of HIV transcripts using an Oxford Nanopore Technologies sequencer and the use of this technique to map m6A at near single nucleotide resolution. This technology also provides the ability to identify splice variants with long RNA reads and thus, can provide high-resolution RNA modification maps that distinguish between overlapping splice variants. The protocols outlined here for m6A also provide a powerful paradigm for studying any other RNA modifications that can be detected on the nanopore platform.

METTL3-mediated m6A methylation of SLC38A1 stimulates cervical cancer growth.

The objective of this study was to better characterize the role of the glutamine transporter SLC38A1 in cervical cancer and explore the underlying mechanisms. Data from public databases and clinical cervical cancer tissue samples were used to assess the expression of SLC38A1 and its prognostic significance. Immunohistochemical staining, qRT-PCR, and Western blotting were used to evaluate the expression of relevant genes and proteins. Cell viability, cell cycle, apoptosis, and intracellular glutamine content were measured using CCK-8, flow cytometry, and biochemical assays. Additionally, the RNA immunoprecipitation (RIP) assay was used to examine the impact of METTL3/IGF2BP3 on the m6A modification of the SLC38A1 3'UTR. Both cervical cancer specimens and cells showed significantly increased expression of SLC38A1 and its expression correlated with an unfavorable prognosis. Knockdown of SLC38A1 inhibited cell viability and cell cycle progression, induced apoptosis, and suppressed tumor growth in vivo. Glutaminase-1 inhibitor CB-839 reversed the effects of SLC38A1 overexpression. METTL3 promoted m6A modification of SLC38A1 and enhanced its mRNA stability through IGF2BP3 recruitment. Moreover, METTL3 silencing inhibited cell viability, cell cycle progression, intracellular glutamine content, and induced apoptosis, but these effects were reversed by SLC38A1 overexpression. In conclusion, METTL3-mediated m6A methylation of SLC38A1 stimulates cervical cancer progression. SLC38A1 inhibition is a potential therapeutic strategy for cervical cancer.

The N6-methyladenosine methylation landscape stratifies breast cancer into two subtypes with distinct immunological characteristics.

N6-methyladenosine (m6A) methylation modification affects the tumorigenesis and metastasis of breast cancer (BC). This study investigated the association between m6A regulator-mediated methylation modification patterns and characterization of the tumour microenvironment in BC, as well as their prognostic importance. Public gene expression data and clinical annotations were collected from The Cancer Genome Atlas (TCGA) database, the Gene Expression Omnibus website and the METABRIC program. We analysed the genetic expression, gene-gene interactions, gene mutations and copy number variations using R software. The data were screened for risk genes using the Cox risk regression model, and we developed an algorithm for risk score and its predictive value. Compared to adjacent normal tissue, we identified 16 differentially expressed m6A regulators in BC, including six writers and 10 readers. Under unsupervised clustering, two distinguished modification patterns were identified, cluster C1 and C2. Compared to m6A cluster C2, cluster C1 was found to be more involved in immune-related pathways, with a relatively higher immune score and stromal score (P < 0.05). Patients were divided into two groups based on their risk scores for survival analysis. The patients in the high-risk score group had significantly worse overall survival than patients in the low-risk score group, (P < 0.0001). The TCGA database validation revealed the same prognostic tendency. In summary, our study showed distinct m6A regulator modification patterns contribute to the immunological heterogeneity and diversity of BC. The development of m6A gene signatures and the m6A score aid in the prognostic prediction of patients with BC.

Identification of 6-methyladenosine sites using novel feature encoding methods and ensemble models.

N6-methyladenosine (6 mA) is the most common internal modification in eukaryotic mRNA. Mass spectrometry and site-directed mutagenesis, two of the most common conventional approaches, have been shown to be laborious and challenging. In recent years, there has been a rising interest in analyzing RNA sequences to systematically investigate mutated locations. Using novel methods for feature development, the current work aimed to identify 6 mA locations in RNA sequences. Following the generation of these novel features, they were used to train an ensemble of models using methods such as stacking, boosting, and bagging. The trained ensemble models were assessed using an independent test set and k-fold cross validation. When compared to baseline predictors, the suggested model performed better and showed improved ratings across the board for key measures of accuracy.

New comparative genomic evidence supporting the proteomic diversification role of A-to-I RNA editing in insects.

Adenosine-to-inosine (A-to-I) RNA editing, resembling A-to-G mutation, confers adaptiveness by increasing proteomic diversity in a temporal-spatial manner. This evolutionary theory named "proteomic diversifying hypothesis" has only partially been tested in very few organisms like Drosophila melanogaster, mainly by observing the positive selection on nonsynonymous editing events. To find additional genome-wide evidences supporting this interesting assumption, we retrieved the genomes of four Drosophila species and collected 20 deep-sequenced transcriptomes of different developmental stages and neuron populations of D. melanogaster. We systematically profiled the RNA editomes in these samples and performed meticulous comparative genomic analyses. Further evidences were found to support the diversifying hypothesis. (1) None of the nonsynonymous editing sites in D. melanogaster had ancestral G-alleles, while the silent editing sites had an unignorable fraction of ancestral G-alleles; (2) Only very few nonsynonymous editing sites in D. melanogaster had corresponding G-alleles derived in the genomes of sibling species, and the fraction of such situation was significantly lower than that of silent editing sites; (3) The few nonsynonymous editing with corresponding G-alleles had significantly more variable editing levels (across samples) than other nonsynonymous editing sites in D. melanogaster. The proteomic diversifying nature of RNA editing in Drosophila excludes the restorative role which favors an ancestral G-allele. The few fixed G-alleles in sibling species might facilitate the adaptation to particular environment and the corresponding nonsynonymous editing in D. melanogaster would introduce stronger advantage of flexible proteomic diversification. With multi-Omics data, our study consolidates the nature of evolutionary significance of A-to-I RNA editing sites in model insects.

N6-methyladenosine-modified circTEAD1 stabilizes Yap1 mRNA to promote chordoma tumorigenesis.

BACKGROUND: Chordoma, a rare bone tumour with aggressive local invasion and high recurrence rate with limited understanding of its molecular mechanisms. Circular RNAs (circRNAs) have been extensively implicated in tumorigenesis, yet their involvement in chordoma remains largely unexplored. N6-methyladenosine (m6A) modification holds a crucial function in regulating protein translation, RNA degradation and transcription. METHODS: Initially, screening and validation of circTEAD1 in chordoma were conducted by high-throughput sequencing. Subsequently, sh-circTEAD1 and an overexpression plasmid were constructed. Colony formation assays, cell counting kit-8, Transwell and wound healing assays were utilized to validate the function of circTEAD1 in vitro. RNA pull-down assays identified the binding proteins of circTEAD1, which underwent verification through RNA immunoprecipitation (RIP). Methylated RIP assays were conducted to detect the m6A binding sites. Following this, luciferase assay, RT-qPCR, RIP and Western blotting analyses were conducted, revealing that Yap1 was the direct target of circTEAD1. Afterwards, the same methods were utilized for the validation of the function of Yap1 in chordoma in vitro. Finally, the regulatory relationship between circTEAD1 and Yap1 in chordoma was verified by an in vivo tumour formation assay. RESULTS: CircTEAD1 was identified as an upregulated circRNA in chordoma specimens, with heightened circTEAD1 expression emerging as a prognostic indicator. In vitro experiments convincingly demonstrated that circTEAD1 significantly promoted chordoma cell invasion, migration and aggressiveness. Furthermore, the analysis revealed that methyltransferase-like 3-mediated m6A modification facilitated the cytoplasmic export of circTEAD1. The circTEAD1/IGF2BP3/Yap1 mRNA RNA-protein ternary complex not only bolstered the stability of Yap1 mRNA but also exerted a pivotal role in driving chordoma tumorigenesis. CONCLUSIONS: In this study, the role of m6A-modified circTEAD1 in chordoma was identified. The findings offer novel insights into the potential molecular targets for chordoma therapy, shedding light on the intricate interplay between circRNAs, m6A modification and Yap1 mRNA in chordoma pathogenesis.

A Mettl16/m6A/mybl2b/Igf2bp1 axis ensures cell cycle progression of embryonic hematopoietic stem and progenitor cells.

Prenatal lethality associated with mouse knockout of Mettl16, a recently identified RNA N6-methyladenosine (m6A) methyltransferase, has hampered characterization of the essential role of METTL16-mediated RNA m6A modification in early embryonic development. Here, using cross-species single-cell RNA sequencing analysis, we found that during early embryonic development, METTL16 is more highly expressed in vertebrate hematopoietic stem and progenitor cells (HSPCs) than other methyltransferases. In Mettl16-deficient zebrafish, proliferation capacity of embryonic HSPCs is compromised due to G1/S cell cycle arrest, an effect whose rescue requires Mettl16 with intact methyltransferase activity. We further identify the cell-cycle transcription factor mybl2b as a directly regulated by Mettl16-mediated m6A modification. Mettl16 deficiency resulted in the destabilization of mybl2b mRNA, likely due to lost binding by the m6A reader Igf2bp1 in vivo. Moreover, we found that the METTL16-m6A-MYBL2-IGF2BP1 axis controlling G1/S progression is conserved in humans. Collectively, our findings elucidate the critical function of METTL16-mediated m6A modification in HSPC cell cycle progression during early embryonic development.

Examining chromatin heterogeneity through PacBio long-read sequencing of M.EcoGII methylated genomes: an m6A detection efficiency and calling bias correcting pipeline.

Recent studies have combined DNA methyltransferase footprinting of genomic DNA in nuclei with long-read sequencing, resulting in detailed chromatin maps for multi-kilobase stretches of genomic DNA from one cell. Theoretically, nucleosome footprints and nucleosome-depleted regions can be identified using M.EcoGII, which methylates adenines in any sequence context, providing a high-resolution map of accessible regions in each DNA molecule. Here, we report PacBio long-read sequence data for budding yeast nuclei treated with M.EcoGII and a bioinformatic pipeline which corrects for three key challenges undermining this promising method. First, detection of m6A in individual DNA molecules by the PacBio software is inefficient, resulting in false footprints predicted by random gaps of seemingly unmethylated adenines. Second, there is a strong bias against m6A base calling as AT content increases. Third, occasional methylation occurs within nucleosomes, breaking up their footprints. After correcting for these issues, our pipeline calculates a correlation coefficient-based score indicating the extent of chromatin heterogeneity within the cell population for every gene. Although the population average is consistent with that derived using other techniques, we observe a wide range of heterogeneity in nucleosome positions at the single-molecule level, probably reflecting cellular chromatin dynamics.

Single-molecule epitranscriptomic analysis of full-length HIV-1 RNAs reveals functional roles of site-specific m6As.

Although the significance of chemical modifications on RNA is acknowledged, the evolutionary benefits and specific roles in human immunodeficiency virus (HIV-1) replication remain elusive. Most studies have provided only population-averaged values of modifications for fragmented RNAs at low resolution and have relied on indirect analyses of phenotypic effects by perturbing host effectors. Here we analysed chemical modifications on HIV-1 RNAs at the full-length, single RNA level and nucleotide resolution using direct RNA sequencing methods. Our data reveal an unexpectedly simple HIV-1 modification landscape, highlighting three predominant N6-methyladenosine (m6A) modifications near the 3' end. More densely installed in spliced viral messenger RNAs than in genomic RNAs, these m6As play a crucial role in maintaining normal levels of HIV-1 RNA splicing and translation. HIV-1 generates diverse RNA subspecies with distinct m6A ensembles, and maintaining multiple of these m6As on its RNAs provides additional stability and resilience to HIV-1 replication, suggesting an unexplored viral RNA-level evolutionary strategy.

m6Aexpress-enet: Predicting the regulatory expression m6A sites by an enet-regularization negative binomial regression model.

As the most abundant mRNA modification, m6A controls and influences many aspects of mRNA metabolism including the mRNA stability and degradation. However, the role of specific m6A sites in regulating gene expression still remains unclear. In additional, the multicollinearity problem caused by the correlation of methylation level of multiple m6A sites in each gene could influence the prediction performance. To address the above challenges, we propose an elastic-net regularized negative binomial regression model (called m6Aexpress-enet) to predict which m6A site could potentially regulate its gene expression. Comprehensive evaluations on simulated datasets demonstrate that m6Aexpress-enet could achieve the top prediction performance. Applying m6Aexpress-enet on real MeRIP-seq data from human lymphoblastoid cell lines, we have uncovered the complex regulatory pattern of predicted m6A sites and their unique enrichment pathway of the constructed co-methylation modules. m6Aexpress-enet proves itself as a powerful tool to enable biologists to discover the mechanism of m6A regulatory gene expression. Furthermore, the source code and the step-by-step implementation of m6Aexpress-enet is freely accessed at https://github.com/tengzhangs/m6Aexpress-enet.

Unraveling Key m6A Modification Regulators Signatures in Postmenopausal Osteoporosis through Bioinformatics and Experimental Verification.

OBJECTIVE: Bone marrow mesenchymal stem cells (BMSCs) show significant potential for osteogenic differentiation. However, the underlying mechanisms of osteogenic capability in osteoporosis-derived BMSCs (OP-BMSCs) remain unclear. This study aims to explore the impact of YTHDF3 (YTH N6-methyladenosine RNA binding protein 3) on the osteogenic traits of OP-BMSCs and identify potential therapeutic targets to boost their bone formation ability. METHODS: We examined microarray datasets (GSE35956 and GSE35958) from the Gene Expression Omnibus (GEO) to identify potential m6A regulators in osteoporosis (OP). Employing differential, protein interaction, and machine learning analyses, we pinpointed critical hub genes linked to OP. We further probed the relationship between these genes and OP using single-cell analysis, immune infiltration assessment, and Mendelian randomization. Our in vivo and in vitro experiments validated the expression and functionality of the key hub gene. RESULTS: Differential analysis revealed seven key hub genes related to OP, with YTHDF3 as a central player, supported by protein interaction analysis and machine learning methodologies. Subsequent single-cell, immune infiltration, and Mendelian randomization studies consistently validated YTHDF3's significant link to osteoporosis. YTHDF3 levels are significantly reduced in femoral head tissue from postmenopausal osteoporosis (PMOP) patients and femoral bone tissue from PMOP mice. Additionally, silencing YTHDF3 in OP-BMSCs substantially impedes their proliferation and differentiation. CONCLUSION: YTHDF3 may be implicated in the pathogenesis of OP by regulating the proliferation and osteogenic differentiation of OP-BMSCs.

DDX21 mediates co-transcriptional RNA m6A modification to promote transcription termination and genome stability.

N6-methyladenosine (m6A) is a crucial RNA modification that regulates diverse biological processes in human cells, but its co-transcriptional deposition and functions remain poorly understood. Here, we identified the RNA helicase DDX21 with a previously unrecognized role in directing m6A modification on nascent RNA for co-transcriptional regulation. DDX21 interacts with METTL3 for co-recruitment to chromatin through its recognition of R-loops, which can be formed co-transcriptionally as nascent transcripts hybridize onto the template DNA strand. Moreover, DDX21's helicase activity is needed for METTL3-mediated m6A deposition onto nascent RNA following recruitment. At transcription termination regions, this nexus of actions promotes XRN2-mediated termination of RNAPII transcription. Disruption of any of these steps, including the loss of DDX21, METTL3, or their enzymatic activities, leads to defective termination that can induce DNA damage. Therefore, we propose that the R-loop-DDX21-METTL3 nexus forges the missing link for co-transcriptional modification of m6A, coordinating transcription termination and genome stability.

Circ_0053943 complexed with IGF2BP3 drives uveal melanoma progression via regulating N6-methyladenosine modification of Epidermal growth factor receptor.

Numerous studies have characterized the critical role of circular RNAs (circRNAs) as regulatory factors in the progression of multiple cancers. However, the biological functions of circRNAs and their underlying molecular mechanisms in the progression of uveal melanoma (UM) remain enigmatic. In this study, we identified a novel circRNA, circ_0053943, through re-analysis of UM microarray data and quantitative RT-PCR. Circ_0053943 was found to be upregulated in UM and to promote the proliferation and metastatic ability of UM cells in both in vitro and in vivo settings. Mechanistically, circ_0053943 was observed to bind to the KH1 and KH2 domains of insulin-like growth factor 2 mRNA-binding protein 3 (IGF2BP3), thereby enhancing the function of IGF2BP3 by stabilizing its target mRNA. RNA sequencing assays identified epidermal growth factor receptor (EGFR) as a target gene of circ_0053943 and IGF2BP3 at the transcriptional level. Rescue assays demonstrated that circ_0053943 exerts its biological function by stabilizing EGFR mRNA and regulating the downstream mitogen-activated protein kinase/extracellular signal-regulated kinase (MAPK/ERK) signaling pathway. Collectively, circ_0053943 may promote UM progression by stabilizing EGFR mRNA and activating the MAPK/ERK signaling pathway through the formation of a circ_0053943/IGF2BP3/EGFR RNA-protein ternary complex, thus providing a potential biomarker and therapeutic target for UM.

Unraveling the cross-talk between N6-methyladenosine modification and non-coding RNAs in breast cancer: Mechanisms and clinical implications.

In recent years, breast cancer (BC) has surpassed lung cancer as the most common malignant tumor worldwide and remains the leading cause of cancer death in women. The etiology of BC usually involves dysregulation of epigenetic mechanisms and aberrant expression of certain non-coding RNAs (ncRNAs). N6-methyladenosine (m6A), the most prevalent RNA modification in eukaryotes, widely exists in ncRNAs to affect its biosynthesis and function, and is an important regulator of tumor-related signaling pathways. Interestingly, ncRNAs can also regulate or target m6A modification, playing a key role in cancer progression. However, the m6A-ncRNAs regulatory network in BC has not been fully elucidated, especially the regulation of m6A modification by ncRNAs. Therefore, in this review, we comprehensively summarize the interaction mechanisms and biological significance of m6A modifications and ncRNAs in BC. Meanwhile, we also focused on the clinical application value of m6A modification in BC diagnosis and prognosis, intending to explore new biomarkers and potential therapeutic targets.

Structural and functional effects of inosine modification in mRNA.

Inosine (I), resulting from the deamination of adenosine (A), is a prominent modification in the human transcriptome. The enzymes responsible for the conversion of adenosine to inosine in human mRNAs are the ADARs (adenosine deaminases acting on RNA). Inosine modification introduces a layer of complexity to mRNA processing and function, as it can impact various aspects of RNA biology, including mRNA stability, splicing, translation, and protein binding. The relevance of this process is emphasized in the growing number of human disorders associated with dysregulated A-to-I editing pathways. Here, we describe the impact of the A-to-I conversion on the structure and stability of duplex RNA and on the consequences of this modification at different locations in mRNAs. Furthermore, we highlight specific open questions regarding the interplay between inosine formation in duplex RNA and the innate immune response.

Quantification of tRNA m1A modification by templated-ligation qPCR.

N1-methyladenosine (m1A) is a widespread modification in all eukaryotic, many archaeal, and some bacterial tRNAs. m1A is generally located in the T loop of cytosolic tRNA and between the acceptor and D stems of mitochondrial tRNAs; it is involved in the tertiary interaction that stabilizes tRNA. Human tRNA m1A levels are dynamically regulated that fine-tune translation and can also serve as biomarkers for infectious disease. Although many methods have been used to measure m1A, a PCR method to assess m1A levels quantitatively in specific tRNAs has been lacking. Here we develop a templated-ligation followed by a qPCR method (TL-qPCR) that measures m1A levels in target tRNAs. Our method uses the SplintR ligase that efficiently ligates two tRNA complementary DNA oligonucleotides using tRNA as the template, followed by qPCR using the ligation product as the template. m1A interferes with the ligation in specific ways, allowing for the quantitative assessment of m1A levels using subnanogram amounts of total RNA. We identify the features of specificity and quantitation for m1A-modified model RNAs and apply these to total RNA samples from human cells. Our method enables easy access to study the dynamics and function of this pervasive tRNA modification.

Regulation of N6-methyladenosine modification in erythropoiesis and thalassemia.

In eukaryotic RNA, N6-methyladenosine (m6A) is a prevalent form of methylation modification. The m6A modification process is reversible and dynamic, written by m6A methyltransferase complex, erased by m6A demethylase, and recognized by m6A binding proteins. Through mediating RNA stability, decay, alternative splicing, and translation processes, m6A 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 m6A modification plays a crucial role in erythropoiesis. In this review, we comprehensively summarize the function of m6A modification in erythropoiesis and further generalize the mechanism of m6A 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.

Phosphorylated SHMT2 Regulates Oncogenesis Through m6A Modification in Lung Adenocarcinoma.

Targeting cancer-specific metabolic processes is a promising therapeutic strategy. Here, this work uses a compound library that directly inhibits metabolic enzymes to screen the potential metabolic targets in lung adenocarcinoma (LUAD). SHIN1, the specific inhibitor of serine hydroxymethyltransferase 1/2 (SHMT1/2), has a highly specific inhibitory effect on LUAD cells, and this effect depends mainly on the overexpression of SHMT2. This work clarifies that mitogen-activated protein kinase 1 (MAPK1)-mediated phosphorylation at Ser90 is the key mechanism underlying SHMT2 upregulation in LUAD and that this phosphorylation stabilizes SHMT2 by reducing STIP1 homology and U-box containing protein 1 (STUB1)-mediated ubiquitination and degradation. SHMT2-Ser90 dephosphorylation decreases S-adenosylmethionine levels in LUAD cells, resulting in reduced N6-methyladenosine (m6A) levels in global RNAs without affecting total protein or DNA methylation. Methylated RNA immunoprecipitation sequencing (MeRIP-Seq) and RNA sequencing (RNA-Seq) analyses further demonstrate that SHMT2-Ser90 dephosphorylation accelerates the RNA degradation of oncogenic genes by reducing m6A modification, leading to the inhibition of tumorigenesis. Overall, this study elucidates a new regulatory mechanism of SHMT2 during oncogenesis and provides a theoretical basis for targeting SHMT2 as a therapeutic target in LUAD.