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Understanding YTHDF2-mediated mRNA degradation by m6A-BERT-Deg.

ABSTRACT: N6-methyladenosine (m6A) is the most abundant mRNA modification within mammalian cells, holding pivotal significance in the regulation of mRNA stability, translation and splicing. Furthermore, it plays a critical role in the regulation of RNA degradation by primarily recruiting the YTHDF2 reader protein. However, the selective regulation of mRNA decay of the m6A-methylated mRNA through YTHDF2 binding is poorly understood. To improve our understanding, we developed m6A-BERT-Deg, a BERT model adapted for predicting YTHDF2-mediated degradation of m6A-methylated mRNAs. We meticulously assembled a high-quality training dataset by integrating multiple data sources for the HeLa cell line. To overcome the limitation of small training samples, we employed a pre-training-fine-tuning strategy by first performing a self-supervised pre-training of the model on 427 760 unlabeled m6A site sequences. The test results demonstrated the importance of this pre-training strategy in enabling m6A-BERT-Deg to outperform other benchmark models. We further conducted a comprehensive model interpretation and revealed a surprising finding that the presence of co-factors in proximity to m6A sites may disrupt YTHDF2-mediated mRNA degradation, subsequently enhancing mRNA stability. We also extended our analyses to the HEK293 cell line, shedding light on the context-dependent YTHDF2-mediated mRNA degradation.

SUBMITTER: Zhang TH

PROVIDER: S-EPMC11018547 | biostudies-literature | 2024 Mar

REPOSITORIES: biostudies-literature

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Understanding YTHDF2-mediated mRNA degradation by m6A-BERT-Deg.

Zhang Ting-He TH Jo Sumin S Zhang Michelle M Wang Kai K Gao Shou-Jiang SJ Huang Yufei Y

Briefings in bioinformatics 20240301 3

N6-methyladenosine (m6A) is the most abundant mRNA modification within mammalian cells, holding pivotal significance in the regulation of mRNA stability, translation and splicing. Furthermore, it plays a critical role in the regulation of RNA degradation by primarily recruiting the YTHDF2 reader protein. However, the selective regulation of mRNA decay of the m6A-methylated mRNA through YTHDF2 binding is poorly understood. To improve our understanding, we developed m6A-BERT-Deg, a BERT model adap ...[more]

PMID: 38622358

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Project description:BackgroundEndometriosis-related infertility is a common worldwide reproductive health concern. Despite ongoing research, the causes of infertility remain unclear. Evidence suggests that epigenetic regulation is crucial in reproduction. However, the role of N6-methyladenosine (m6A) modification of RNA in endometriosis-related infertility requires further investigation.MethodsWe examined the expression of m6A and methyltransferase-like 3 (METTL3) in endometrial samples taken from normal fertile women in the proliferative phase (the NP group) or the mid-secretory phase (the NS group) or from women with endometriosis-related infertility at the mid-secretory phase (the ES group). We treated primary endometrial stromal cells (ESCs) with medroxyprogesterone acetate and 8-Bromo-cyclic adenosine monophosphate for in vitro decidualization and detected the expression of m6A, METTL3, and decidual markers. We analyzed the expression of m6A, METTL3, and forkhead box O1 (FOXO1) in ESCs from normal fertile women (the ND group) or women with endometriosis-related infertility (the ED group). We also assessed the expression of m6A, METTL3, and decidual markers, as well as the embryo adhesion rate, upon METTL3 overexpression or knockdown. Additionally, we investigated the role of METTL3 in embryo implantation in vivo by applying mice with endometriosis. Furthermore, we performed RNA stability assays, RNA immunoprecipitation (RIP), and methylated RIP assays to explore the mechanisms underlying the regulation of FOXO1 by METTL3-mediated m6A.ResultsThe expression of m6A and METTL3 was reduced only in the NS group; the NP and ES groups demonstrated increased m6A and METTL3 levels. m6A and METTL3 levels decreased in ESCs with prolonged decidual treatment. Compared to the ND group, m6A and METTL3 levels in the ED group increased after decidual treatment, whereas the expression of FOXO1 decreased. METTL3 overexpression suppressed the expression of decidual markers and embryo implantation in vitro; METTL3 knockdown exhibited the opposite effect. Inhibition of METTL3 promoted embryo implantation in vivo. Furthermore, we observed that METTL3-mediated m6A regulated the degradation of FOXO1 mRNA through YTHDF2, a m6A binding protein.ConclusionsMETTL3-regulated m6A promotes YTHDF2-mediated decay of FOXO1 mRNA, thereby affecting cellular decidualization and embryo implantation. These findings provide novel insights into the development of therapies for women with endometriosis-related infertility.

Project description:BackgroundN6-methyladenosine (m6A) methylation is the most abundant chemical posttranscriptional modification of mRNA, and it is associated with the regulation of the immune response to tumors. However, the function of m6A modification in the immune response to endometrial cancer (EC) remains unknown. Our study investigated the immunological role of methyltransferase-like 3 (METTL3) in EC and the underlying molecular mechanism.MethodsWe investigated the correlation between the expression of METTL3 and CD8 by using an endometrial tissue microarray cohort. Next, we investigated the role and mechanism of METTL3 in the immune response to EC using a mouse tumor model and a CD8+ T cell-EC cell coculture system after METTL3 overexpression or depletion. Additionally, RNA immunoprecipitation (RIP), methylated RIP, and RNA stability experiments were used to investigate the mechanism underlying the function of METTL3 in immunosurveillance of EC.ResultsMETTL3 levels were downregulated in EC patients, low levels of METTL3 were correlated with poor prognosis in EC patients. There was a positive correlation between METTL3 expression and CD8 expression. Overexpression of METTL3 in the EC cell and CD8+ T cell coculture system inhibited EC cell proliferation, migration, and promoted CD8+ T-cell proliferation, and in vivo, METTL3 overexpression increased CD8+ T cell proportions and inhibited EC progression; however, genetic depletion of METTL3 exerted the opposite effects. NLR family CARD domain-containing 5 (NLRC5) was identified as a target of METTL3-mediated m6A modification. The degradation of NLRC5 was increased by YTH domain-containing family 2 (YTHDF2).ConclusionsOverall, METTL3, YTHDF2, and NLRC5 have potential to be the diagnostic and prognostic biomarkers for EC. METTL3 facilitated the m6A modifications of NLRC5 and inhibited its degradation through a YTHDF2-dependent mechanism in EC. Genetic overexpression of METTL3 attenuated the immune evasion of EC by promoting NLRC5-mediated immunosurveillance, suggesting that the METTL3/YTHDF2/NLRC5 axis is a promising target of immunotherapy in EC.

Project description:BackgroundHepatic fibrosis may progress to liver cirrhosis and eventually cause death. Epithelial-mesenchymal transition (EMT) of hepatocytes plays critical roles in hepatic fibrosis. Exploring the mechanisms underlying EMT is crucial for a better understanding of hepatic fibrosis pathogenesis.MethodsHepatocyte EMT wad induced with TGF-β1 and evaluated by Western blotting and immunofluorescence staining. Methylated RNA immunoprecipitation (MeRIP) was applied to assess N6-methyladenosine (m6A) modification. RIP and RNA pull-down assays were performed to analyze the interaction between circABHD3, YTHDF2 and YPEL3 mRNA. MEOX1-mediated transcription of ABHD3 was examined by luciferase and chromatin immunoprecipitation (ChIP). Mice were intraperitoneally injected with CCl4 or treated with bile duct ligation (BDL) surgery for hepatic fibrosis induction. Liver injury and collagen deposition were examined with hematoxylin and eosin (HE), Masson, and Sirius Red staining. Alanine transaminase (ALT), aspartate transaminase (AST) and hydroxyproline (HYP) were examined using ELISA.ResultsCircABHD3 was upregulated in in vitro and in vivo models of hepatic fibrosis and patients. Knockdown of circABHD3 inhibited TGF-β1-induced expression of fibrosis markers, EMT and mitochondrial impairment in hepatocytes. MEOX1 could directly bind to the promoter of ABHD3 to facilitate its transcription and subsequent circABHD3 generation. Knockdown of MEOX1 suppressed TGF-β1-induced EMT and mitochondrial impairment through suppression of circABHD3. CircABHD3 destabilized YPEL3 mRNA via promoting YTHDF2-dependent recognition of m6A-modified YPEL3 mRNA to trigger β-catenin signaling activation. Furthermore, circABHD3 silencing-mediated inhibition of EMT and mitochondrial impairment was counteracted by YPEL3 knockdown and activation of β-catenin signaling. Depletion of circABHD3 significantly reduced EMT, mitochondrial impairment and hepatic fibrosis via promoting YPEL3 expression and suppressing β-catenin signaling in vivo.ConclusionMEOX1-mediated generation of circABHD3 promotes EMT and mitochondrial impairment by enhancing YTHDF2-mediated degradation of YPEL3 mRNA and activating downstream β-catenin signaling, thus exacerbating hepatic fibrosis.

Project description:BackgroundCurdione is a sesquiterpene isolated from Curcumae Rhizoma that possesses high biological activity and extensive pharmacological effects. As a traditional Chinese medicine, Curcumae Rhizoma can inhibit the development of many types of cancer, especially colorectal cancer. However, the anti-colorectal mechanism of its monomer curdione remains unclear.MethodsColorectal cancer (CRC) cells were treated with curdione at doses of 12.5 μM, 25 μM, and 50 μM, and then the cells' activity was measured with methyl thiazolyl tetrazolium (MTT). Nude mice were administered different doses of curdione subcutaneously and oxaliplatin by tail vein injection, and then hematoxylin-eosin (HE) staining was adopted to examine tumor histology. Moreover, flow cytometry was applied to detect reactive oxygen species in cells and tissues. Kits were employed to detect the levels of iron ions, malondialdehyde, lipid hydroperoxide, and glutathione. Polymerase chain reaction (PCR) and Western blotting were adopted to detect ferroptosis and m6A modification-related factors. A methylation spot hybridization assay was performed to measure changes in overall methylation. SLC7A11 and HOXA13 were measured by MeRIP-qPCR. The shRNA-METTL14 plasmid was constructed to verify the inhibitory effect of curdione on CRC.ResultsA dose-dependent decrease in activity was observed in curdione-treated cells. Curdione increased the accumulation of reactive oxygen species in CRC cells and tumor tissues, greatly enhanced the levels of malondialdehyde, lipid hydroperoxide and Fe2+, and lowered the activity of glutathione. According to the qPCR and Western blot results, curdione promoted the expression of METTL14 and YTHDF2 in CRC cells and tissues, respectively, and decreased the expression of SLC7A11, SLC3A2, HOXA13, and glutathione peroxidase 4. Additionally, in animal experiments, the curdione-treated group showed severe necrosis of tumor cells, as displayed by HE staining. Furthermore, compared with the control group, levels of m6A modifying factors (namely, SLC7A11 and HOXA13) were increased in the tissues after drug intervention. METTL14 knockdown was followed by an increase in CRC cell activity and glutathione levels. However, the levels of reactive oxygen species, malondialdehyde, and iron ions decreased. The expression levels of SLC7A11, SLC3A2, HOXA13, and GPX4 were all increased after METTL14 knockdown.ConclusionThe results suggest that curdione induces ferroptosis in CRC by virtue of m6A methylation.

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Understanding YTHDF2-mediated mRNA degradation by m6A-BERT-Deg.

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Understanding YTHDF2-mediated mRNA degradation by m6A-BERT-Deg.

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