bims-rimeca Biomed News
on RNA methylation in cancer
Issue of 2021‒06‒27
nineteen papers selected by
Sk Ramiz Islam
Saha Institute of Nuclear Physics
  1. Exploration of the Role of m6 A RNA Methylation Regulators in Malignant Progression and Clinical Prognosis of Ovarian Cancer.
  2. N6-Methyladenosine RNA Modification in Inflammation: Roles, Mechanisms, and Applications.
  3. Prognostic Implication of the m6A RNA Methylation Regulators in Rectal Cancer.
  4. Pan-Cancer Prognostic, Immunity, Stemness, and Anticancer Drug Sensitivity Characterization of N6-Methyladenosine RNA Modification Regulators in Human Cancers.
  5. Targeting the m6A RNA modification pathway blocks SARS-CoV-2 and HCoV-OC43 replication.
  6. Data mining analysis of the prognostic impact of N6-methyladenosine regulators in patients with endometrial adenocarcinoma.
  7. RNA modifications in hematopoietic malignancies: A new research frontier.
  8. ALKBH5-mediated m6A-demethylation of USP1 regulated T-cell acute lymphoblastic leukemia cell glucocorticoid resistance by Aurora B.
  9. Deep and accurate detection of m6A RNA modifications using miCLIP2 and m6Aboost machine learning.
  10. Integrative genomic analysis of N6-methyladenosine-single nucleotide polymorphisms (m6A-SNPs) associated with breast cancer.
  11. METTL3 promotes tumour development by decreasing APC expression mediated by APC mRNA N6-methyladenosine-dependent YTHDF binding.
  12. m6A demethylase FTO induces NELL2 expression by inhibiting E2F1 m6A modification leading to metastasis of non-small cell lung cancer.
  13. m6A-Induced LncRNA MEG3 Suppresses the Proliferation, Migration and Invasion of Hepatocellular Carcinoma Cell Through miR-544b/BTG2 Signaling.
  14. METTL3/N6-methyladenosine/ miR-21-5p promotes obstructive renal fibrosis by regulating inflammation through SPRY1/ERK/NF-κB pathway activation.
  15. Melatonin Attenuates Chromium (VI)-Induced Spermatogonial Stem Cell/Progenitor Mitophagy by Restoration of METTL3-Mediated RNA N6-Methyladenosine Modification.
  16. METTL3 induces bone marrow mesenchymal stem cells osteogenic differentiation and migration through facilitating M1 macrophage differentiation.
  17. M5C regulator-mediated methylation modification patterns and tumor microenvironment infiltration characterization in lung adenocarcinoma.
  18. The RNA m6A reader YTHDF2 controls NK cell antitumor and antiviral immunity.
  19. N6-Methyladenosine Modification of LncRNA DUXAP9 Promotes Renal Cancer Cells Proliferation and Motility by Activating the PI3K/AKT Signaling Pathway.
  1. Front Genet. 2021 ;12 650554
    Exploration of the Role of m6 A RNA Methylation Regulators in Malignant Progression and Clinical Prognosis of Ovarian Cancer.
    Qinglv Wei, Dan Yang, Xiaoyi Liu, Hongyan Zhao, Yu Yang, Jing Xu, Tao Liu, Ping Yi.
      Ovarian cancer is the most deadly gynecologic malignancy worldwide and it is warranted to dissect the critical gene regulatory network in ovarian cancer. N6-methyladenosine (m6A) RNA methylation, as the most prevalent RNA modification, is orchestrated by the m6A RNA methylation regulators and has been implicated in malignant progression of various cancers. In this study, we investigated the genetic landscape and expression profile of the m6A RNA methylation regulators in ovarian cancer and found that several m6A RNA methylation regulators were frequently amplified and up-regulated in ovarian cancer. Utilizing consensus cluster analysis, we stratified ovarian cancer samples into four clusters with distinct m6A methylation patterns and patients in these subgroups displayed the different clinical outcomes. Moreover, multivariate Cox proportional hazard model was used to screen the key m6A regulators associated with the prognosis of ovarian cancer and the last absolute shrinkage and selection operator (LASSO) Cox regression was used to construct the gene signature for prognosis prediction. The survival analysis exhibited the risk-gene signature could be used as independent prognostic markers for ovarian cancer. In conclusion, m6A RNA methylation regulators are associated with the malignant progression of ovarian cancer and could be a potential in prognostic prediction for ovarian cancer.
    Keywords:  LASSO Cox regression; TCGA; TME; consensus clustering; m6A; ovarian cancer; prognosis
    DOI:  https://doi.org/10.3389/fgene.2021.650554
  2. Front Cell Dev Biol. 2021 ;9 670711
    N6-Methyladenosine RNA Modification in Inflammation: Roles, Mechanisms, and Applications.
    Jiahui Luo, Tao Xu, Kai Sun.
      N6-methyladenosine (m6A) is the most prevalent internal mRNA modification. m6A can be installed by the methyltransferase complex and removed by demethylases, which are involved in regulating post-transcriptional expression of target genes. RNA methylation is linked to various inflammatory states, including autoimmunity, infection, metabolic disease, cancer, neurodegenerative diseases, heart diseases, and bone diseases. However, systematic knowledge of the relationship between m6A modification and inflammation in human diseases remains unclear. In this review, we will discuss the association between m6A modification and inflammatory response in diseases, especially the role, mechanisms, and potential clinical application of m6A as a biomarker and therapeutic target for inflammatory diseases.
    Keywords:  N6-methyladenosine; RNA modification; epigenetics; inflammation; inflammatory disease
    DOI:  https://doi.org/10.3389/fcell.2021.670711
  3. Front Genet. 2021 ;12 604229
    Prognostic Implication of the m6A RNA Methylation Regulators in Rectal Cancer.
    Yajie Chen, Shanshan Wang, William C Cho, Xiang Zhou, Zhen Zhang.
      N6-methyladenosine (m6A) is a very common and abundant RNA modifications occurring in nearly all types of RNAs. Although the dysregulated expression of m6A regulators is implicated in cancer progression, our understanding of the prognostic value of the m6A regulators in rectal cancer is still quite limited. In this study, we analyzed the RNA expression levels of the 17 m6A regulator genes of 95 rectal cancer and 10 normal rectal samples from the The Cancer Genome Atlas Rectum Adenocarcinoma (TCGA-READ) dataset. Lasso regression analysis was conducted to build a prognostic model and calculate the risk score. The rectal cancer patients were then devided into the high-risk and low-risk groups according to the mean risk score. The prognostic value of the identified model was separately evaluated in the TCGA-READ and GSE87211 datasets. GSEA was conducted to analyze the functional difference of high-risk and low-risk rectal cancer patients. Our analysis revealed that rectal cancer patients with lower expression of YTHDC2 and METTL14 had a remarkable worse overall survival (P < 0.05). The prognostic value of the model was validated in GSE87211 datasets, with AUC = 0.612 for OS and AUC = 0.651 for RFS. Furthermore, the m6A modification-based risk score system is associated with activation of distinct signaling pathways, such as DNA repair, epithelial-mesenchymal transition, G2M checkpoint and the MYC pathway, that may contribute to the progression of rectal cancer. In conclusion, our findings demonstrated that the m6A RNA methylation regulators, specifically YTHDC2 and METTL14, were significantly down-regulated and might be potential prognostic biomarkers in rectal cancer.
    Keywords:  METTL14; N6-methyladenosine; YTHDC2; prognosis; rectal cancer
    DOI:  https://doi.org/10.3389/fgene.2021.604229
  4. Front Mol Biosci. 2021 ;8 644620
    Pan-Cancer Prognostic, Immunity, Stemness, and Anticancer Drug Sensitivity Characterization of N6-Methyladenosine RNA Modification Regulators in Human Cancers.
    Rui Li, Yun-Hong Yin, Xiu-Li Ji, Xiao Liu, Jian-Ping Li, Yi-Qing Qu.
      N6-methyladenosine RNA modification plays a significant role in the progression of multiple tumorigenesis. Our study identified the imperative role of m6A regulators in the tumor immune microenvironment, survival, stemness score, and anticancer drug sensitivity of pan-cancer. The Wilcox test was to identify the differential expression between 17 m6A regulators across 33 TCGA cancer types and their normal tissues from UCSC Xena GDC pan-cancer. Survival analysis of m6A-related regulators in 33 TCGA cancer types was identified using the "survival" and "survminer" package. The Spearman correlation test and Pearson correlation test were used to identify the correlation relationship between m6A regulators expression and tumor microenvironment, tumor stem cell score, and drug sensitivity of anticancer drugs. ConsensusPathDB was used for exploring m6A regulators functional enrichment. The 17 (METTL3, WTAP, METTL14, RBM15, RBM15B, VIRMA, HNRNPC, HNRNPA2B1, YTHDC1, ZC3H13, YTHDF1, YTHDC2, YTHDF2, IGF2BP3, IGF2BP1, FTO, and ALKBH5) m6A regulators were differentially expressed in 18 TCGA cancer types and adjacent normal tissues. Correlation analysis indicated that the relationship between the expression of 17 m6A regulators and tumor microenvironment indicated that the higher expression of m6A regulators, the higher the degree of tumor stem cells. The anticancer drug sensitivity analysis indicated that ZC3H13 expression had a positive relationship with anticancer drugs such as selumetinib, dabrafenib, cobimetinib, trametinib, and hypothemycin (p < 0.001). YTHDF2 expression was significantly negatively correlated with the anticancer drug dasatinib (p < 0.001). The pan-cancer immune subtype analysis showed that the 17 m6A regulators were significantly different in immune subtype C1 (wound healing), C3 (inflammatory), C2 (IFN-gamma dominant), C5 (immunological quiet), C4 (lymphocyte depleted), and C6 (TGF-beta dominant) (p < 0.001). Our study provides a comprehensive insight for revealing the significant role of m6A regulators in the tumor immune microenvironment, stemness score, and anticancer drug sensitivity of human cancers.
    Keywords:  drug sensitivity; m6A regulators; overall survival; pan-cancer analysis; tumor immune microenvironment
    DOI:  https://doi.org/10.3389/fmolb.2021.644620
  5. Genes Dev. 2021 Jun 24.
    Targeting the m6A RNA modification pathway blocks SARS-CoV-2 and HCoV-OC43 replication.
    Hannah M Burgess, Daniel P Depledge, Letitia Thompson, Kalanghad Puthankalam Srinivas, Rebecca C Grande, Elizabeth I Vink, Jonathan S Abebe, Wesley P Blackaby, Alan Hendrick, Mark R Albertella, Tony Kouzarides, Kenneth A Stapleford, Angus C Wilson, Ian Mohr.
      N6-methyladenosine (m6A) is an abundant internal RNA modification, influencing transcript fate and function in uninfected and virus-infected cells. Installation of m6A by the nuclear RNA methyltransferase METTL3 occurs cotranscriptionally; however, the genomes of some cytoplasmic RNA viruses are also m6A-modified. How the cellular m6A modification machinery impacts coronavirus replication, which occurs exclusively in the cytoplasm, is unknown. Here we show that replication of SARS-CoV-2, the agent responsible for the COVID-19 pandemic, and a seasonal human β-coronavirus HCoV-OC43, can be suppressed by depletion of METTL3 or cytoplasmic m6A reader proteins YTHDF1 and YTHDF3 and by a highly specific small molecule METTL3 inhibitor. Reduction of infectious titer correlates with decreased synthesis of viral RNAs and the essential nucleocapsid (N) protein. Sites of m6A modification on genomic and subgenomic RNAs of both viruses were mapped by methylated RNA immunoprecipitation sequencing (meRIP-seq). Levels of host factors involved in m6A installation, removal, and recognition were unchanged by HCoV-OC43 infection; however, nuclear localization of METTL3 and cytoplasmic m6A readers YTHDF1 and YTHDF2 increased. This establishes that coronavirus RNAs are m6A-modified and host m6A pathway components control β-coronavirus replication. Moreover, it illustrates the therapeutic potential of targeting the m6A pathway to restrict coronavirus reproduction.
    Keywords:  HCoV-OC43; N6-methyladenosine; RNA modification; SARS-CoV-2; coronavirus; direct RNA sequencing; nanopore sequencing; virus–host interactions
    DOI:  https://doi.org/10.1101/gad.348320.121
  6. J Cancer. 2021 ;12(15): 4729-4738
    Data mining analysis of the prognostic impact of N6-methyladenosine regulators in patients with endometrial adenocarcinoma.
    Junyu Zhai, Shang Li, Yu Li, Yanzhi Du.
      We reanalyzed the expression of 16 acknowledged N6-methyladenosine (m6A) RNA regulators in 406 endometrial adenocarcinoma patients and 19 controls using The Cancer Genome Atlas (TCGA) dataset, and further verified our results using Gene Expression Omnibus (GEO) dataset and real-time quantitative polymerase chain reaction. Thirteen m6A RNA methylation regulators were differentially expressed between patients with endometrial adenocarcinoma and controls. FTO, RBM15, and YTHDF1, were identified as independent prognostic markers and closely associated with International Federation of Gynecology and Obstetrics grade in endometrial cancer patients. GEO dataset also verified the differential expression of FTO and RBM15 between patients with endometrial adenocarcinoma and hyperplasia. Functional enrichment and ingenuity pathway analysis network suggested that FTO and RBM15 contributed to the survival of patients with endometrial adenocarcinoma via the regulation of connective tissue development, catabolic process, RNA stability, oxidative demethylation, temperature homeostasis, and energy metabolism through IGF1, IRS1, RBM24, LARP1, and CBFA2T3. The decreased FTO expression and increased RBM15 expression in endometrial adenocarcinoma from our validation cohort was consistent with in silico analysis using TCGA and GEO datasets. In conclusion, m6A methylation regulators, especially FTO, RBM15, and YTHDF1, are critical in the progression and prognosis of endometrial adenocarcinoma.
    Keywords:  Endometrial Adenocarcinoma; Energy Metabolism; N6-methyladenosine; RNA-Binding Protein
    DOI:  https://doi.org/10.7150/jca.50868
  7. Blood. 2021 Jun 22. pii: blood.2019004263. [Epub ahead of print]
    RNA modifications in hematopoietic malignancies: A new research frontier.
    Ying Qing, Rui Su, Jianjun Chen.
      Both protein-coding and noncoding RNAs can be decorated with a wealth of chemical modifications and such modifications coordinately orchestrate gene expression during normal hematopoietic differentiation and development. However, aberrant expression and/or dysfunction of the relevant RNA modification modulators/regulators ("writers", "erasers", and "readers") drive the initiation and progression of hematopoietic malignancies, and targeting these dysregulated modulators holds potent therapeutic potential for the treatment of hematopoietic malignancies. In this review, we summarize current progress in the understanding of the biological functions and underlying mechanisms of RNA modifications in normal and malignant hematopoiesis, with a focus on the N6-methyladenosine (m6A) modification, and discuss the therapeutic potential of targeting RNA modifications for the treatment of hematopoietic malignancies, especially acute myeloid leukemia (AML).
    DOI:  https://doi.org/10.1182/blood.2019004263
  8. Mol Carcinog. 2021 Jun 25.
    ALKBH5-mediated m6A-demethylation of USP1 regulated T-cell acute lymphoblastic leukemia cell glucocorticoid resistance by Aurora B.
    Hongtao Gong, Liu Liu, Lina Cui, Hongyan Ma, Liyun Shen.
      Recent studies evidence that ubiquitin-specific proteases (USPs) are associated with the occurrence and chemoresistance of T-cell acute lymphoblastic leukemia (T-ALL). N6 -methyladenosine (m6A) demethylase AlkB homolog 5 (ALKBH5) exerts a carcinogenic effect in human cancers and improves the mRNA stability of USPs. Whether ubiquitin-specific protease 1 (USP1) controls chemoresistance of T-ALL is unknown. Our study demonstrated that USP1 expression was upregulated in glucocorticoid (GC)-resistant T-ALL patients and cells (CEM-C1). High expression of USP1 was correlated to the poor prognosis in T-ALL patients. Silencing USP1 increased CEM-C1 cell sensitivity to dexamethasone (Dex), reduced cell invasion, promoted cell apoptosis, and ameliorated glucocorticoid receptor (GR) expression. USP1 mediated T-ALL chemoresistance by interacting with and deubiquitination of Aurora B. Overexpression of USP1 reversed the amelioration effect of Aurora B inhibitor on CEM-C1 cell resistance to Dex. Mechanistically, ALKBH5 enhanced USP1 expression by reducing m6A level and mRNA stability in USP1 mRNA transcript. Downregulation of ALKBH5 reduced the levels of USP1 and Aurora B, facilitated CEM-C1 cell sensitivity to Dex, apoptosis, and GR expression, suppressed cell invasion. However, overexpression of USP1 reversed all the effects of ALKBH5 on CEM-C1 cells. In vivo results showed that tail vein injection of sh-USP1 resulted in a significant prolongation of mouse survival, suppressed tumor growth, maintained the normal weight of mice, reduced USP1 expression and facilitated GR expression. In conclusion, inhibition of ALKBH5-mediated m6A modification decreased USP1 expression and downregulation of USP1 ameliorated GC resistance of T-ALL through suppressing Aurora B expression and elevating GR level.
    Keywords:  Aurora B; N6-methyladenosine demethylase; T-cell acute lymphoblastic leukemia; chemoresistance; ubiquitin-specific protease 1
    DOI:  https://doi.org/10.1002/mc.23330
  9. Nucleic Acids Res. 2021 Jun 22. pii: gkab485. [Epub ahead of print]
    Deep and accurate detection of m6A RNA modifications using miCLIP2 and m6Aboost machine learning.
    Nadine Körtel, Cornelia Rücklé, You Zhou, Anke Busch, Peter Hoch-Kraft, F X Reymond Sutandy, Jacob Haase, Mihika Pradhan, Michael Musheev, Dirk Ostareck, Antje Ostareck-Lederer, Christoph Dieterich, Stefan Hüttelmaier, Christof Niehrs, Oliver Rausch, Dan Dominissini, Julian König, Kathi Zarnack.
      N6-methyladenosine (m6A) is the most abundant internal RNA modification in eukaryotic mRNAs and influences many aspects of RNA processing. miCLIP (m6A individual-nucleotide resolution UV crosslinking and immunoprecipitation) is an antibody-based approach to map m6A sites with single-nucleotide resolution. However, due to broad antibody reactivity, reliable identification of m6A sites from miCLIP data remains challenging. Here, we present miCLIP2 in combination with machine learning to significantly improve m6A detection. The optimized miCLIP2 results in high-complexity libraries from less input material. Importantly, we established a robust computational pipeline to tackle the inherent issue of false positives in antibody-based m6A detection. The analyses were calibrated with Mettl3 knockout cells to learn the characteristics of m6A deposition, including m6A sites outside of DRACH motifs. To make our results universally applicable, we trained a machine learning model, m6Aboost, based on the experimental and RNA sequence features. Importantly, m6Aboost allows prediction of genuine m6A sites in miCLIP2 data without filtering for DRACH motifs or the need for Mettl3 depletion. Using m6Aboost, we identify thousands of high-confidence m6A sites in different murine and human cell lines, which provide a rich resource for future analysis. Collectively, our combined experimental and computational methodology greatly improves m6A identification.
    DOI:  https://doi.org/10.1093/nar/gkab485
  10. Bioengineered. 2021 Dec;12(1): 2389-2397
    Integrative genomic analysis of N6-methyladenosine-single nucleotide polymorphisms (m6A-SNPs) associated with breast cancer.
    Zixue Xuan, Yiwen Zhang, Jinying Jiang, Xiaowei Zheng, Xiaoping Hu, Xiuli Yang, Yanfei Shao, Guobing Zhang, Ping Huang.
      Due to the important role of N6-methyladenosine (m6A) in breast cancer, single nucleotide polymorphisms (SNPs) in genes with m6A modification may also be involved in breast cancer pathogenesis. In this study, we used a public genome-wide association study dataset to identify m6A-SNPs associated with breast cancer and to further explore their potential functions. We found 113 m6A-SNPs associated with breast cancer that reached the genome-wide suggestive threshold (5.0E-05), and 86 m6A-SNPs had eQTL signals. Only six genes were differentially expressed between controls and breast cancer cases in GEO datasets (GSE15852, GSE115144, and GSE109169), and the SNPs rs4829 and rs9610915 were located next to the m6A modification sites in the 3'UTRs of TOM1L1 and MAFF, respectively. In addition, we found that polyadenylate-binding protein cytoplasmic 1 might have a potential interaction with rs4829 (TOM1L1) and rs9610915 (MAFF). In summary, these findings indicated that the SNPs rs4829 and rs9610915 are potentially associated with breast cancer because they had eQTL signals, altered gene expression, and were located next to the m6A modification sites in the 3'UTRs of their coding genes. However, further studies are still needed to clarify how genetic variation affects the epigenetic modification, m6A, and its subsequent functions in the pathogenesis of breast cancer.
    Keywords:  GWAS; breast cancer; m6A; m6Avar; single nucleotide polymorphisms (SNPs)
    DOI:  https://doi.org/10.1080/21655979.2021.1935406
  11. Nat Commun. 2021 06 21. 12(1): 3803
    METTL3 promotes tumour development by decreasing APC expression mediated by APC mRNA N6-methyladenosine-dependent YTHDF binding.
    Wei Wang, Fei Shao, Xueying Yang, Juhong Wang, Rongxuan Zhu, Yannan Yang, Gaoxiang Zhao, Dong Guo, Yingli Sun, Jie Wang, Qi Xue, Shugeng Gao, Yibo Gao, Jie He, Zhimin Lu.
      The adenomatous polyposis coli (APC) is a frequently mutated tumour suppressor gene in cancers. However, whether APC is regulated at the epitranscriptomic level remains elusive. In this study, we analysed TCGA data and separated 200 paired oesophageal squamous cell carcinoma (ESCC) specimens and their adjacent normal tissues and demonstrated that methyltransferase-like 3 (METTL3) is highly expressed in tumour tissues. m6A-RNA immunoprecipitation sequencing revealed that METTL3 upregulates the m6A modification of APC, which recruits YTHDF for APC mRNA degradation. Reduced APC expression increases the expression of β-catenin and β-catenin-mediated cyclin D1, c-Myc, and PKM2 expression, thereby leading to enhanced aerobic glycolysis, ESCC cell proliferation, and tumour formation in mice. In addition, downregulated APC expression correlates with upregulated METTL3 expression in human ESCC specimens and poor prognosis in ESCC patients. Our findings reveal a mechanism by which the Wnt/β-catenin pathway is upregulated in ESCC via METTL3/YTHDF-coupled epitranscriptomal downregulation of APC.
    DOI:  https://doi.org/10.1038/s41467-021-23501-5
  12. Mol Ther Oncolytics. 2021 Jun 25. 21 367-376
    m6A demethylase FTO induces NELL2 expression by inhibiting E2F1 m6A modification leading to metastasis of non-small cell lung cancer.
    Yanyun Wang, Man Li, Lin Zhang, Yitong Chen, Shoudan Zhang.
      Non-small cell lung cancer (NSCLC) represents one of the primary causes of cancer-related mortality all over the world. Following our initial finding of the upregulated expression of E2F transcription factor-1 (E2F1) in the NSCLC-related microarray, this study aimed to explore the regulatory role of E2F1 and underlying mechanism in NSCLC development. NSCLC cell viability, migration, and invasion were evaluated utilizing Cell Counting Kit 8 (CCK-8), 5-ethynyl-2'-deoxyuridine (EdU), wound-healing, and Transwell assays. Loss- and gain-function assays were performed to determine the effects of the fat mass and obesity-associated protein (FTO)/E2F1/neural epidermal growth factor-like 2 (NELL2) axis on NSCLC cell behaviors in vitro and NSCLC tumor growth in vivo. E2F1 was highly expressed in both NSCLC tissues and cells. E2F1 augmented the viability, migration, and invasion of NSCLC cells, which was attributable to E2F1 transcriptionally activating NELL2. FTO upregulated the expression of E2F1 by inhibiting the m6A modification of E2F1. The FTO/E2F1/NELL2 axis modulated NSCLC cell viability, migration, and invasion in vitro as well as affected NSCLC tumor growth and metastasis in vivo. The FTO/E2F1/NELL2 axis may impart pro-tumorigenic effects on the cell behavior of NSCLC cells and thus accelerate NSCLC progression.
    Keywords:  E2F transcription factor-1; demethylase; fat mass and obesity-associated protein; m6A modification; neural epidermal growth factor-like 2; non-small cell lung cancer; tumor growth; tumor metastasis
    DOI:  https://doi.org/10.1016/j.omto.2021.04.011
  13. Onco Targets Ther. 2021 ;14 3745-3755
    m6A-Induced LncRNA MEG3 Suppresses the Proliferation, Migration and Invasion of Hepatocellular Carcinoma Cell Through miR-544b/BTG2 Signaling.
    Jian Wu, Runhua Pang, Minan Li, Bin Chen, Junhai Huang, Yaqing Zhu.
      Objective: Hepatocellular carcinoma (HCC) is one of the most common malignant tumors. Long non-coding RNA plays an important role in the development of HCC. This study analyzed the impact of MEG3 on malignant behavior of HCC and explored its possible molecular mechanism.Methods: Expression of MEG3 in HCC tissues and cell lines was measured by qRT-PCR. Transfection efficiency of MEG3 was verified by qRT-PCR. Cell proliferation, transwell migration, invasion and cell cloning assays were used to detect the effect of MEG3 on the proliferation, migration and invasion ability of HCC cells. The bioinformatics analysis was applied to predict the binding between miR-544b and MEG3 as well as BTG2. Luciferase reporter assay was performed to verify their interaction. Finally, the m6A modification of MEG3 by METTL3 was identified through RIP experiments.
    Results: MEG3 was lowly expressed in HCC tissues and cells. Overexpression of MEG3 inhibits the proliferation, migration and invasion of HCC cells. MiR-544b can be sponged by MEG3, and overexpression of miR-544b reverses the anti-cancer effect of MEG3. We further confirmed that BTG2 gene is the target gene of miR-544b. Epigenetic studies have shown that METTL3-mediated N6-methyladenosine modification led to MEG3 downregulation.
    Conclusion: In HCC, MEG3 and BTG2 are lowly expressed while miR-544b is highly expressed. MEG3 regulates the expression of BTG2 through miR-544b, thus affecting the malignant behavior of HCC. METTL3 regulates the m6A modification of MEG3 and its expression. This study clarified the role of MEG3/miR-544b/BTG2 axis in HCC and also provided new targets for HCC research.
    Keywords:  BTG2; HCC; N6-methyladenosine; lncRNA MEG3; miR-544b
    DOI:  https://doi.org/10.2147/OTT.S289198
  14. J Cell Mol Med. 2021 Jun 24.
    METTL3/N6-methyladenosine/ miR-21-5p promotes obstructive renal fibrosis by regulating inflammation through SPRY1/ERK/NF-κB pathway activation.
    Erpeng Liu, Lei Lv, Yonghao Zhan, Yuan Ma, Jinjin Feng, Yulin He, Yibo Wen, Yanping Zhang, Qingsong Pu, Fengping Ji, Xinghuan Yang, Jian Guo Wen.
      Renal fibrosis induced by urinary tract obstruction is a common clinical occurrence; however, effective treatment is lacking, and a deeper understanding of the mechanism of renal fibrosis is needed. Previous studies have revealed that miR-21 impacts liver and lung fibrosis progression by activating the SPRY1/ERK/NF-kB signalling pathway. However, whether miR-21 mediates obstructive renal fibrosis through the same signalling pathway has not been determined. Additionally, studies have shown that N6-methyladenosine (m6 A) modification-dependent primary microRNA (pri-microRNA) processing is essential for maturation of microRNAs, but its role in the maturation of miR-21 in obstructive renal fibrosis has not yet been investigated in detail. To address these issues, we employed a mouse model of unilateral ureteral obstruction (UUO) in which the left ureters were ligated for 3, 7 and 14 days to simulate the fibrotic process. In vitro, human renal proximal tubular epithelial (HK-2) cells were transfected with plasmids containing the corresponding sequence of METTL3, miR-21-5p mimic or miR-21-5p inhibitor. We found that the levels of miR-21-5p and m6 A modification in the UUO model groups increased significantly, and as predicted, the SPRY1/ERK/NF-kB pathway was activated by miR-21-5p, confirming that miR-21-5p plays an important role in obstructive renal fibrosis by enhancing inflammation. METTL3 was found to play a major catalytic role in m6 A modification in UUO mice and drove obstructive renal fibrosis development by promoting miR-21-5p maturation. Our research is the first to demonstrate the role of the METTL3-m6 A-miR-21-5p-SPRY1/ERK/NF-kB axis in obstructive renal fibrosis and provides a deeper understanding of renal fibrosis.
    Keywords:  METTL3; N6-methyladenosine (m6A); Spry1/ERK/NF-κB; miR-21-5p; renal fibrosis; urinary tract obstruction
    DOI:  https://doi.org/10.1111/jcmm.16603
  15. Front Cell Dev Biol. 2021 ;9 684398
    Melatonin Attenuates Chromium (VI)-Induced Spermatogonial Stem Cell/Progenitor Mitophagy by Restoration of METTL3-Mediated RNA N6-Methyladenosine Modification.
    Yinghua Lv, Tianjiao Li, Manman Yang, Lihong Su, Zhendong Zhu, Sihang Zhao, Wenxian Zeng, Yi Zheng.
      Spermatogonial stem cells (SSCs) are the basis of spermatogenesis, and any damage to SSCs may result in spermatogenic disorder and male infertility. Chromium (Cr) (VI) is a proven toxin, mutagen, and carcinogen, perpetually detrimental to environmental organisms due to its intricate and enduring detoxification process in vivo. Despite this, the deleterious effects of Cr (VI) on SSCs and the underlying mechanisms remain poorly understood. In this study, we identified that Cr (VI) impaired male reproductive system in mouse testes and induced mitochondrial dynamic imbalance and mitophagy in SSCs/progenitors. Cr (VI) also downregulated the RNA N6-methyladenosine (m6A) modification levels in mitochondrial dynamic balance and mitophagy genes in SSCs/progenitors. Inspiringly, the toxic effects of Cr (VI) could be relieved by melatonin pretreatment. Melatonin alleviated Cr (VI)-induced damage to male reproductive system and autophagy in mouse testes. Melatonin also attenuated Cr (VI)-induced cell viability loss and reactive oxygen species (ROS) generation, as well as mitochondrial dynamic disorders and mitophagy in SSCs/progenitors. The protective roles of melatonin against Cr (VI)-induced mitophagy were exerted by restoration of METTL3-mediated RNA m6A modification and activation of mitochondrial fusion proteins MFN2 and OPA1, as well as inhibition of the mitophagy BNIP3/NIX receptor pathway. Thus, our study provides novel insights into the molecular mechanisms for RNA m6A modification underlying the gene regulatory network responsible for mitochondrial dynamic balance, and also lays new experimental groundwork for treatment of Cr (VI)-induced damage to male fertility.
    Keywords:  RNA m6A modification; chromium (VI); melatonin; mitophagy; spermatogonial stem cells
    DOI:  https://doi.org/10.3389/fcell.2021.684398
  16. Am J Transl Res. 2021 ;13(5): 4376-4388
    METTL3 induces bone marrow mesenchymal stem cells osteogenic differentiation and migration through facilitating M1 macrophage differentiation.
    Hong Lei, Mingyu He, Xiaoqi He, Guanghui Li, Yang Wang, Yuelin Gao, Gege Yan, Quan Wang, Tao Li, Guoxin Liu, Weijie Du, Ye Yuan, Lei Yang.
      Despite the crucial role of m6A methyltransferase METTL3 in multiple diseases onset and progression, there are still lacking hard evidence proving that METTL3 could affect macrophage polarization in the stage of bone repair. Here, we aimed to explore the potential involvement of METTL3 in bone repair through modulating macrophage polarization and decipher the underlying cellular/molecular mechanisms. Here we treated RAW 264.7 cells and BM-derived primary macrophages (BMDM) with lipopolysaccharide (LPS) to induce M1 differentiation. METTL3 expression was upregulated in pro-inflammatory macrophages (M1) as compared with macrophages (M0). And overexpression of METTL3 promoted the expression of IL-6 and iNOS secretion by M1 macrophage. In the coculture condition, M1 macrophages with forced expression of METTL3 significantly enhanced migration ability of BMSCs, and also remarkably facilitated osteogenesis ability of BMSCs; the opposite was true when expression of METTL3 was knockdown. In addition, the m6A-RIP microarray suggested that METTL3 silencing significantly reduce the m6A modification of DUSP14, HDAC5 and Nfam1. Furthermore, the findings showed that expression of HADC5 was downregulated in M1 macrophages with METTL3 knockdown, while the DUSP14 expression had slight change and Nfam1 expression was very low. In contrast, METTL3 overexpression promoted HDAC5 expression, indicating that HDAC5 is the critical target gene of METTL3. Under such a theme, we proposed that METTL3 overexpression might be a new approach of replacement therapy for the treatment of bone repair.
    Keywords:  BMSCs; METTL3; macrophage; migration; osteogenic differentiation
  17. Transl Lung Cancer Res. 2021 May;10(5): 2172-2192
    M5C regulator-mediated methylation modification patterns and tumor microenvironment infiltration characterization in lung adenocarcinoma.
    Hui Chen, Xiao-Lin Ge, Zhao-Yue Zhang, Ming Liu, Rui-Yan Wu, Xiao-Fei Zhang, Li-Ping Xu, Hong-Yan Cheng, Xin-Chen Sun, Hong-Cheng Zhu.
      Background: In recent years, immunotherapy has made great progress, and the regulatory role of epigenetics has been verified. However, the role of 5-methylcytosine (m5C) in the tumor microenvironment (TME) and immunotherapy response remains unclear.Methods: Based on 11 m5C regulators, we evaluated the m5C modification patterns of 572 lung adenocarcinoma (LUAD) patients. The m5C score was constructed by principal component analysis (PCA) algorithms in order to quantify the m5C modification pattern of individual LUAD patients.
    Results: Two m5C methylation modification patterns were identified according to 11 m5C regulators. The two patterns had a remarkably distinct TME immune cell infiltration characterization. Next, 226 differentially expressed genes (DEGs) related to the m5C phenotype were screened. Patients were divided into three different gene cluster subtypes based on these genes, which had different TME immune cell infiltration and prognosis characteristics. The m5C score was constructed to quantify the m5C modification pattern of individual LUAD patients. We found that the high m5C score group had a better prognosis. The role of the m5C score in predicting prognosis was also verified in the dataset GSE31210.
    Conclusions: Our study revealed that m5C modification played a significant role in TME regulation of LUAD. Investigation of the m5C regulation mode may have some implications for tumor immunotherapy in the future.
    Keywords:  M5C; immunotherapy; lung adenocarcinoma (LUAD); tumor microenvironment (TME)
    DOI:  https://doi.org/10.21037/tlcr-21-351
  18. J Exp Med. 2021 Aug 02. pii: e20210279. [Epub ahead of print]218(8):
    The RNA m6A reader YTHDF2 controls NK cell antitumor and antiviral immunity.
    Shoubao Ma, Jiazhuo Yan, Tasha Barr, Jianying Zhang, Zhenhua Chen, Li-Shu Wang, Joseph C Sun, Jianjun Chen, Michael A Caligiuri, Jianhua Yu.
      N 6-methyladenosine (m6A) is the most prevalent posttranscriptional modification on RNA. NK cells are the predominant innate lymphoid cells that mediate antiviral and antitumor immunity. However, whether and how m6A modifications affect NK cell immunity remain unknown. Here, we discover that YTHDF2, a well-known m6A reader, is upregulated in NK cells upon activation by cytokines, tumors, and cytomegalovirus infection. Ythdf2 deficiency in NK cells impairs NK cell antitumor and antiviral activity in vivo. YTHDF2 maintains NK cell homeostasis and terminal maturation, correlating with modulating NK cell trafficking and regulating Eomes, respectively. YTHDF2 promotes NK cell effector function and is required for IL-15-mediated NK cell survival and proliferation by forming a STAT5-YTHDF2 positive feedback loop. Transcriptome-wide screening identifies Tardbp to be involved in cell proliferation or survival as a YTHDF2-binding target in NK cells. Collectively, we elucidate the biological roles of m6A modifications in NK cells and highlight a new direction to harness NK cell antitumor immunity.
    DOI:  https://doi.org/10.1084/jem.20210279
  19. Front Oncol. 2021 ;11 641833
    N6-Methyladenosine Modification of LncRNA DUXAP9 Promotes Renal Cancer Cells Proliferation and Motility by Activating the PI3K/AKT Signaling Pathway.
    Lei Tan, Yiming Tang, Hongbo Li, Pengju Li, Yunlin Ye, Junjie Cen, Chengpeng Gui, Junhang Luo, Jiazheng Cao, Jinhuan Wei.
      Most localized human renal clear cell carcinoma (ccRCC)-related deaths result from cancer recurrence and metastasis. However, the precise molecular mechanisms largely remain unknown. In recent years, an increasing number of long noncoding RNAs (lncRNAs) have been shown to be vital regulators of tumorigenesis. In this study, we characterized a lncRNA DUXAP9 and the upregulation of DUXAP9 was analyzed by quantitative real-time PCR in 112 pairs of localized ccRCC tumor tissues compared with adjacent normal tissues. Kaplan-Meier curves showed that patients of localized ccRCC with high DUXAP9 expression had poorer overall survival (P<0.01) and progression-free survival (P<0.05) than cases with low DUXAP9 expression. Multivariate Cox regression analysis also showed that high DUXAP9 expression was an independent risk factor for poor prognosis in localized ccRCC (p<0.05). DUXAP9 knockdown in renal cancer cells inhibited renal cancer cells proliferation and motility capacities in vitro and reversed epithelial-mesenchymal transition (EMT), whereas overexpression of DUXAP9 promoted renal cancer cells proliferation and motility capacities in vitro and induced EMT. Pull-down, RNA immunoprecipitation and RNA stability assays (involving actinomycin D) showed that DUXAP9 was methylated at N6-adenosine and binds to IGF2BP2, which increases its stability. DUXAP9 activate PI3K/AKT pathway and Snail expression in renal cancer cells. DUXAP9 may be useful as a prognostic marker and/or therapeutic target in localized ccRCC.
    Keywords:  Akt; EMT; N6-methyladenosine; ccRCC; lncRNA
    DOI:  https://doi.org/10.3389/fonc.2021.641833
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