bims-rimeca 2022-04-24 papers

bims-rimeca

Biomed News

on RNA methylation in cancer

Issue of 2022–04–24
nine papers selected by
Sk Ramiz Islam, Saha Institute of Nuclear Physics

METTL3-mediated m6A modification of STEAP2 mRNA inhibits papillary thyroid cancer progress by blocking the Hedgehog signaling pathway and epithelial-to-mesenchymal transition.
Methyltransferase 3, N6-adenosine-methyltransferase complex catalytic subunit-induced long intergenic non-protein coding RNA 1833 N6-methyladenosine methylation promotes the non-small cell lung cancer progression via regulating heterogeneous nuclear ribonucleoprotein A2/B1 expression.
METTL3 Promotes the Progression of Lung Cancer via Activating PI3K/AKT/mTOR Pathway.
The "m6A writer" METTL3 and the "m6A reader" IGF2BP2 regulate cutaneous T-cell lymphomas (CTCL) progression via CDKN2A.
An Alternatively Spliced Variant of METTL3 Mediates Tumor Suppression in Hepatocellular Carcinoma.
Primary sequence-assisted prediction of m6A RNA methylation sites from Oxford nanopore direct RNA sequencing data.
Risk Score Prediction Model of Prognosis in GC Patients by Age and Gender Combined With m6A Modification Genes FTO and RBM15.
Lipopolysaccharide facilitates immune escape of hepatocellular carcinoma cells via m6A modification of lncRNA MIR155HG to upregulate PD-L1 expression.
YTHDF1 Negatively Regulates Treponema pallidum-Induced Inflammation in THP-1 Macrophages by Promoting SOCS3 Translation in an m6A-Dependent Manner.

Cell Death Dis. 2022 Apr 18. 13(4): 358

METTL3-mediated m6A modification of STEAP2 mRNA inhibits papillary thyroid cancer progress by blocking the Hedgehog signaling pathway and epithelial-to-mesenchymal transition.

Yue Zhu, Xinzhi Peng, Qianlei Zhou, Langping Tan, Cheng Zhang, Shaojian Lin, Miaoyun Long.

Papillary thyroid cancer (PTC) is a common endocrine system malignancy all over the world. Aberrant expression of six transmembrane epithelial antigen of the prostate 2 (STEAP2) has been functionally associated with cancer progression in many cancers. Nevertheless, its biological function in PTC is still unclear. Here, we found that PTC tissues had preferentially downregulated STEAP2 as compared with noncancerous tissues. Low STEAP2 expression correlated with aggressive clinicopathological characteristics and dismal prognosis in patients with PTC. We performed gain- and loss-of-function experiments, including cell proliferation assay (Cell Counting Kit-8 assay), EdU (5-ethynyl-2'-deoxyuridine) and colony formation assays, transwell migration, and invasion assays, and constructed a nude mouse xenograft tumor model. The results demonstrated that STEAP2 overexpression inhibited PTC cell proliferation, migration, and invasion in vitro and inhibited lung metastasis and tumorigenicity in vivo. Conversely, silencing STEAP2 yielded the opposite results in vitro. Mechanistically, bioinformatics analysis combined with validation experiments identified STEAP2 as the downstream target of methyltransferase-like 3 (METTL3)-mediated N6-methyladenosine (m6A) modification. METTL3 stabilized STEAP2 mRNA and regulated STEAP2 expression positively in an m6A-dependent manner. We also showed that m6A-mediated STEAP2 mRNA translation initiation relied on a pathway dependent on the m6A reader protein YTHDF1. Rescue experiments revealed that silencing STEAP2 partially rescued the tumor-suppressive phenotype induced by METTL3 overexpression. Lastly, we verified that the METTL3-STEAP2 axis functions as an inhibitor in PTC by suppressing epithelial-mesenchymal transition and the Hedgehog signaling pathway. Taken together, these findings strongly suggest that METTL3-mediated STEAP2 m6A modification plays a critical tumor-suppressive role in PTC progression. The METTL3-STEAP2 axis may be a potential therapeutic molecular target against PTC.

DOI: https://doi.org/10.1038/s41419-022-04817-6
Bioengineered. 2022 Apr;13(4): 10493-10503

Methyltransferase 3, N6-adenosine-methyltransferase complex catalytic subunit-induced long intergenic non-protein coding RNA 1833 N6-methyladenosine methylation promotes the non-small cell lung cancer progression via regulating heterogeneous nuclear ribonucleoprotein A2/B1 expression.

Dongliang Li, Zejun Fu, Chaoqun Dong, Yongming Song.

  Long intergenic non-protein coding RNA 1833 (LINC01833) exhibits elevated expression in the non-small cell lung cancer (NSCLC) tissues, while its molecular mechanism in NSCLC progression remains elusive. Herein, the proliferation, migration, invasion as well as apoptosis of NSCLC cells were assessed. The potential N6-methyladenosine (m6A) modification site was predicted by the m6aVar tool. RNA pulldown and m6A-specific immunoprecipitation assays were used to detect the interaction between LINC01833 and methyltransferase 3, N6-adenosine-methyltransferase complex catalytic subunit (METTL3). RNA pull-down together with mass spectrometry were performed to assess the binding relationship between LINC01833 and heterogeneous nuclear ribonucleoprotein A2/B1 (HNRNPA2B1) in NSCLC. Tumor xenograft mice model was established, and the tumor size and weight were measured. The results demonstrated that LINC01833 expression was elevated in NSCLC samples. Overexpression of LINC01833 promoted proliferative, migratory, and invasive abilities and inhibited HCC827 cell apoptosis. LINC01833 knockdown inhibited tumor growth in mice. LINC01833 is further demonstrated to be modulated by METTL3, which is highly expressed in NSCLC samples. In addition, RNA pulldown and m6A-specific immunoprecipitation assays indicated that LINC01833 might form a complex with HNRNPA2B1. In conclusion, m6A transferase METTL3-induced LINC01833 m6A methylation promotes NSCLC progression through modulating HNRNPA2B1 expression. Our findings indicated that LINC01833 might be a therapeutic target for NSCLC.

Keywords:  HNRNPA2B1; LINC01833; METTL3; Non-small cell lung cancer; m6A methylation; molecular mechanism

DOI:  https://doi.org/10.1080/21655979.2022.2061305
Clin Exp Pharmacol Physiol. 2022 Apr 17.

METTL3 Promotes the Progression of Lung Cancer via Activating PI3K/AKT/mTOR Pathway.

Feng-Wu Cheng, Li-Ming Peng, Dan Luo.

  Methyltransferase-like 3 (METTL3) catalyzes N6-methyladenosine (m6 A) modification on mRNA and participates in a wide range of biological functions via epigenetically regulating gene expression. Recent studies suggested that dysregulation of METTL3 is associated with multiple human cancers; however, the role of METTL3 in lung cancer remains unclear. In present study, through transcriptome analysis of lung cancer patients, we found that METTL3 is overexpressed in lung cancer patients and is associated with poor patient survival. More importantly, combining both in vitro and in vivo models, we revealed that in lung cancer cells, METTL3 overexpression activates PI3K/AKT/mTOR pathway and mTOR-mediated protein synthesis. Mechanistically, METTL3 promotes PI3K expression by introducing m6 A modification in PI3K 3' untranslated region (3' UTR). Elevated PI3K level then activates downstream AKT and mTOR signaling pathway and results in rapid cancer cell proliferation and metastasis. Taken together, our study reveals that METTL3-mediated m6 A methylation promotes lung cancer progression via activating PI3K/AKT/mTOR pathway.

Keywords:  METTL3; PI3K; lung cancer; m6A; mTOR

DOI:  https://doi.org/10.1111/1440-1681.13647
Hematol Oncol. 2022 Apr 21.

The "m6A writer" METTL3 and the "m6A reader" IGF2BP2 regulate cutaneous T-cell lymphomas (CTCL) progression via CDKN2A.

Xinchen Wang, Maogui Hu, Lu Yu, Xiaoyan Wang, Xinlu Jiang, Guihong Zhang, Kaiyang Ding.

  It has been established that Cutaneous T-Cell lymphomas (CTCL) are caused by the monoclonal proliferation of T lymphocytes in the skin. This heterogeneous group of diseases represents a significant source of distress to patients since the diagnosis and treatment are often challenging. As one of the most abundant internal modifications in mRNA in higher eukaryotes, N6-methyladenosine (m6A) is widely recognized to affect the development and progression of cancers. However, knowledge on the involvement of m6A in CTCL is still limited. In this work, we revealed the role of METTL3-mediated m6A modification in CTCL progression. ELISA, western blot, and qRT-PCR assays demonstrated that METTL3 was significantly downregulated in CTCL cells both in vivo and in vitro. CCK-8, EdU, flow cytometry, and transwell assays showed that the decline in METTL3 levels was responsible for CTCL cell proliferation and migration. Furthermore, using small interfering RNAs (siRNAs) against METTL3 and the RIP assay, we showed that CDKN2A was a key regulator during this process in vitro and in vivo, and insufficient methylation modification blocked the interaction between CDKN2A and m6A reader IGF2BP2, resulting in mRNA degradation. To the best of our knowledge, this is the first study to depict the role of m6A in CTCL development and provide potential bio-targets for therapy. This article is protected by copyright. All rights reserved.

Keywords:  CDKN2A; IGF2BP2; METTL3; cutaneous T-Cell lymphomas; m6A modification

DOI:  https://doi.org/10.1002/hon.3005
Genes (Basel). 2022 Apr 11. pii: 669. [Epub ahead of print]13(4):

An Alternatively Spliced Variant of METTL3 Mediates Tumor Suppression in Hepatocellular Carcinoma.

Rui-Yao Xu, Zhan Ding, Qing Zhao, Tiao-Ying Ke, Shu Chen, Xing-Yu Wang, Yao-Yun Wang, Meng-Fei Sheng, Wei Wang, Ni Long, Yu-Xian Shen, Yong-Zhen Xu, Wei Shao.

  Many post-transcriptional mRNA processing steps play crucial roles in tumorigenesis and the progression of cancers, such as N6-methyladenosine (m6A) modification and alternative splicing. Upregulation of methyltransferase-like 3 (METTL3), the catalytic core of the m6A methyltransferase complex, increases m6A levels and results in significant effects on the progression of hepatocellular carcinoma (HCC). However, alternative splicing of METTL3 has not been fully investigated, and the functions of its splice variants remain unclear. Here, we analyzed both our and online transcriptomic data, obtaining 13 splice variants of METTL3 in addition to canonical full-length METTL3-A in HCC cell lines and tissues. Validated by RT-qPCR and Western blotting, we found that METTL3-D, one of the splice variants expressing a truncated METTL3 protein, exhibits higher levels than METTL3-A in normal human livers but lower levels than METTL3-A in HCC tumor tissues and cell lines. Further functional assays demonstrated that METTL3-D expression decreased cellular m6A modification, inhibited the proliferation, migration, and invasion of HCC cells, and was negatively associated with the malignancy of patient tumors, exhibiting functions opposite to those of full-length METTL3-A. This study demonstrates that the METTL3-D splice variant is a tumor suppressor that could potentially be used as a target for HCC therapy.

Keywords:  METTL3; RNA m6A; hepatocellular carcinoma; splice variants; tumor suppressor

DOI:  https://doi.org/10.3390/genes13040669
Methods. 2022 Apr 13. pii: S1046-2023(22)00089-5. [Epub ahead of print]

Primary sequence-assisted prediction of m6A RNA methylation sites from Oxford nanopore direct RNA sequencing data.

Yuxin Zhang, Daiyun Huang, Zhen Wei, Kunqi Chen.

  Traditional epitranscriptome profiling approach relies on specific antibodies or chemical treatments to capture modified RNA molecules and then applies high throughput sequencing to identify their transcriptomic locations. However, due to the lack of suitable or high-quality antibodies, only a small proportion of the 170 documented RNA modifications were profiled with those approaches. Direct sequencing of native RNA molecules using Oxford Nanopore Technologies (ONT) enabled straight inspection of RNA modifications and offered a promising alternative solution. N6-methyladenosine (m6A) is known to cause characteristic changes and increased base call errors of ONT signals compared with non-modified adenosines, based on which, the m6A sites can be identified directly from ONT signals. Meanwhile, a number of studies have shown that it is possible to predict m6A sites from RNA primary sequences. Using the m6A sites revealed by Illumina technology as benchmark, we showed that, the accuracy of ONT-based m6A site prediction can be further increased by integrating additional information from the primary sequences of RNA (AUROC of 0.918), compared with using ONT signals only (AUROC 0.878 using Base call error features, and 0.804 using Tombo features), providing a new perspective for more reliable mining of the relatively noisy ONT signals.

Keywords:  Machine learning; Oxford nanopore technique; RNA modification

DOI:  https://doi.org/10.1016/j.ymeth.2022.04.003
Front Cell Dev Biol. 2022 ;10 710708

Risk Score Prediction Model of Prognosis in GC Patients by Age and Gender Combined With m6A Modification Genes FTO and RBM15.

Limin Yue, Rongguang Zhang, Shuaiyin Chen, Guangcai Duan.

  Background: Gastric cancer (GC) has a high mortality rate. N6-methyladenosine (m6A) is involved in the development of GC. Age and gender are associated with GC incidence and survival. This study aimed to explore the risk score prediction model of prognosis in GC patients by age and gender combined with m6A modification genes. Methods: Data on m6A modification gene expression and clinical information downloaded from the Cancer Genome Atlas (TCGA) database were used to construct the risk score prediction model. Cox and least absolute shrinkage and selection operator (LASSO) regression were performed to identify clinical characteristics and m6A modification genes associated with prognosis. A risk score prediction model was established based on multivariate Cox regression analysis. The Gene Expression Omnibus (GEO) database was used to validate this model. Results: Most of the m6A modification genes were upregulated in GC tumor tissues compared with that in normal tissues and were correlated with clinical characteristics including grade, stage status, and T status. The risk score prediction model was established based on age, gender, FTO, and RBM15. GC patients were divided into high- or low-risk groups based on the median risk score. Patients with a high risk score had poor prognosis. Multivariate Cox regression indicated that risk score was an independent prognostic factor for GC patients. The data from GSE84437 verified the predictive value of this model. Conclusion: The risk score prediction model based on age and gender combined with m6A modification genes FTO and RBM15 was an independent prognostic factor for GC patients.

Keywords:  FTO; N6-methyladenosine; RBM15; gastric cancer; prognostic factor

DOI:  https://doi.org/10.3389/fcell.2022.710708
Cell Biol Toxicol. 2022 Apr 19.

Lipopolysaccharide facilitates immune escape of hepatocellular carcinoma cells via m6A modification of lncRNA MIR155HG to upregulate PD-L1 expression.

Lirong Peng, Banglun Pan, Xiaoxia Zhang, Zengbin Wang, Jiacheng Qiu, Xiaoqian Wang, Nanhong Tang.

  Recent studies have suggested that the initiation and progression of hepatocellular carcinoma (HCC) are closely associated with lipopolysaccharide (LPS) of intestinal bacteria. However, the role of LPS in immune regulation of HCC remains largely unknown. An orthotopic Hepa1-6 tumor model of HCC was constructed to analyze the effect of LPS on the expression of immune checkpoint molecules PD-1 and PD-L1. Then we verified the regulation of PD-L1 by LPS in HCC cells. Based on the previous finding that lncRNA MIR155HG regulates PD-L1 expression in HCC cells, we analyzed the relationship of LPS signaling pathway molecules with PD-L1 and MIR155HG by bioinformatics. The molecular mechanism of MIR155HG regulating PD-L1 expression induced by LPS was investigated by RNA pull-down followed by mass spectrometry, RNA immunoprecipitation, fluorescence in situ hybridization, and luciferase reporter assay. Finally, the HepG2 xenograft model was established to determine the role of MIR155HG on PD-L1 expression in vivo. We showed that LPS induced PD-1 and PD-L1 expression in mouse tumor tissues and induced PD-L1 expression in HCC cells. Mechanistically, upregulation of METTL14 by LPS promotes the m6A methylation of MIR155HG, which stabilizes MIR155HG relying on the "reader" protein ELAVL1 (also known as HuR)-dependent pathway. Moreover, MIR155HG functions as a competing endogenous RNA (ceRNA) to modulate the expression of PD-L1 by miR-223/STAT1 axis. Our results suggested that LPS plays a critical role in immune escape of HCC through METTL14/MIR155HG/PD-L1 axis. This study provides a new insight for understanding the complex immune microenvironment of HCC. 1. LPS plays a critical role in immune escape of HCC, especially HCC with cirrhosis. 2. Our study reveals that LPS regulates PD-L1 by m6A modification of lncRNA in HCC. 3. MIR155HG plays an important role in LPS induced PD-L1 expression. 4. LPS-MIR155HG-PD-L1 regulatory axis provides a new target for the treatment of HCC.

Keywords:  Hepatocellular carcinoma; Lipopolysaccharide; Long noncoding RNA; METTL14; PD-L1

DOI:  https://doi.org/10.1007/s10565-022-09718-0
Front Immunol. 2022 ;13 857727

YTHDF1 Negatively Regulates Treponema pallidum-Induced Inflammation in THP-1 Macrophages by Promoting SOCS3 Translation in an m6A-Dependent Manner.

Zhijia Li, Muzhou Teng, Yinbo Jiang, Litian Zhang, Xi Luo, Yuhui Liao, Bin Yang.

   Background: Previous studies have confirmed that the bacterium Treponema pallidum (TP) or its proteins provide signals to macrophages that induce an inflammatory response; however, little is known about the negative regulation of this macrophage-mediated inflammatory response during syphilis infection or the underlying mechanism. Recent evidence suggests the role of the RNA modification, N6-adenosine methylation (m6A), in regulating the inflammatory response and pathogen-host cell interactions. Therefore, we hypothesized that m6A plays a role in the regulation of the inflammatory response in macrophages exposed to TP.
Methods: We first assessed m6A levels in TP-infected macrophages differentiated from the human monocyte cell line THP-1. The binding and interaction between the m6A "writer" methyltransferase-like 3 (METTL3) or the m6A "reader" YT521-B homology (YTH) domain-containing protein YTHDF1 and the suppressor of cytokine signaling 3 (SOCS3), as a major regulator of the inflammatory response, were explored in differentiated TP-infected THP-1 cells as well as in secondary syphilitic lesions from patients. The mechanisms by which YTHDF1 and SOCS3 regulate the inflammatory response in macrophages were assessed.
Results and Conclusion: After macrophages were stimulated by TP, YTHDF1 was upregulated in the cells. YTHDF1 was also upregulated in the syphilitic lesions compared to adjacent tissue in patients. YTHDF1 recognizes and binds to the m6A methylation site of SOCS3 mRNA, consequently promoting its translation, thereby inhibiting the JAK2/STAT3 pathway, and reducing the secretion of inflammatory factors, which results in anti-inflammatory regulation. This study provides the first demonstration of the role of m6A methylation in the pathological process of syphilis and further offers new insight into the pathogenesis of TP infection.

Keywords:  Treponema pallidum; inflammation; m6A methylation; macrophage; syphilis

DOI:  https://doi.org/10.3389/fimmu.2022.857727