bims-rimeca 2021-12-05 papers

bims-rimeca

Biomed News

on RNA methylation in cancer

Issue of 2021–12–05
ten papers selected by
Sk Ramiz Islam, Saha Institute of Nuclear Physics

The N6-Methyladenosine- (m6A-) Associated Genes Act as Strong Key Biomarkers for the Prognosis of Pancreatic Adenocarcinoma.
Function of N6-Methyladenosine Modification in Tumors.
Knockdown RBM15 Inhibits Colorectal Cancer Cell Proliferation and Metastasis Via N6-Methyladenosine (m6A) Modification of MyD88 mRNA.
The RNA N6-Methyladenosine Demethylase FTO Promotes Head and Neck Squamous Cell Carcinoma Proliferation and Migration by Increasing CTNNB1.
Enzymatic deamination of the epigenetic nucleoside N6-methyladenosine regulates gene expression.
ALKBH5-mediated m6A modification of lncRNA KCNQ1OT1 triggers the development of LSCC via upregulation of HOXA9.
HRD1-mediated METTL14 degradation regulates m6A mRNA modification to suppress ER proteotoxic liver disease.
Analysis of Multi-Layer RNA Modification Patterns for the Characterization of Tumor Immune Microenvironment in Hepatocellular Carcinoma.
Ezh2 promotes TRβ lysine methylation-mediated degradation in hepatocellular carcinoma.
ALKBH5-mediated m6A demethylation of KCNK15-AS1 inhibits pancreatic cancer progression via regulating KCNK15 and PTEN/AKT signaling.

Comput Math Methods Med. 2021 ;2021 8715823

The N6-Methyladenosine- (m6A-) Associated Genes Act as Strong Key Biomarkers for the Prognosis of Pancreatic Adenocarcinoma.

Fei Li, Ping Zhang.

Background: Pancreatic adenocarcinoma (PAAD) has become the major cause of cancer-related deaths globally. The m6A (N6-methyladenosine) alteration plays a crucial function in carcinogenesis and tumor progression. The role of genes related to m6A and their expression level in pancreatic cancer is not identified yet. The objective of this research analysis is a demonstration of the m6A RNA methylation regulators based as biomarkers for the PAAD diagnosis.
Methods: About 23 extensively reported m6A RNA methylation regulators were identified through the Cancer Genome Atlas (TCGA) database. This identification was based on consensus clustering analysis, protein-protein integration (PPI) analysis, risk prognostic model, Cox-regression analysis, String Spearman analysis, and LASSO Cox-regression.
Results: Herein, we conclude that 23 m6A methylation regulators have a strong link with the clinical and molecular characteristics of PAAD. The three subgroups (1/2) of pancreatic adenocarcinoma were identified using the clustering of 23 m6A regulators. Subgroup cluster 2 had a lower survival rate than the subgroup of cluster 1, and the difference in grades between the two groups was substantial. An assessment was performed using the 23 reported m6A methylation regulators. Eight of these can be used as independent PAAD prognostic markers. The consequences of variable IGF2BP3 expression in PAAD were then investigated further.
Conclusions: The key finding of this study was that the m6A methylation regulator gene has the main role in pancreatic tumors, and it may be used as a biomarker in the prognosis of the PAAD and for therapy purposes.

DOI: https://doi.org/10.1155/2021/8715823
J Oncol. 2021 ;2021 6461552

Function of N6-Methyladenosine Modification in Tumors.

Nan Zhang, Yuxin Zuo, Yu Peng, Lielian Zuo.

N6-Methyladenosine (m6A) modification is a dynamic and reversible methylation modification at the N6-position of adenosine. As one of the most prevalent posttranscriptional methylation modifications of RNA, m6A modification participates in several mRNA processes, including nuclear export, splicing, translation, and degradation. Some proteins, such as METTL3, METTL14, WTAP, ALKBH5, FTO, and YTHDF1/2/3, are involved in methylation. These proteins are subdivided into writers (METTL3, METTL14, WTAP), erasers (ALKBH5, FTO), and readers (YTHDF1/2/3) according to their functions in m6A modification. Several studies have shown that abnormal m6A modification occurs in tumors, including colorectal cancer, liver cancer, breast cancer, nasopharyngeal carcinoma, and gastric cancer. The proteins for m6A modification are involved in tumor proliferation, angiogenesis, metastasis, immunity, and other processes. Herein, the roles of m6A modification in cancer are discussed, which will improve the understanding of tumorigenesis, as well as the diagnosis, treatment, and prognosis of tumors.

DOI: https://doi.org/10.1155/2021/6461552
Cancer Biother Radiopharm. 2021 Nov 25.

Knockdown RBM15 Inhibits Colorectal Cancer Cell Proliferation and Metastasis Via N6-Methyladenosine (m6A) Modification of MyD88 mRNA.

Zhen Zhang, Yan Mei, Mingxing Hou.

  Colorectal cancer (CRC) is one of the most common cancers worldwide. In this study, we explored the role of RNA binding motif protein 15 (RBM15)-mediated MyD88 mRNA N6-methyladenosine (m6A) in CRC development. Cell proliferation, apoptosis, and invasion were detected by EdU, Annexin V-FITC/PI staining, and Transwell assays, respectively. RBM15 and MyD88 expression was detected by RT-qPCR and Western blot. RNA-seq, RIP-seq, and MeRIP-seq were used for RBM15 downstream target gene prediction and expression detection. In this research, we confirmed that RBM15 was highly expressed in CRC tissues and was negatively correlated with overall and disease-free survival rate. Silencing RBM15 significantly inhibited the proliferative and invasive abilities and promoted cell apoptosis in the CRC cell lines (SW480 and HCT116). Moreover, tumor growth and CRC liver metastasis were inhibited by silencing RBM15 in vivo. m6A methylation level was decreased in RBM15-silenced SW480 and HCT116 cells. MyD88 was the target mRNA of RBM15-mediated m6A methylation in CRC. MyD88 was lowly expressed in CRC and negatively correlated with RBM15 expression. Taken together, RBM15 silencing inhibited the CRC growth and metastasis in vitro and in vivo. RBM15 mediated m6A methylation modification of MyD88 mRNA in CRC cells.

Keywords:  MyD88; RBM15; RNA methylation; colorectal cancer; m6A

DOI:  https://doi.org/10.1089/cbr.2021.0226
Int J Gen Med. 2021 ;14 8785-8795

The RNA N6-Methyladenosine Demethylase FTO Promotes Head and Neck Squamous Cell Carcinoma Proliferation and Migration by Increasing CTNNB1.

Yu Zhang, Lixiao Chen, Xiaoliang Wu, Zhenfeng Sun, Fei Wang, Baoxin Wang, Pin Dong.

   Objective: In this study, we aimed to investigate the role of RNA N6-methyladenosine demethylase fat mass and obesity-associated protein (FTO) in head and neck squamous cell carcinoma (HNSCC).
Methods: Clinical data downloaded from The Cancer Genome Atlas (TCGA) database were used to analyze the relationship between mRNA levels of FTO, METTL3, METTL14, and ALKBH5, and the overall survival in cancer and para-cancer datasets. FTO expression in tumor and normal tissues was compared using immunohistochemistry, and its relationship with overall survival was analyzed based on the Kaplan-Meier method. The FaDu cell line with high FTO levels was chosen from five HNSCC cell lines for further experiments. FTO was verified as an oncogene in HNSCC by in vitro loss-of-function and overexpression studies, cell proliferation assay, wound healing assay, and identification of expression changes of epithelial-mesenchymal transition (EMT)-related markers. Catenin beta 1 (CTNNB1) was confirmed as a downstream target gene of FTO with additional methods like the GEPIA online tool, qRT-PCR, Western blotting, and dot blot assay.
Results: We found that FTO expression was significantly upregulated in HNSCC datasets and tissues. Increased FTO expression indicated a trend towards poor prognosis and was found to promote disease proliferation and migration. Mechanistically, cell proliferation assay, wound healing assay, and identification of expression changes of EMT-related markers demonstrated that FTO could act as an oncogene in HNSCC. FTO expression was significantly correlated with CTNNB1 expression. Moreover, it exerted a tumorigenic effect by increasing CTNNB1 expression in an m6A-dependent manner.
Conclusion: FTO promotes head and neck squamous cell carcinoma proliferation and migration by increasing CTNNB1 in an m6A-dependent manner.

Keywords:  CTNNB1; FTO; HNSCC; m6A

DOI:  https://doi.org/10.2147/IJGM.S339095
Nucleic Acids Res. 2021 Nov 24. pii: gkab1124. [Epub ahead of print]

Enzymatic deamination of the epigenetic nucleoside N6-methyladenosine regulates gene expression.

Zhuoran Jiang, Chao Wang, Zixin Wu, Kun Chen, Wei Yang, Hexiang Deng, Heng Song, Xiang Zhou.

N6-methyladenosine (m6A) modification is the most extensively studied epigenetic modification due to its crucial role in regulating an array of biological processes. Herein, Bsu06560, formerly annotated as an adenine deaminase derived from Bacillus subtilis 168, was recognized as the first enzyme capable of metabolizing the epigenetic nucleoside N6-methyladenosine. A model of Bsu06560 was constructed, and several critical residues were putatively identified via mutational screening. Two mutants, F91L and Q150W, provided a superiorly enhanced conversion ratio of adenosine and N6-methyladenosine. The CRISPR-Cas9 system generated Bsu06560-knockout, F91L, and Q150W mutations from the B. subtilis 168 genome. Transcriptional profiling revealed a higher global gene expression level in BS-F91L and BS-Q150W strains with enhanced N6-methyladenosine deaminase activity. The differentially expressed genes were categorized using GO, COG, KEGG and verified through RT-qPCR. This study assessed the crucial roles of Bsu06560 in regulating adenosine and N6-methyladenosine metabolism, which influence a myriad of biological processes. This is the first systematic research to identify and functionally annotate an enzyme capable of metabolizing N6-methyladenosine and highlight its significant roles in regulation of bacterial metabolism. Besides, this study provides a novel method for controlling gene expression through the mutations of critical residues.

DOI: https://doi.org/10.1093/nar/gkab1124
J Cell Mol Med. 2021 Dec 01.

ALKBH5-mediated m6A modification of lncRNA KCNQ1OT1 triggers the development of LSCC via upregulation of HOXA9.

Yushan Li, Bingrui Yan, Xin Wang, Qiuying Li, Xuan Kan, Jingting Wang, Yanan Sun, Peng Wang, Linli Tian, Ming Liu.

  It has been shown that N6-methyladenosine (m6A) modification is involved in the development of complex human diseases, especially in the development of cancer. Our research investigated the role and mechanism of the m6A modification of lncRNA KCNQ1 overlapping transcript 1 (KCNQ1OT1) in Laryngeal squamous cell carcinoma (LSCC) progression. Microarray analysis was used to quantitatively detect the m6A apparent transcriptional modification level of lncRNA in LSCC tissue. Methylated RNA immunoprecipitation-qPCR (MeRIP-qPCR), in situ hybridization (ISH) and quantitative real-time PCR (qRT-PCR) were used to examine the m6A modification and expression of KCNQ1OT1. In addition, in vivo and in vitro experiments have tested the effects of KCNQ1OT1 knockdown on the proliferation, invasion and metastasis of LSCC. Mechanically, we found the N6-methyladenosine (m6A) demethylase ALKBH5 mediates KCNQ1OT1 expression via an m6A-YTHDF2-dependent manner and KCNQ1OT1 could directly bind to HOXA9 to further regulate the proliferation, invasion and metastasis of LSCC cells. In general, our research indicates that ALKBH5-mediated m6A modification of KCNQ1OT1 triggers the development of LSCC via upregulation of HOXA9.

Keywords:  ALKBH5; HOXA9; laryngeal squamous cell cancer; lncRNA KCNQ1OT1; m6A methylation

DOI:  https://doi.org/10.1111/jcmm.17091
Mol Cell. 2021 Nov 23. pii: S1097-2765(21)00948-5. [Epub ahead of print]

HRD1-mediated METTL14 degradation regulates m6A mRNA modification to suppress ER proteotoxic liver disease.

Juncheng Wei, Bryan T Harada, Dan Lu, Ruihua Ma, Beixue Gao, Yanan Xu, Elena Montauti, Nikita Mani, Shuvam M Chaudhuri, Shana Gregory, Samuel E Weinberg, Donna D Zhang, Richard Green, Chuan He, Deyu Fang.

Accumulation of unfolded or misfolded proteins in the endoplasmic reticulum (ER) lumen triggers an unfolded protein response (UPR) for stress adaptation, the failure of which induces cell apoptosis and tissue/organ damage. The molecular switches underlying how the UPR selects for stress adaptation over apoptosis remain unknown. Here, we discovered that accumulation of unfolded/misfolded proteins selectively induces N6-adenosine-methyltransferase-14 (METTL14) expression. METTL14 promotes C/EBP-homologous protein (CHOP) mRNA decay through its 3' UTR N6-methyladenosine (m6A) to inhibit its downstream pro-apoptotic target gene expression. UPR induces METTL14 expression by competing against the HRD1-ER-associated degradation (ERAD) machinery to block METTL14 ubiquitination and degradation. Therefore, mice with liver-specific METTL14 deletion are highly susceptible to both acute pharmacological and alpha-1 antitrypsin (AAT) deficiency-induced ER proteotoxic stress and liver injury. Further hepatic CHOP deletion protects METTL14 knockout mice from ER-stress-induced liver damage. Our study reveals a crosstalk between ER stress and mRNA m6A modification pathways, termed the ERm6A pathway, for ER stress adaptation to proteotoxicity.

DOI: https://doi.org/10.1016/j.molcel.2021.10.028
Front Cell Dev Biol. 2021 ;9 761391

Analysis of Multi-Layer RNA Modification Patterns for the Characterization of Tumor Immune Microenvironment in Hepatocellular Carcinoma.

Jiyuan Xing, Shen Shen, Zihui Dong, Xiaobo Hu, Lixia Xu, Xiaorui Liu, Qinggang Li, Yize Zhang, Gangying Cui, Zujiang Yu.

  Background: RNA modifications have emerged as important posttranscriptional changes in multiple tumor cellular processes and tumorigenesis, including hepatocellular carcinoma (HCC). However, the potential roles and the interaction between regulators of RNA modifications and the tumor microenvironment (TME) are unclear in HCC. Methods: The gene expression profiles of 26 RNA modification "writers" were investigated in the TCGA cohort. The unsupervised clustering approach was used to class these RNA modification regulators. The characteristics of immune cell infiltration from TME for each cluster was tested by the CIBERSORT method. Additionally, we established a scoring model to evaluate the RNA modification characteristics of individual tumors. The associations between the scoring model and genetic as well as clinical characteristics, drug sensitivity, and response to immunotherapy were also analyzed. Results: We mapped the somatic mutations and somatic copy number variation of the RNA modification regulators. The expression of all selected regulators was detected, and two modification patterns were identified that featured distinct immune cell infiltration characteristics. Subsequently, we developed a score model (termed as WM-Score model). Furthermore, the survival analysis showed that the WM-Score value was associated with HCC patient prognosis. The results of the ROC curves analysis and multivariate analysis all confirmed that the WM-Score value was strongly associated with anti-cancer drug resistance and therapeutic efficacy of immunotherapy, thus could be used as an independent risk factor in HCC. Conclusion: Our research identified two RNA modification patterns characterized by distinct TME, and the WM-Score model was developed that might serve as reliable prognostic and immunotherapeutic effect predictor of HCC.

Keywords:  RNA methylation modification; TME; hepatocellular carcinoma; immunotherapy; prognosis

DOI:  https://doi.org/10.3389/fcell.2021.761391
Genes Genomics. 2021 Dec 01.

Ezh2 promotes TRβ lysine methylation-mediated degradation in hepatocellular carcinoma.

Su Chan Park, Ji Min Lee.

   BACKGROUND: Post-translational modification (PTM) of proteins controls various cellular functions of transcriptional regulators and participates in diverse signal transduction pathways in cancer. The thyroid hormone (triiodothyronine, T3) plays a critical role in metabolic homeostasis via its direct interaction with the thyroid hormone receptor beta (TRβ). TRβ is involved in physiological processes, such as cell growth, differentiation, apoptosis, and maintenance of metabolic homeostasis through transcriptional regulation of target genes.
OBJECTIVE: This study was performed to characterize the specific PTM of TRβ is an active control mechanism for the proteasomal degradation of TRβ in transcriptional signaling pathways in hepatocellular carcinoma cells.
METHODS: Based on a previous study, we predicted that the lysine methyltransferase and methylation sites of TRβ by comparing the amino acid sequences of histone H3 and TRβ. Methyl-acceptor site of TRβ was confirmed by point mutation. TRβ protein stability was evaluated by ubiquitination assay with MG132. For glucose starvation, HepG2 cells were incubated in media without D-glucose. Proliferation-related proteins were detected by western blotting. MicroRNA level and autophagy marker were measured by real-time qPCR.
RESULTS: The presence of enhancer of zeste homolog 2 (Ezh2), a methyltransferase of H3 lysine 27, as a methyltransferase of TRβ also revealed that direct lysine methylation and consequent stimulated protein degradation of TRβ underlies the negative correlation between Ezh2 and TRβ. Notably, glucose starvation significantly increased lysine methylation, and methylated TRβ showed further protein instability leading to an increase in the proliferation and growth of hepatocellular carcinoma cells.
CONCLUSIONS: TRβ functions as a tumor suppressor in various cancers; therefore, we evaluated the effect of TRβ degradation on oncogenesis during glucose starvation. These data clearly define a functional model and provide a link between metabolism and cancer by regulating methyl-dependent protein levels of tumor suppressors. Taken together, maintaining TRβ against methyl-dependent degradation is considered a possible therapeutic target for cancer progression.

Keywords:  Hepatocellular carcinoma; Metabolic homeostasis; Post-translational modification; Thyroid hormone; Transcriptional regulator; Tumor suppressor

DOI:  https://doi.org/10.1007/s13258-021-01196-8
Cell Death Dis. 2021 Dec 01. 12(12): 1121

ALKBH5-mediated m6A demethylation of KCNK15-AS1 inhibits pancreatic cancer progression via regulating KCNK15 and PTEN/AKT signaling.

Yuan He, HongQin Yue, Ying Cheng, Zhilong Ding, Zhen Xu, Chunyang Lv, Zheng Wang, Jing Wang, Chenglong Yin, Huihui Hao, Chuang Chen.

Long noncoding RNAs (lncRNAs) are regarded as crucial regulators in tumor progression. Potassium two pore domain channel subfamily K member 15 and WISP2 antisense RNA 1 (KCNK15-AS1) has been confirmed to inhibit the migration and invasion of pancreatic cancer (PC) cells. However, its downstream mechanism and effect on other cellular functions in PC remain unknown. This study probed the function and potential mechanism of KCNK15-AS1 in PC cell growth. RT-qPCR and western blot were employed to measure gene expression in PC cells. ISH was applied to analyze KCNK15-AS1 expression in PC tissues. Functional assays were utilized to evaluate PC cell proliferation, apoptosis, migration and EMT. Mechanical experiments were adopted to detect gene interaction in PC cells. The obtained data indicated that KCNK15-AS1 was down-regulated in PC cells and tissues. Overexpressing KCNK15-AS1 hindered cell proliferation, migration and EMT while facilitated cell apoptosis in PC. Mechanically, alkylation repair homolog protein 5 (ALKBH5) was verified to induce m6A demethylation of KCNK15-AS1 to mediate KCNK15-AS1 up-regulation. KCNK15-AS1 combined with KCNK15 5'UTR to inhibit KCNK15 translation. Moreover, KCNK15-AS1 recruited MDM2 proto-oncogene (MDM2) to promote RE1 silencing transcription factor (REST) ubiquitination, thus transcriptionally upregulating phosphatase and tensin homolog (PTEN) to inactivate AKT pathway. In conclusion, our study first confirmed that KCNK15-AS1 hinders PC cell growth by regulating KCNK15 and PTEN, suggesting KCNK15-AS1 as a potential biomarker of PC.

DOI: https://doi.org/10.1038/s41419-021-04401-4