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?-catenin represses miR455-3p to stimulate m6A modification of HSF1 mRNA and promote its translation in colorectal cancer.

ABSTRACT: BACKGROUND:Heat shock transcription factor1 (HSF1) was overexpressed to promote glutaminolysis and activate mTOR in colorectal cancer (CRC). Here, we investigated the mechanism for cancer-specific overexpression of HSF1. METHODS:HSF1 expression was analyzed by chromatin immunoprecipitation, qRT-PCR, immunohistochemistry staining and immunoblotting. HSF1 translation was explored by polysome profiling and nascent protein analysis. Biotin pulldown and m6A RNA immunoprecipitation were applied to investigate RNA/RNA interaction and m6A modification. The relevance of HSF1 to CRC was analyzed in APCmin/+ and APCmin/+ HSF1+/-mice. RESULTS:HSF1 expression and activity were reduced after the inhibition of WNT/?-catenin signaling by pyrvinium or ?-catenin knockdown, but elevated upon its activation by lithium chloride (LiCl) or ?-catenin overexpression. There are much less upregulated genes in HSF1-KO MEF treated with LiCl when compared with LiCl-treated WT MEF. HSF1 protein expression was positively correlated with ?-catenin expression in cell lines and primary tissues. After ?-catenin depletion, HSF1 mRNA translation was impaired, accompanied by the reduction of its m6A modification and the upregulation of miR455-3p, which can interact with 3'-UTR of HSF1 mRNA to repress its translation. Interestingly, inhibition of miR455-3p rescued ?-catenin depletion-induced reduction of HSF1 m6A modification and METTL3 interaction. Both the size and number of tumors were significantly reduced in APCmin/+ mice when HSF1 was genetically knocked-out or chemically inhibited. CONCLUSIONS:?-catenin suppresses miR455-3p generation to stimulate m6A modification and subsequent translation of HSF1 mRNA. HSF1 is important for ?-catenin to promote CRC development. Targeting HSF1 could be a potential strategy for the intervention of ?-catenin-driven cancers.

SUBMITTER: Song P

PROVIDER: S-EPMC7446108 | biostudies-literature | 2020 Aug

REPOSITORIES: biostudies-literature

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β-catenin represses miR455-3p to stimulate m6A modification of HSF1 mRNA and promote its translation in colorectal cancer.

Song Ping P Feng Lifeng L Li Jiaqiu J Dai Dongjun D Zhu Liyuan L Wang Chaoqun C Li Jingyi J Li Ling L Zhou Qiyin Q Shi Rongkai R Wang Xian X Jin Hongchuan H

Molecular cancer 20200824 1

<h4>Background</h4>Heat shock transcription factor1 (HSF1) was overexpressed to promote glutaminolysis and activate mTOR in colorectal cancer (CRC). Here, we investigated the mechanism for cancer-specific overexpression of HSF1.<h4>Methods</h4>HSF1 expression was analyzed by chromatin immunoprecipitation, qRT-PCR, immunohistochemistry staining and immunoblotting. HSF1 translation was explored by polysome profiling and nascent protein analysis. Biotin pulldown and m6A RNA immunoprecipitation were ...[more]

PMID: 32838807

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Project description:Gastric cancer (GC) is the fifth most common cancer and the third deadliest cancer in the world, and the occurrence and development of GC are influenced by epigenetics. Methyltransferase-like 3 (METTL3) is a prominent RNA n6-adenosine methyltransferase (m6A) that plays an important role in tumor growth by controlling the work of RNA. This study aimed to reveal the biological function and molecular mechanism of METTL3 in GC. The expression level of METTL3 in GC tissues and cells was detected by qPCR, Western blot and immunohistochemistry, and the expression level and prognosis of METTL3 were predicted in public databases. CCK-8, colony formation, transwell and wound healing assays were used to study the effect of METTL3 on GC cell proliferation and migration. In addition, the enrichment effect of METTL3 on DEK mRNA was detected by the RIP experiment, the m6A modification effect of METTL3 on DEK was verified by the MeRIP experiment and the mRNA half-life of DEK when METTL3 was overexpressed was detected. The dot blot assay detects m6A modification at the mRNA level. The effect of METTL3 on cell migration ability in vivo was examined by tail vein injection of luciferase-labeled cells. The experimental results showed that METTL3 was highly expressed in GC tissues and cells, and the high expression of METTL3 was associated with a poor prognosis. In addition, the m6A modification level of mRNA was higher in GC tissues and GC cell lines. Overexpression of METTL3 in MGC80-3 cells and AGS promoted cell proliferation and migration, while the knockdown of METTL3 inhibited cell proliferation and migration. The results of in vitro rescue experiments showed that the knockdown of DEK reversed the promoting effects of METTL3 on cell proliferation and migration. In vivo experiments showed that the knockdown of DEK reversed the increase in lung metastases caused by the overexpression of METTL3 in mice. Mechanistically, the results of the RIP experiment showed that METTL3 could enrich DEK mRNA, and the results of the MePIP and RNA half-life experiments indicated that METTL3 binds to the 3'UTR of DEK, participates in the m6A modification of DEK and promotes the stability of DEK mRNA. Ultimately, we concluded that METTL3 promotes GC cell proliferation and migration by stabilizing DEK mRNA expression. Therefore, METTL3 is a potential biomarker for GC prognosis and a therapeutic target.

Project description:Introduction: Middle ear cholesteatoma is characterized by the hyperproliferation of keratinocytes. In recent decades, N6-methyladenosine (m6A) modification has been shown to play an essential role in the pathogenesis of many proliferative diseases. However, neither the m6A modification profile nor its potential role in the pathogenesis of middle ear cholesteatoma has currently been investigated. Therefore, this study aimed to explore m6A modification patterns in middle ear cholesteatoma. Materials and methods: An m6A mRNA epitranscriptomic microarray analysis was performed to analyze m6A modification patterns in middle ear cholesteatoma tissue (n = 5) and normal post-auricular skin samples (n = 5). Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses were performed to predict the potential biological functions and signaling pathways underlying the pathogenesis of middle ear cholesteatoma. Subsequently, m6A modification levels were verified by methylated RNA immunoprecipitation–qPCR (MeRIP–qPCR) in middle ear cholesteatoma tissue and normal skin samples, respectively. Results: A total of 6,865 distinctive m6A-modified mRNAs were identified, including 4,620 hypermethylated and 2,245 hypomethylated mRNAs, as well as 9,162 differentially expressed mRNAs, including 4,891 upregulated and 4,271 downregulated mRNAs, in the middle ear cholesteatoma group relative to the normal skin group. An association analysis between methylation and gene expression demonstrated that expression of 1,926 hypermethylated mRNAs was upregulated, while expression of 2,187 hypomethylated mRNAs and 38 hypermethylated mRNAs was downregulated. Moreover, GO analysis suggested that differentially methylated mRNAs might influence cellular processes and biological behaviors, such as cell differentiation, biosynthetic processes, regulation of molecular functions, and keratinization. KEGG pathway analysis demonstrated that the hypermethylated transcripts were involved in 26 pathways, including the Hippo signaling pathway, the p53 signaling pathway, and the inflammatory mediator regulation of transient receptor potential (TRP) channels, while the hypomethylated transcripts were involved in 13 pathways, including bacterial invasion of epithelial cells, steroid biosynthesis, and the Hippo signaling pathway. Conclusion: Our study presents m6A modification patterns in middle ear cholesteatoma, which may exert regulatory roles in middle ear cholesteatoma. The present study provides directions for mRNA m6A modification-based research on the epigenetic etiology and pathogenesis of middle ear cholesteatoma.

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?-catenin represses miR455-3p to stimulate m6A modification of HSF1 mRNA and promote its translation in colorectal cancer.

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β-catenin represses miR455-3p to stimulate m6A modification of HSF1 mRNA and promote its translation in colorectal cancer.

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