N6-methyladenosine (m6A) modification is the most common chemical modification in mammalian mRNAs, and it plays important roles by regulating several cellular processes. Previous studies report that m6A is implicated in modulating tumorigenesis and progression. However, dysregulation of m6A modification and effect of m6A demethylase fat-mass and obesity-associated protein (FTO) on glucose metabolism has not been fully elucidated in papillary thyroid cancer (PTC).Quantitative real-time PCR (qRT-PCR), western blotting and immunohistochemistry were performed to explore the expression profile of FTO in PTC tissues and adjacent non-cancerous thyroid tissues. Effects of FTO on PTC glycolysis and growth were investigated through in vitro and in vivo experiments. Mechanism of FTO-mediated m6A modification was explored through transcriptome-sequencing (RNA-seq), methylated RNA immunoprecipitation sequencing (MeRIP-seq), MeRIP-qPCR, luciferase reporter assays, RNA stability assay and RNA immunoprecipitation assay.FTO expression was significantly downregulated in PTC tissues. Functional analysis showed that FTO inhibited PTC glycolysis and growth. Further analyses were conducted to explore FTO-mediated m6A modification profile in PTC cells and Apolipoprotein E (APOE) was identified as the target gene for FTO-mediated m6A modification using RNA-seq and MeRIP-seq. FTO knockdown significantly increased APOE mRNA m6A modification and upregulated its expression. FTO-mediated m6A modification of APOE mRNA was recognized and stabilized by the m6A reader IGF2BP2. The findings showed that APOE also promoted tumor growth through glycolysis in PTC. Analysis showed that FTO/APOE axis inhibits PTC glycolysis by modulating IL-6/JAK2/STAT3 signaling pathway.FTO acts as a tumor suppressor to inhibit tumor glycolysis in PTC. The findings of the current study showed that FTO inhibited expression of APOE through IGF2BP2-mediated m6A modification and may inhibit glycolytic metabolism in PTC by modulating IL-6/JAK2/STAT3 signaling pathway, thus abrogating tumor growth. 

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FTO suppresses glycolysis and growth of papillary thyroid cancer via decreasing stability of APOE mRNA in an N6-methyladenosine-dependent manner

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N6-Methyladenosine (m6A) modification is the most abundant RNA modification in eukaryotic cells. IGF2BP3, a well-known m6A reader, is deregulated in many cancers, but its role in nasopharyngeal carcinoma (NPC) remains unclear. In this work, IGF2BP3 was upregulated in NPC tissues and cells. The high level of IGF2BP3 was positively related to late clinical stages, node metastasis, and poor outcomes. Moreover, IGF2BP3 accelerated NPC cell tumor progression and metastasis in vitro and vivo. Upstream mechanism analyses indicated that the high expression of IGF2BP3 in head and neck tumors was mainly due to mRNA level amplification. Luciferase assay and chromatin immunoprecipitation assay (CHIP) depicted that MYC was effectively bound to the promoter of IGF2BP3, thereby improving its transcriptional activity. Results also showed that IGF2BP3 was not only positively correlated with KPNA2 expression but also modulated the expression of KPNA2. m6A RNA immunoprecipitation (MeRIP) and RNA stability experiments verified that silencing IGF2BP3 significantly inhibited the m6A modification level of KPNA2, thereby stabilizing the mRNA stability of KPNA2. Rescue experiments proved that the effect of inhibiting or overexpressing IGF2BP3 on NPC cells was partly reversed by KPNA2. Collectively, MYC-activated IGF2BP3 promoted NPC cell proliferation and metastasis by influencing the stability of m6A-modified KPNA2. Our findings offer new insights that IGF2BP3 may serve as a new molecular marker and potential therapeutic target for NPC treatment. 

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MYC-activated RNA N6-methyladenosine reader IGF2BP3 promotes cell proliferation and metastasis in nasopharyngeal carcinoma

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The N6-methyladenosine (m6A) RNA modification can regulate autophagy to modulate the growth and development of tumors, but the mechanism of m6A modification for the regulation of autophagy in hepatocellular carcinoma cells (HCC) remains unclear. In the study, the knockdown of the Wilms’ tumor 1-associating protein (WTAP) was made in HCC to study the correlation between m6A modification and autophagy. A fluorescent confocal microscopy analysis showed that the knockdown of WTAP could facilitate the autophagy of HCC. A Western blot analysis showed that the level of p-AMPK was decreased in WTAP-knockdown HCC cells. Additionally, LKB1, the upstream kinase of AMPK, was regulated by WTAP and it could mediate the phosphorylation of AMPK in an m6A-dependent manner. Further studies revealed that the knockdown of WTAP could reduce the level of LKB1 mRNA with m6A. This could result in the increased stability of LKB1 mRNA to promote its expression. The knockdown of WTAP could upregulate the level of autophagy and inhibit HCC proliferation. However, the overexpression of WTAP could resist autophagic cell death.

m6A mRNA Methylation Regulates LKB1 to Promote Autophagy of Hepatoblastoma Cells through Upregulated Phosphorylation of AMPK.

The m6A RNA methylation is the most prevalent internal modification in mammalian mRNAs which plays critical biological roles by regulating vital cellular processes Dysregulations of the m6A modification due to aberrant expression of its regulatory proteins are frequently observed in many pathological conditions, particularly in cancer Normal cells undergo malignant transformation via activation or modulation of different oncogenic signaling pathways through complex mechanisms Accumulating evidence showing regulation of oncogenic signaling pathways at the epitranscriptomic level has added an extra layer of the complexity In particular, recent studies demonstrated that, in many types of cancers various oncogenic signaling pathways are modulated by the m6A modification in the target mRNAs as well as noncoding RNA transcripts m6A modifications in these RNA molecules control their fate and metabolism by regulating their stability, translation or subcellular localizations In this review we discussed recent exciting studies on oncogenic signaling pathways that are modulated by the m6A RNA modification and/or their regulators in cancer and provided perspectives for further studies The regulation of oncogenic signaling pathways by the m6A modification and its regulators also render them as potential druggable targets for the treatment of cancer

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Chengfeng Yang

Papers

The m 6 A RNA methylation regulates oncogenic signaling pathways driving cell malignant transformation and carcinogenesis