ALKBH5-mediated m6A demethylation of lncRNA PVT1 plays an oncogenic role in osteosarcoma

Abstract

Background: Osteosarcoma (OS) is one of the most common malignant bone tumors. Plasmacytoma variant translocation 1 (PVT1) is a well-known oncogenic long noncoding RNA (lncRNA). However, to date, the regulatory mechanism of PVT1 upregulation in OS remains unknown.Methods: qRT-PCR was carried out to test the expression level of PVT1 and ALKBH5. RNA immunoprecipitation (RIP) and RNA pull-down assays were performed to detect the interaction of PVT1 with ALKBH5 and YTHDF2. Methylated RNA immune-precipitation (MeRIP) was used to examine the N 6 -methyladenosine (m 6 A) modification of PVT1 transcript. Results:In this study, we found that PVT1 expression was upregulated in OS tissues and cells and significantly related with clinical stage, tumor size, and prognosis of patients with OS. Further investigation revealed that N 6 -methyladenosine (m 6 A) demethylase ALKBH5 could associate with PVT1 and suppress its degradation. ALKBH5 decreased the m 6 A modification of PVT1, thus inhibiting the binding of reader protein YTHDF2 in PVT1. Functionally, ALKBH5-mediated PVT1 upregulation promoted the OS cell proliferation in vitro and tumor growth in vivo. Conclusions:Our study suggests that ALKBH5-mediated m 6 A modification of PVT1 contributes to OS tumorigenesis.

The biological function of m6A demethylase ALKBH5 and its role in human disease

“…As for other cancers, ALKBH5 was significantly downregulated in clear cell renal cell carcinoma (ccRCC) compared to normal tissue [45]. It was also affirmed that ALKBH5 took a great part in contributing to osteosarcoma tumorigenesis [40]. ALKBH5, directly regulated by human RNA helicase DDX3, decreased m6A methylation in FOXM1 and NANOG nascent transcript and contributed to chemoresistance of oral squamous cell carcinoma(OSCC) [46].…”

“…Moreover, ALKBH5 demethylated lncRNA KCNK15-AS1, which was downregulated in pancreatic cancer tissues compared with the normal tissues, thus elevating pancreatic cancer motility, including migration and invasion [26]. In addition, ALKBH5 suppressed the degradation of lncRNA plasmacytoma variant translocation 1 (PVT1), which was up-regulated in osteosarcoma tissues and cells, thus promoting the osteosarcoma cell proliferation in vitro and tumor growth in vivo through inhibiting the binding of m6A reader YTHDF2 in PVT1 [40]. ALKBH5 decreased the level of lncRNA nuclear paraspeckle assembly transcript 1 (NEAT1), which was overexpressed in gastric cancer cells and tissue, and took a great part in the invasion and metastasis of gastric cancer [19].…”

“…Chu et al [32] and Feng et al [33] respectively identified the crystal structure of zebrafish ALKBH5 (fALKBH5) and human ALKBH5, which promoted the understanding of the substrate recognition specificity of ALKBH5 and offered a foundation for the development of inhibitors selective for NAOX. In addition, Xu et al [34] affirmed Clear cell renal cell carcinoma Downregulated --- [45] Osteosarcoma --Proliferation and tumor growth PVT1 [40] Oral squamous cell carcinoma --Chemoresistance FOXM1, NANOG [46] the m6A binding pocket of ALKBH5 and the key residues related with m6A recognition through mutagenesis and isothermal titration calorimetry binding experiments. Furthermore, m6A served as a 'conformational marker' , which induced different conformational outcomes in RNAs depending on sequence context and critically impacted its interactions with ALKBH5 [35].…”

Multifaceted Functions and Novel Insight Into the Regulatory Role of RNA N6-Methyladenosine Modification in Musculoskeletal Disorders

“…The aberrantly expressed genes were associated with pluripotency regulation of the OSCs ( Wang J. et al, 2019 ). Further, it has been shown that lncRNA plasmacytoma variant translocation 1 (PVT1) transcript was upregulated because of m 6 A methylation decrease mediated by ALKBH5, which reduced the binding of reader protein YTHDF2 in PVT1, subsequently lessening the degradation of PVT1 and promoting tumorigenesis of OS ( Chen et al, 2020 ). METTL3 was found localizing in both cytoplasm and nucleus of OS cells, which is also on the map of OS modulation.…”

Joint analysis of lncRNA m6A methylome and lncRNA/mRNA expression profiles in gastric cancer

“…For instance, METTL3-mediated m 6 A modification led to LINC00958 upregulation through stabilizing its RNA transcript, and LINC00968 sponged miR-3619-5p to upregulate HDGF expression thereby facilitating the lipogenesis and progression of hepatocellular carcinoma (HCC) [ 27 ]. Another investigation revealed that m 6 A demethylase ALKBH5 could suppress the degradation of lncRNA PVT1, and its overexpression promoted osteosarcoma cell proliferation in vitro and tumor growth in vivo [ 28 ]. Similar phenomenon could also be observed in GC-related lncRNAs with m 6 A [ 29 ].…”