N(6)-Methyladenosine-Modified ATP8B1-AS1 Exerts Oncogenic Roles in Hepatocellular Carcinoma via Epigenetically Activating MYC

PURPOSE: N(6)-methyladenosine (m(6)A) modification has shown critical roles in regulating mRNA fate. Non-coding RNAs also have important roles in various diseases, including hepatocellular carcinoma (HCC). However, the potential influences of m(6)A modification on non-coding RNAs are still unclear. In this study, we identified a novel m(6)A-modified ATP8B1-AS1 and aimed to investigate the effects of m(6)A on the expression and role of ATP8B1-AS1 in HCC. METHODS: qPCR was performed to measure the expression of related genes. The correlation between gene expression and prognosis was analyzed using public database. m(6)A modification level was measured using MeRIP and single-base elongation- and ligation-based qPCR amplification method. The roles of ATP8B1-AS1 in HCC were investigated using in vitro and in vivo functional assays. The mechanisms underlying the roles of ATP8B1-AS1 were investigated by ChIRP and ChIP assays. RESULTS: ATP8B1-AS1 is highly expressed in HCC tissues and cell lines. High expression of ATP8B1-AS1 is correlated with poor overall survival of HCC patients. ATP8B1-AS1 is m(6)A modified and the 792 site of ATP8B1-AS1 is identified as an m(6)A modification site. m(6)A modification increases the stability of ATP8B1-AS1 transcript. m(6)A modification level of ATP8B1-AS1 is increased in HCC tissues and cell lines, and correlated with poor overall survival of HCC patients. ATP8B1-AS1 promotes HCC cell proliferation, migration, and invasion, which were abolished by the mutation of m(6)A-modified 792 site. Mechanistic investigation revealed that m(6)A-modified ATP8B1-AS1 interacts with and recruits m(6)A reader YTHDC1 and histone demethylase KDM3B to MYC promoter region, leading to the reduction of H3K9me2 level at MYC promoter region and activation of MYC transcription. Functional rescue assays showed that depletion of MYC largely abolished the oncogenic roles of ATP8B1-AS1. CONCLUSION: m(6)A modification level of ATP8B1-AS1 is increased and correlated with poor prognosis in HCC. m(6)A-modified ATP8B1-AS1 exerts oncogenic roles in HCC via epigenetically activating MYC expression.