The hepatitis B virus X protein (HBx) has been implicated in HBV-associated development of hepatocellular carcinom (HCC) by both epigenetic modifications and genetic regulations. Recently, long noncoding RNAs (lncRNAs) were found to be dysregulated in a variety of tumors and play critical regulatory roles in cancer biology. In the present study, we aimed to investigate the effects of HBx on lncRNAs expression and the underlying molecular mechanisms in hepatocarcinogenesis.
From differential expression profile of lncRNAs induced by HBx, lncRNA HBAL, which is HBx associated lncRNA, was a candidate to assess its potential function by silencing and enforced expressing lncRNA in vitro and in vivo. Cell proliferation ability of transfected cells was detected by CCK-8 assays, and cell-cycle and apoptosis were measured by flow cytometry analysis. Interaction of lncRNA HBAL with EZH2 was determined by RIP assay. The binding of EZH2 and 3MeK27H3 level in the exon 1 of p27 were measured through CHIP assay. RNA fractions of nuclear and cytosolic were separated using the PARIS Kit.
Among of upregulated 379 and downregulated 724 lncRNAs induced by HBx, an upregulated lncRNA induced by HBx was selected to explore its function in HBV-related hepatocellular carcinogenesis. In HCC tissues, the expression levels of HBAL is positively associated with HBx indicates its important roles in HCC. Enforced HBAL promotes HCC cells growth through altering cell apoptosis rate by caspase 3,8 cleaved and increase G1/S transition in vitro. In animal model, knockdown of lncRNA HBAL significantly inhibited the HCC tumor growth in vivo. Screening cell cycle regulators expression profile, CDK2 was elevated in HBAL transfected cells, while, p27, a negative regulator of CDK2, was epigenetic repressed via recruiting EZH2, a key component of PRC2. An inverse correlation between the mRNA expression of p27 and lncRNA HBAL was observed in HCC tissues.
This study is the first time report that HBx upregulates lncRNA HBAL expression to promote tumor cells growth, indicating that lncRNA HBAL is a potential oncogenic lncRNA involved in tumor progression. These data provides a broader perspective into the role of HBx in hepatocarcinogenesis.
All authors have declared no conflicts of interest.