HBV-encoded miR-2 functions as an oncogene by downregulating TRIM35 but upregulating RAN in liver cancer cells

BACKGROUND: Hepatitis B virus (HBV) infection has been well established as a high-risk factor for the carcinogenesis of hepatocellular carcinoma (HCC). Cellular microRNA (miRNA) is involved in tumorigenesis by accelerating the malignant phenotype in HCC. However, whether HBV can encode miRNAs that contribute to HCC is not entirely clear. METHODS: In this study, an miRNA encoded by HBV (HBV-miR-2) was identified by Solexa sequencing in HBV-positive HCC specimens and further verified in serum samples from HCC patients with HBV infection and in HBV-positive HCC cell lines. To evaluate the roles of HBV-miR-2 in liver cancer cells, we determined cell viability and migration/invasion ability by gain-of-function experiment in HBV(−) liver cancer cells (HepG2 and Huh7) and loss-of-function experiments in Huh7 cells stably expressing HBV-miR-2 (Huh7/HBV-miR-2 cells) and HepG2.2.15 cells. Furthermore, to elucidate the mechanism by which HBV-miR-2 work on cell malignancy, we identified and studied the effect of two target genes (TRIM35 and RAN) of HBV-miR-2 in liver cancer cells. FINDINGS: We revealed that HBV-miR-2 promoted HCC cell growth ability by suppressing apoptosis and promoting migration and invasion by enhancing the epithelial-mesenchymal transition (EMT), functioning as an oncogene in the development of HBV-related HCC. Furthermore, we demonstrated that HBV-miR-2 suppresses the expression of TRIM35 but enhances RAN expression by targeting their 3′-untranslated regions (3’UTR) and that the ectopic expression of TRIM35 or knockdown of RAN counteracted the malignant phenotypes induced by HBV-miR-2. INTERPRETATION: Our findings indicate that an HBV-encoded miRNA, HBV-miR-2, promotes oncogenic activity by downregulating TRIM35 expression and upregulating RAN expression in liver cancer cells, likely providing insight into tumorigenesis in HBV-related liver cancer. FUND: This work was supported in part by the National Natural Science Foundation of China (No: 81830094; 91629302; 31270818) and the Natural Science Foundation of Tianjin (No: 12JCZDJC25100).