RESUMEN
Hepatocellular carcinoma (HCC) is one of the most prevalent cancers. It has been demonstrated that various cellular microRNAs (miRNAs) play an important role in HCC development. Here, we analyzed the miRNA profile in HCC tissues by Solexa sequencing, and we identified a novel microRNA, miR-HCC1, which is upregulated in HCC tissues. Further experiments showed that miR-HCC1 promoted HCC cell proliferation in vivo and in vitro, and migration and invasion resulting from the epithelial-mesenchymal transition (EMT) process. Nuclear factor I/X (NFIX), which inhibited cell proliferation, migration and invasion in HCC cells, was identified as a direct and functional target of miR-HCC1. Furthermore, lymphoid enhancer binding factor 1 (LEF1), a transcription factor, was shown to bind the promoter of miR-HCC1 and activate its expression. Collectively, these results indicate that LEF1-upregulated miR-HCC1 functions as an oncogene through the negative regulation of NFIX expression, which links the LEF1/miR-HCC1/NFIX axis to contribute to cell proliferation, migration and invasion of HCC cells and could provide novel insights into miRNA function and hepatocarcinogenesis and potential biomarkers for HCC.
RESUMEN
BACKGROUND: Occurrence and progression of hepatocellular carcinoma (HCC) are associated with hepatitis B virus (HBV) infection. miR-1269b is up-regulated in HCC cells and tissues. However, the regulation of miR-1269b expression by HBV and the mechanism underlying the oncogenic activity of miR-1269b in HCC are unclear. METHODS: Reverse transcription quantitative PCR (RT-qPCR) was used to measure the expression of miR-1269b and target genes in HCC tissues and cell lines. Western blot analysis was used to assess the expression of miR-1269b target genes and related proteins. Using luciferase reporter assays and EMSA, we identified the factors regulating the transcriptional level of miR-1269b. Colony formation, flow cytometry and cell migration assays were performed to evaluate the phenotypic changes caused by miR-1269b and its target in HCC cells. RESULTS: We demonstrated that the expression levels of pre-miR-1269b and miR-1269b in HBV-positive HepG2.2.15 cells were dramatically increased compared with HBV-negative HepG2 cells. HBx was shown to facilitate translocation of NF-κB from the cytoplasm to the nucleus, and NF-κB binds to the promoter of miR-1269b to enhance its transcription. miR-1269b targets and up-regulates CDC40, a cell division cycle 40 homolog. CDC40 increases cell cycle progression, cell proliferation and migration. Rescue experiment indicated that CDC40 promotes malignancy induced by miR-1269b in HCC cells. CONCLUSION: We found that HBx activates NF-κB to promote the expression of miR1269b, which augments CDC40 expression, contributing to malignancy in HCC. Our findings provide insights into the mechanisms underlying HBV-induced hepatocarcinogenesis.
Asunto(s)
Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/patología , Proteínas de Ciclo Celular/metabolismo , Movimiento Celular/genética , MicroARNs/metabolismo , FN-kappa B/metabolismo , Factores de Empalme de ARN/metabolismo , Transactivadores/metabolismo , Regulación hacia Arriba/genética , Regiones no Traducidas 3'/genética , Secuencia de Bases , Ciclo Celular/genética , Proliferación Celular/genética , Regulación Neoplásica de la Expresión Génica , Células Hep G2 , Humanos , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/patología , MicroARNs/genética , Fenotipo , Regiones Promotoras Genéticas , Unión Proteica/genética , Transcripción Genética , Proteínas Reguladoras y Accesorias ViralesRESUMEN
Increasing evidence has lent support to the notion that miRNAs regulate hepatocellular carcinoma (HCC) cell proliferation by directly targeting cell cycle-related genes. Among these genes, we identified PRPF4B, a CDK-like kinase, as a new target of miR-371-5p. Over-expression of miR-371-5p and knockdown of PRPF4B promotes cell growth by accelerating the G1/S transition in HCC cell lines. Moreover, miR-371-5p promotes tumor growth of QGY-7703 cells in vivo. Conversely, inhibition of miR-371-5p yields an opposing effect. Ectopic expression of PFPF4B abolishes the malignant phenotypes caused by miR-371-5p. Furthermore, contrary to PRPF4B, miR-371 was up-regulated in HCC tissues. Collectively, we highlight the significance of miR-371-5p and PRPF4B in cell cycle progression and hepatocarcinogenesis.
