Mechanisms Of Epstein-Barr Virus-encoded BARF1 And Its Variant To Affect The Biological Behaviors Of Nasopharyngeal Carcinoma Cell Lines

Background and objective: The Epstein-Barr virus(EBV)-encoded BARF1 gene is considered as one of the virus oncogene and its transformation role has been demonstrated in various cell lines. BARF1 is frequently expressed in nasopharyngeal carcinoma(NPC), indicating that BARF1 gene may play an important role in NPC. Our previous study showed that BARF1 and its variant, V29 A, increase the anti-apoptosis and migration abilities of NPC cell line CNE1. This study aims to further explore the oncogenic functions of BARF1 and its variant in EBV-negative NPC cell lines HONE1 and investigate the mechanisms of BARF1 to affect the biological behaviors of NPC cell lines.Methods: The cell biological behavior analysis were conducted including MTT assay, colony formation assay, anti-apoptosis assay, cell migration and invasion assay and in vivo tumorigenicity test in prototype BARF1 and V29 A variant BARF1 transfected cells under the control of lentivirus vector transfection group. The differentially expressed genes between the BARF1 transfected cells and the vector transfected cells were detected by cDNA microarray technique. Functional ontologies and pathway analysis for the BARF1-modulated genes was applied, and the BARF1-related cancer pathways and key genes were validated in NPC cell lines and xenograft tumors.Results:(1) Expression of prototype BARF1 and variant BARF1 in HONE1 cell lines increased cell proliferation and colony formation ability(P<0.05), and enhanced the cell anti-apoptosis, migration and invasion ability(P<0.05). No difference was found between prototype and variant BARF1-expressing cells.(2) BARF1 expression in HONE1 cells did not enhance the in vivo tumorigenicity in nude mice.(3) The prototype BARF1 and variant BARF1 transfected cells demonstrated similar expression profile and they showed 317 differentially expressed genes compared with vector transfected cells, including 94 up-regulated genes and 223 down-regulated genes. The BARF1-modulated genes were involved in cell adhesion, cell signal transduction, cell apoptosis, cell proliferation, cell migration and invasion, and participanted in many pathways, such as cytokine-cytokine receptor interaction, ErbB signaling pathway and pathways in cancer.(4) Real-time PCR showed that CDH1, TP63, MIR205 HG, GJA1 and S100A2 were down-regulated in BARF1 transfected cells and xenograft tumors, while ZEB2 was up-regulated, which were consistent with the results of microarray.Conclusion:(1) Both the prototype BARF1 and variant BARF1 enhance the oncogenic ability in NPC cell lines HONE1. The V29 A variant of BARF1 has no significant effect on carcinogenic ability of BARF1 gene.(2) BARF1 modulates the expression of many cancer-related genes.(3) BARF1 gene may promote cell proliferation in NPC cell lines by down-regulating the expression of CDH1, GJA1 and S100A2, or increase the ability of anti-apoptosis, cell migration and invasion by down-regulating the expression of TP63 and MIR205 and up-regulating the expression of ZEB2.