Effects Of Epstein-Barr Virus-encoded BARF1Gene And Its Variant On The Biological Behaviors Of Nasopharyngeal Carcinoma Cell

Background and objective:The Epstein-Barr virus (EBV)-encoded BARF1gene is considered as one of the virus oncogene and its transformation role has been demonstrated in various cell lines. BARF1is frequently expressed in nasopharyngeal carcinoma (NPC) and EBV-associated gastric carcinoma tissues, indicating that BARF1gene may play an important role in epithelial cell tumorigenesis such as NPC and EBV-associated gastric carcinoma. In our previous study, a variant of BARF1(V29A) was identified in Northern China, which showed a relatively higher frequency in NPC than healthy donors. This study aims to investigate the oncogenic functions of BARF1and its variant in EBV-negative NPC cell lines.Methods:The lentivirus vector which expressed BARF1prototype gene (pLenti6.3-B95-8) or BARF1variant gene (pLenti6.3-V29A) was constructed and transfected into EBV-negative NPC cell line CNE, respectively. After screened by Blasticidin and confirmed by RT-PCR and qRT-PCR, NPC cell line with stable expression of BARF1prototype gene or variant gene was established. The cell biological behavior analysis were conducted including MTT assay, cell cycle analysis by flow cytometric, colony formation assay, anti-apoptosis observation induced by anti-cancer drug diamminedichloroplatinum (DDP) and migration test under the control of vacant lentivirus vector transfection group (pLenti6.3).Results:(1) NPC cell line with stable expression of BARFl prototype gene or variant gene was successfully established.(2) Both the prototype BARF1-expressing cells and variant BARF1-expressing cells did not show higher proliferation than non-expressing cells by MTT assay, colony formation assay and cell cycle analysis (P>0.05), and also no difference was found between prototype and variant BARF1-expressing cells (P>0.05).(3) At48h induced by DDP, the prototype BARF1-expressing cells and variant BARF1-expressing cells showed higher cell viability and lower apoptosis rate than the non-expressing cells (P<0.05), while the difference between prototype and variant BARF1-expressing cells was not significant in cell viability and apoptosis rate (P>0.05).(4) The prototype BARF1-expressing cells and variant BARF1-expressing cells led to stronger migration capability than the non-expressing cells (P<0.05), while the prototype and variant BARF1-expressing cells had no significant difference in cell migration capability (P>0.05).Conclusion:(1)BARF1gene has no significant effect on the proliferation of EBV-negative NPC cell line CNE, but it can increase its anti-apoptosis and migration abilities, suggesting BARFl enhance the oncogenic ability in CNE cells and may play important roles in the development and progress of NPC.(2) The V29A variant of BARF1has no significant effect on carcinogenic ability of BARF1gene.