Natural Variations in BRLF1 Promoter Contribute to the Elevated Reactivation Level of Epstein-Barr Virus in Endemic Areas of Nasopharyngeal Carcinoma.

Epstein-Barr virus (EBV) infection is a crucial risk factor for nasopharyngeal carcinoma (NPC), but the mechanism for its elevated activation level in NPC endemic areas remains unclear. This study aims to identify the EBV natural variations contributed to the different reactivation potential between NPC endemic and non-endemic areas. 1030 subjects were recruited in China, including 303 healthy individuals from two NPC non-endemic areas, 483 healthy people from three endemic areas and 244 NPC patients. Among which, saliva DNA samples from 244 participants were sequenced for the EBV immediate early (IE) genes of BRLF1 and BZLF1, their promoters were included; the rest 786 subjects were used for the validation of significant variations among three different populations. Haplotype and population structure analysis were conducted. Dual-luciferase assay was used to detect the promoter activity. A total of 246 distinct variations were detected, 29 showed significant difference in the frequencies between healthy people from NPC endemic area and non-endemic area. Population structure analysis clustered EBV strains into 9 subgroups mostly in accordance with the geographical origin of samples. Interestingly, two EBV genotypes, Rp-V1 and Rp-V2, were identified according to the linkage relationship of the variations in BRLF1 promoter (Rp). Rp-V1 has higher frequency in NPC endemic areas than in non-endemic areas (52.38% vs 18.15%, P = 2.07 × 10-14), and was associated with higher oral EBV DNA levels (adjusted OR = 1.64, 95% CI = 1.21-2.24, P = .002), suggesting a more powerful activation ability of Rp-V1 than that of the prototype Rp-of the EBV strain; On the contrary, Rp-V2 has higher frequency in NPC non-endemic areas than in endemic areas (18.48% vs 0.38%, P = 1.17 × 10-7), might represent a reduced activation potential of EBV. Further dual-luciferase assay showed Rp-V1 has higher promoter activity while compared with Rp-V2 (P < .0001). Notably, Rp-V1 impaired the transcription repression effect of YY1 while Rp-V2 strengthened the transcription repression effect of EBF1 on Rp. In addition, significant differences of Rta 393-407 CTL epitope which may influence the recognition of Rta by CD8+ T cells were detected between healthy people from NPC endemic area and non-endemic area. This study identified natural variations in cis-acting elements (YY1 and EBF1) of EBV Rp altering Rp transcription activities, which may contribute to the elevated EBV activation level in NPC endemic areas than non-endemic areas.

Zhang JB ，Huang SY ，Wang TM ，Dong SQ ，He YQ ，Zheng XH ，Li XZ ，Wang F ，Jianbing M ，Jia WH ... -  《EBioMedicine》

Expression of Epstein-Barr virus lytic gene BRLF1 in nasopharyngeal carcinoma: potential use in diagnosis.

Feng P ，Ren EC ，Liu D ，Chan SH ，Hu H ... -  《JOURNAL OF GENERAL VIROLOGY》

Epstein-Barr Virus BRLF1 Induces PD-L1 Expression in Nasopharyngeal Carcinoma Cells.

Wu CC ，Chen MS ，Lee TY ，Huang TS ，Cho DY ，Chen JY ... -  《-》

Inhibition of Epstein-Barr virus reactivation in nasopharyngeal carcinoma cells by dietary sulforaphane.

Epstein-Barr virus (EBV) has been associated with several human malignancies including nasopharyngeal carcinoma (NPC). Reactivation of latent EBV has been considered to contribute to the carcinogenesis of NPC. Blocking the EBV lytic cycle has been shown effective in the treatment of EBV-associated diseases. We have searched for natural dietary compounds inhibiting EBV reactivation in NPC cells. Among them, sulforaphane (SFN) was found to be effective in the inhibition of EBV reactivation in latent EBV-positive NPC cells, NA and HA. SFN is a histone deacetylase (HDAC) inhibitor and has been recognized as an antioxidant and antitumor compound for chemoprevention. However, its antiviral effect is less well elucidated. In this study, after determination of the cytotoxicity of SFN on various epithelial cells, we showed that SFN treatment inhibits EBV reactivation, rather than induction, by detection of EBV lytic gene expression in EBV-positive NPC cells. We also determined that the number of cells supporting the EBV lytic cycle is decreased using immunofluorescence and flow cytometric analysis. Moreover, we have found that this inhibitory effect decreases virus production. To elucidate the inhibitory mechanism of SFN on the EBV lytic cycle, luciferase reporter assays were carried out on the Zta and Rta promoters. The results show that SFN inhibits transactivation activity of the EBV immediate-early gene Rta but not Zta. Together, our results suggest that SFN has the capability to inhibit EBV lytic cycle and the potential to be taken as a dietary compound for prevention of EBV reactivation.

Wu CC ，Chuang HY ，Lin CY ，Chen YJ ，Tsai WH ，Fang CY ，Huang SY ，Chuang FY ，Lin SF ，Chang Y ，Chen JY ... -  《-》

A single nucleotide polymorphism in the Epstein-Barr virus genome is strongly associated with a high risk of nasopharyngeal carcinoma.

Feng FT ，Cui Q ，Liu WS ，Guo YM ，Feng QS ，Chen LZ ，Xu M ，Luo B ，Li DJ ，Hu LF ，Middeldorp JM ，Ramayanti O ，Tao Q ，Cao SM ，Jia WH ，Bei JX ，Zeng YX ... -  《-》