雙特異性去磷酸酶於EB病毒感染細胞中之調控與生物功能

Abstract

Epstein-Barr virus (EBV) is an oncogenic virus that infects B cells and epithelial cells, and can transform B cells to continuously proliferative lymphoblastoid cell lines (LCLs). Serial lines of evidence indicated that EBV infection is highly correlated with a variety of lymphoproliferative diseases and human carcinomas. In order to maintain long-term persistence in host cells, EBV can utilize its numerous gene products to activate many signaling pathways, including NF-κB, MAPK, PI3K/Akt, thereby influencing cell division, growth, or cell apoptosis. Previously, we have used cDNA microarray to compare the expression pattern of cellular genes in CD19+ B cells with B95.8 strain EBV-transformed LCLs, and preliminarily found that EBV affects the expression of many dual specificity phosphatases (DUSPs), such as DUSP1, DUSP6 and DUSP8. Given that some DUSPs were known to be the negative regulator in MAPK signaling, therefore, this study will further investigate the interaction between EBV and DUSPs, and also the biological functions of DUSPs in EBV-infected cells. First, we use RT-Q-PCR and RT-PCR to confirm that the expression levels of DUSP1, DUSP2, DUSP6, and DUSP8, are reduced in LCLs, compared to those in CD19+ B cells. Further, we found that EBV lytic transactivator Zta can downregulate the transcription and translation of DUSP2 in Burkitt’s lymphoma cell line, BJAB cells, and in human kidney epithelial cells, HEK293T cells. Meanwhile, we have also discovered that EBV latent membrane protein 1 (LMP1) can inhibit the transcription and translation of DUSP8 in BJAB and another Burkitt’s lymphoma cell line, Ramos cells, Hodgkin lymphoma cell line, L428 cells, and also HEK293T cells. Using luciferase reporter assay, we demonstrated that Zta can regulate the expression of DUSP2 through inhibiting its promoter activity. On the other hand, LMP1, in addition to regulate the promoter activity of DUSP8, can also activate the downstream NF-κB signaling through its C-terminal region, CTAR1 and CTAR2 domain, to inhibit the expression of DUSP8. To investigate the possible roles of DUSPs in EBV-infected B cells, we transduce DUSP2 or DUSP8 to BJAB and LCLs, and found that DUSP8 can downregulate the activation of p38 MAPK and JNK in cells. Surprisingly, we also discovered that the exogenous expression of DUSP8 in BJAB cells diminishes cell growth. Besides, we found that exogenous expression of DUSP2 dramatically changes cell morphology and inhibits cell proliferation in LCLs. However, it seems that DUSP8 does not affect cell proliferation and cell cycle transition in LCLs. This study reveals that EBV gene products, Zta and LMP1, have the ability to regulate the expression of DUSPs in EBV-infected cells, thereby promoting continuous growth of cells to maintain long-term persistence. However, further investigations will be needed to dissect the detail correlation between EBV and DUSPs, and the exact roles of DUSPs in cells.