Upregulation of LMP1 Expression by Histone Deacetylase Inhibitors in an EBV Carrying NPC Cell Line JUN NISHIKAWA, 1,2 LORAND L. KIS, 1 ANQUAN LIU, 1 XIANGNING ZHANG, 1 MIKI TAKAHARA, 1 KENTARO BANDOBASHI, 1 CSABA KISS, 1 NOEMI NAGY, 1 KIWAMU OKITA, 2 GEORGE KLEIN 1 & EVA KLEIN 1, * 1 Microbiology and Tumor Biology Center, Karolinska Institute, S-171 77 Stockholm, Sweden 2 Department of Gastroenterology & Hepatology, Yamaguchi University, School of Medicine, Minami-Kogushi 1-1-1, Ube, Yamaguchi 755-8505, Japan Received August 27, 2003; Accepted September 14, 2003 Abstract. Objectives: In about 60% of Epstein–Barr virus (EBV) carrying nasopharyngeal carcinomas (NPC) LMP1 expressing cells can be detected. The frequency of LMP1 positive cells and the expression level varies from cell to cell in the different tumors. Cell lines derived from EBV positive NPCs loose the virus during in vitro culture. The in vitro infected NPC cell line TWO3-EBV used in our study carries the neomycin-resistance gene containing EBV and expresses low level of LMP1. With this cell line it was thus possible to study the regulation of LMP1 expression by modification of chromatin acetylation state. Study design: The TWO-EBV cell line was treated with n-butyrate (NB) or trichostatin A (TSA). Results: Shown by immunoblotting, the LMP1 level was elevated in the treated samples. Already 2 h after TSA exposure LMP1 expression was higher and it increased up to 24 h. Immunofluorescence staining showed that nearly all cells were LMP1 positive. Neither EBNA2 nor BZLF1 were induced. Tested first 2 h after the treatment, acetylated histone H3 and H4 were already detectable, and their level increased up to 8 h. Chromatin immunoprecipitation (ChIP) verified that the LMP1-promoter (LMP1p) (ED-L1) was acety- lated after TSA treatment. Conclusion: EBV carrying epithelial cells do not express EBNA-2. We showed that LMP1 expression was upregulated by histone deacetylase inhibitors in an in vitro infected, EBV carrier NPC cell line. Key words: Epstein–Barr virus (EBV), histone acetylation, histone deacetylase inhibitor, latent membrane protein 1(LMP1), nasopharyngeal carcinoma (NPC) Introduction Epstein–Barr virus (EBV) genome can be detected in several malignancies, both of lymphoid and epithelial cell origin [1]. The expression of the vi- rally encoded proteins differs in the various tumor cell types. In the in vitro transformed B cells 9 EBV encoded proteins are expressed (six nuclear; EBNAs and three membrane associated; LMPs). B cell malignancies exhibit 3 patterns of EBV la- tency, such as type I – expressing only EBNA1 protein – in Burkitt lymphomas (BL); type II – expressing EBNA1 and LMP1, LMP2s – in Hodgkin lymphoma (HL), and type III – ex- pressing EBNA1-6 and LMP1 and LMP2s – in immunoblastic lymphomas which arise in immu- nocompromised patients. In EBV carrying T/NK cell lymphomas and in nasopharyngeal carcino- mas (NPC), the cell virus interaction is type II [2]. *Author for all correspondence: E-mail: Eva.Klein@mtc.ki.se Virus Genes 28:1, 121–128, 2004 Ó 2004 Kluwer Academic Publishers. Manufactured in The Netherlands.