Cytostatic Effect of Epstein–Barr Virus Latent Membrane Protein-1 Analyzed Using Tetracycline-Regulated Expression in B Cell Lines

AbstractTetracycline-regulated vectors were used to obtain inducible expression in stable transfected B cell lines of two Epstein–Barr virus (EBV) latent genes, LMP1 and EBNA2. The transfected genes were tightly repressed by low, nontoxic concentrations of tetracycline (≤1 μg/ml) and, following removal of tetracycline, were induced to levels comparable to or up to 3× that of EBV-transformed normal lymphoblastoid cell lines. In transfected DG75 cells, induced expression of LMP1, but not of EBNA2, led to the expected upregulation of various cell surface markers, including: CD40, CD54, CD58, and HLA class I. A novel observation was that both LMP1 and EBNA2 independently caused the downregulation of surface IgM, an effect mirrored in EBV-positive Burkitt lymphoma lines undergoing phenotypic drift during the transition from latency I to latency III in which both LMP1 and EBNA2 are upregulated. Most remarkably, induced LMP1 expression almost completely inhibited cell growth for 4 to 5 days, after which the cells recovered a limited proliferative capacity. The cytostatic effect of LMP1 was observed in all three B cell lines studied: DG75, BJAB, and Akata. Further analysis showed that induction of LMP1 coincided with a reduction in the levels of c-myc, and that the cytostatic effect was due to an accumulation of cells at the G2/M phase of the cell cycle. These data suggest a novel function for the LMP1 oncogene in controlling the proliferation of EBV-infected cells by regulating progress through G2/M phase