B-CELL PROLIFERATION AND INDUCTION OF EARLY G(1)-REGULATING PROTEINS BY EPSTEIN-BARR-VIRUS MUTANTS CONDITIONAL FOR EBNA2

Infection of primary B-lymphocytes by Epstein-Barr virus (EBV) leads t o growth transformation of these B-cells in vitro. EBV nuclear antigen 2 (EBNA2), one of the first genes expressed after EBV infection of B- cells, is a transcriptional activator of viral and cellular genes and is essential for the transforming potential of the virus. We generated conditional EBV mutants by expressing EBNA2 as chimeric fusion protei n ninth the hormone binding domain of the estrogen receptor on the gen etic background of the virus. Growth transformation of primary normal B-cells by mutant virus resulted in estrogen-dependent lymphoblastoid cell lines expressing the chimeric EBNA2 protein. In the absence of es trogen about half of the cells enter a quiescent non-proliferative sta te whereas the others die by apoptosis. EBNA2 is thus required not onl y for initiation but also for maintenance of transformation. Growth ar rest occurred at G(1) and G(2) stages of the cell cycle, indicating th at functional EBNA2 is required at different restriction points of the cell cycle. Growth arrest is reversible for G(1)/G(0) cells as indica ted by the sequential accumulation and modification of cell cycle regu lating proteins. EBV induces the same cell cycle regulating proteins a s polyclonal stimuli in primary B-cells. These data suggest that EBV i s using a common pathway for B-cell activation bypassing the requireme nt for antigen, T-cell signals and growth factors.