Epstein–Barr virus reprograms human B lymphocytes immediately in the prelatent phase of infection

Significance More than 50 years ago, Epstein–Barr virus (EBV) was found to infect resting human B lymphocytes and to induce their continuous proliferation in vitro. This seminal finding clearly mimics aspects of EBV’s oncogenic potential in vivo but how EBV activates primary human B lymphocytes and how lymphoblastoid cell lines (LCLs) evolve from the EBV-infected lymphocytes is uncertain. We studied this model and found that the EBV-infected naïve B lymphocytes undergo a germinal center-like activation and differentiation program within only a few days after infection to emerge with transcriptomic and phenotypic features resembling plasmablasts and early plasma cells. In vitro in established LCLs, EBV “locks” the infected B cells in a stage of differentiation that is only transient in vivo. Epstein–Barr virus (EBV) is a human tumor virus and a model of herpesviral latency. The virus efficiently infects resting human B lymphocytes and induces their continuous proliferation in vitro, which mimics certain aspects of EBV’s oncogenic potential in vivo. How lymphoblastoid cell lines (LCLs) evolve from the infected lymphocytes is uncertain. We conducted a systematic time-resolved longitudinal study of cellular functions and transcriptional profiles of newly infected naïve primary B lymphocytes. EBV reprograms the cells comprehensively and globally. Rapid and extensive transcriptional changes occur within 24 h and precede any metabolic and phenotypic changes. Within 72 h, the virus activates the cells, changes their phenotypes with respect to cell size, RNA, and protein content, and induces metabolic pathways to cope with the increased demand for energy, supporting an efficient cell cycle entry on day 3 postinfection. The transcriptional program that EBV initiates consists of 3 waves of clearly discernable clusters of cellular genes that peak on day 2, 3, or 4 and regulate RNA synthesis, metabolic pathways, and cell division, respectively. Upon onset of cell doublings on day 4, the cellular transcriptome appears to be completely reprogrammed to support the proliferating cells, but 3 additional clusters of EBV-regulated genes fine-tune cell signaling, migration, and immune response pathways, eventually. Our study reveals that more than 11,000 genes are regulated upon EBV infection as naïve B cells exit quiescence to enter a germinal center-like differentiation program, which culminates in immortalized, proliferating cells that partially resemble plasmablasts and early plasma cells.