We study two human tumor viruses, EBV and KSHV, to understand how they maintain the tumor they cause and to develop effective therapies for those tumors.
Epstein-Barr Virus (EBV) contributes causally to multiple lymphomas and carcinomas throughout the world. We have developed a general genetic approach to identify viral genes and their functions needed to maintain recently isolated lymphoma cells in culture (Vereide and Sugden, Blood, 2010). In this approach lymphoma cells are engineered to express conditionally a dominant negative derivative of the viral protein, EBNA1, which can inhibit viral genome replication forcing the loss of EBV from the lymphoma cells. This loss induces apoptosis in Type 1 Burkitt’s Lymphoma (BL) cells, Wp-restricted BL cells, and Post-Transplant Lymphoproliferative Disease (PTLD) cells (ibid) which can be prevented in the first case by expression of EBV’s BART miRNAs and in the latter two cases by expression of the cellular anti-apoptotic gene, BclXL (ibid; Vereide et al. Oncogene, 2013). Surprisingly, all three classes of rescued lymphoma cells show dramatic defects in proliferation which provide robust assays to identify and characterize the viral genes that drive these lymphoma cells to proliferate. We have found that a subset of EBV’s miRNAs contribute to this proliferation and are searching for the mRNAs they target.
We also study the synthesis and partitioning of the plasmid genomes of EBV and Kaposi’s Sarcoma Herpesvirus (KSHV) to understand these events generally. (KSHV is also a human tumor virus.) To do so we have developed new methods to visualize these plasmid replicons in live cells by tagging them with a polymer of LacO sites that can be bound by the Lac repressor fused to td tomato. This approach allows the detection of newly synthesized viral DNA which increases in its florescent intensity as it binds more repressor (Nanbo et al. EMBO J. 2007; Norby et al. J.Vis Exp. 2012). We follow these DNAs through multiple cell cycles in order to elucidate their mechanisms of partitioning.