Studies on the ribosomal protein L22: Linking RNA-binding to subcellular localization and relocalization to Epstein-Barr virus pathogenesis

Epstein-Barr virus (EBV) is a ubiquitous human herpesvirus that establishes a lifelong latent infection in B-lymphocytes and is associated with a variety of human malignancies, including Burkitt lymphoma (BL). EBV is maintained latently in infected cells and only a subset of viral genes are expressed during latency. Included are the Epstein-Barr virus encoded RNAs, EBER-1 and EBER-2. Expression of these non-coding nuclear RNAs has been shown to mediate a number of phenotypic effects, including the ability to enhance the tumorigenic potential of EBV-negative BL cells. The mechanisms for this and other EBER-associated effects, however, have not been elucidated. Although the EBERs have been shown to interact with a number of cellular proteins in vitro, no in vivo interactions with endogenous cellular proteins have been confirmed. EBER-1 interacts with the cellular ribosomal protein L22. Previous reports have also demonstrated that L22 is relocalized during EBV infection, although the mechanism for relocalization is not known. In addition, L22 has been shown to be relocalized during Herpes Simplex virus infection and to interact with the genome of Hepatitis C virus, suggesting that L22 plays a critical role during viral replication. The studies undertaken in this dissertation were designed to characterize the interaction between EBER-1 and L22 on a molecular and functional level, as well as to evaluate the role of L22 during viral infection. RNA-binding assays and fluorescence localization studies were used to study the binding of RNA ligands by L22 and to link RNA-binding to the subcellular localization of this protein. Next, fluorescence localization studies were used to establish that EBER-1 is sufficient for L22 relocalization, and mutant EBER-1 RNAs incapable of L22 binding were utilized to evaluate the consequence of the EBER-1-L22 interaction during viral infection. Finally, a number of preliminary experiments were conducted to evaluate the role of L22 during normal cellular processes as well as during other viral infections. The data presented here demonstrate that the interaction between EBER-1 and L22 is a bona fide in vivo interaction and that the relocalization of L22 during EBV infection is a key contributor to the tumorigenic potential of EBV-infected BL cells.