The Role of Cis-acting Elements, RNA Modification, and Cellular Localization of mRNA Transcripts During KSHV Lytic Infection

Abstract

Exploitation and control of the cellular environment is the cornerstone of successful viral infection, with struggle for control over gene expression machinery being a driving force for the evolutionary arms race between viruses and their hosts. During infection, different viruses utilize a wide range of strategies to prioritize viral transcripts over host transcripts. In both Coronavirus and Kaposi’s sarcoma associated herpesvirus (KSHV) this is driven by a wide-spread RNA decay event which leads to the destruction of host RNA as driven by viral proteins. In KSHV, this process is driven by multiple factors during the duration of KSHV lytic infection with a viral endoribonuclease, SOX, being the primary trigger and a process known as hyperadenylation, triggered by SOX activity and SOX nuclear relocalization. In Coronavirus, this process is driven by the nsp1 protein, which has variable activity between different Coronaviruses, but specifically targets and degrades host transcripts, primarily through interactions with the ribosome. However, in both conditions, there exists subsets of transcripts that are resistant to these viral decay events, of both host and viral origin. These resistant transcripts are protected from decay through a myriad of different mechanisms related to RNA secondary structure, RNA modifications, and cellular localization. In this dissertation, we will explore the role of these different factors in two separate viral systems to investigate the impact of viral activity on RNA stability, to better our understanding of viral-host interplay in the RNA landscape. In the first chapter, we will explore the impact of four different Coronavirus nsp1 proteins and how the different classifications of viruses utilize their homologs in a variety of ways to impact the host transcriptional landscape. In the second chapter, we will explore work dedicated to understanding our primary escapee, human Interleukin-6 (IL-6), and the impact of RNA secondary structure, nuclear export, and virally induced RNA modification on this transcript. Finally, in the third chapter, we extend exploration into the polyA landscape of transcripts during KSHV lytic reactivation to better understand the impact of hyperadenylation on the host transcriptome. To achieve this we performed PolyA-seq to explore the changes in polyA tail length as well as subcellular localization of transcripts as well as experiments exploring the impact of CRM1 inhibition on the stability and localization of these transcripts.Doctor of Philosophy (Ph.D.