Ubiquitin and ubiquitin like (Ubl) proteins such as SUMO-1 and SUMO-2/3 have wide ranging effects on protein stability, localization, interactions, and overall function. Many viruses have evolved mechanisms to influence cellular processes through manipulation of the ubiquitin proteasome system. Kaposi?s sarcoma herpesvirus (KSHV) encodes proteins with E3 ubiquitin ligase activity, SUMO ligase activity and interacts with the cellular ubiquitin proteasome system, with reported effects on signaling, apoptosis, immune response and cell cycle regulation. KSHV is a human tumor virus associated with three different cancers and two inflammatory syndromes. Both latency and lytic replication play roles in establishing and maintaining disease, therefore it is important to understand the mechanisms that regulate this transition. Our long-term goal is to identify and characterize cellular pathways targeted by KSHV through viral manipulation of the ubiquitin and ubiquitin-like modification systems. This will contribute to our understanding of the cell biology of KSHV infection and enable identification of targets for further study and ultimately intervention. We have carried out a comparative analysis of the ubiquitome in TREx BCBL1 cells as well as cells expressing RTA alone. We have identified 193 differentially ubiquitinated sites in 146 proteins in cells transfected with RTA, 274 sites in 206 proteins in doxycycline treated TREx BCBL1 cells, and 209 sites in 98 proteins in both data sets, representing proteins with RTA induced ubiquitination alterations.
In AIM1 of this grant, we propose to carry out validation and initial characterization of our findings in concert with the development of a CURE (course- based undergraduate research experience).
In AIM 2, undergraduate and masters students in my laboratory will evaluate outcomes in infected cells in terms of lytic reactivation, genome replication and infectious virus production. These studies will have a significant impact as we will identify and characterize novel strategies of viral evasion and manipulation of host cellular processes while providing undergraduate research experiences to a diverse population of students.
We propose to examine how KSHV manipulates the ubiquitin and ubiquitin-like modification systems to regulate the transition from latency to lytic replication. Kaposi?s sarcoma herpesvirus (KSHV) is the causative agent of Kaposi?s sarcoma and is associated with primary effusion lymphoma and multicentric Castleman?s disease as well as two inflammatory diseases: KSHV inflammatory cytokine syndrome and immune reconstitution syndrome-KS. The results of the proposed experiments have the potential to inform the development of novel chemotherapeutics.
