The Function of EBV LMP1 CTAR3 in Sumoylation and Oncogenesis

Bentz, Gretchen

Abstract

Candidate and Environment: My career goal is to become a leading independent scientist in the field of herpesvirology. During the mentored phase of the Pathway to Independence Award, I will work to further establish my publication record in the field of EBV, laying the groundwork for the research I will do as a faculty member. This project will provide training in new techniques [BAC technology, a humanized mouse model system, investigating protein-protein and protein-DNA interactions] that will be applicable to the research performed as an independent investigator. The expertise of Dr. Pagano as well as other faculty members at UNC will be invaluable in learning the necessary skills to become a successful independent researcher. Research: EBV, a ubiquitous ?-herpesvirus, is associated with several lymphoid and epithelial malignancies. Latent membrane protein-1 (LMP1), the principal viral oncoprotein, is a constitutively active membrane signaling protein that regulates multiple signal transduction pathways via its cytoplasmic C-terminal activating regions (CTAR). In contrast to CTAR1 and CTAR2, the function for CTAR3 is not well defined. I recently published that LMP1 CTAR3 is necessary and sufficient for LMP1 to interact with the SUMO-conjugating enzyme Ubc9 and induce the sumoylation of cellular proteins. Disruption of the LMP1-Ubc9 interaction resulted in changes in cellular behaviors associated with oncogenesis and LMP1 and loss of LMP1-induced sumoylation. Further understanding how LMP1 CTAR3 functions and mechanisms by which it functions is my next priority. I hypothesize that CTAR3 is a self-regulatory domain that helps modulate the oncogenic potential of LMP1.
In Specific Aim 1, to be completed during the mentored phase, I will determine biological functions of LMP1 CTAR3 in vivo. Using wildtype and LMP1 CTAR3 deletion viruses to infect humanized mice, a new animal model developed at LCCC for EBV infection and disease, I will examine the tumorigenic and immunogenic potentials of the viruses. The role of CTAR3 is the intracellular trafficking and stability of LMP1 will also e tested.
In Specific Aim 2, to be completed during the independent phase, I will examine mechanisms by which CTAR3 functions. Initial targets of LMP1-induced sumoylation include IRF7, STAT1, and the SUMO protease SENP1. I will investigate the role of LMP1 CTAR3 in the sumoylation of these proteins and how sumoylation regulates protein function. Finally, because protein sumoylation can help inhibit the latent-lytic switch, I will examine in LMP1 CTAR3 contributes to LMP1-mediated repression of lytic replication by inducing protein sumoylation, specifically of PML, ZEB1, and ZEB2. The proposed work will greatly increase our understanding of functions for LMP1 CTAR3, specifically its role in regulating protein sumoylation, and how CTAR3 functions, expanding knowledge of how LMP1 acts as an oncoprotein.

Public Health Relevance

Epstein-Barr virus is associated with several malignancies. The principal viral oncoprotein, latent membrane protein-1 (LMP1), contains several C-terminal regulatory regions (CTARs), but not much is known about the function of CTAR3. Therefore, understanding how LMP1 CTAR3 contributes to LMP1-mediate cellular changes will offer insight into how LMP1 acts as an oncoprotein. The written critiques and criteria scores of individual reviewers are provided in essentially unedited form in the Critique section below. Please note that these critiques and criteria scores were prepared prior to the meeting and may not have been revised subsequent to any discussions at the review meeting. The Resume and Summary of Discussion section above summarizes the final opinions of the committee.