Latent Membrane Protein 1 Is a Novel Determinant of Epstein-Barr Virus Genome Persistence and Reactivation

Epstein-Barr virus (EBV) is a ubiquitous gammaherpesvirus that persistently infects humans, with nearly 95% seropositivity in adults. Infection in differentiating epithelia is permissive, but EBV-associated nasopharyngeal carcinoma (NPC) tumors harbor a clonal and nonproductive latent infection. However, in explanted NPC cultures and epithelial cell lines, episomal EBV genomes are frequently lost. The resulting unstable infection has hampered efforts to study the determinants of EBV persistence and latency in epithelial oncogenesis. The EBV nuclear antigen 1 (EBNA1) protein is required for tethering EBV episomes to cellular DNA and for mitotic segregation to daughter cells. Expression of EBNA1 does not ensure faithful partitioning of EBV episomes or replicons, suggesting that additional regulatory mechanisms have yet to be elucidated. The EBV latent membrane protein 1 (LMP1) is an oncogenic signaling protein expressed in latent and lytic cycles. This study identified that LMP1 contributes to the loss of EBV genomes in latently infected cells and promotes differentiation-induced lytic replication in a polarized air-liquid interface (ALI) culture model. Deletion of LMP1 in recombinantly infected 293 cells promoted the retention of EBV genomes in passaged cells, which was in part localized to a conserved PXQXT motif in the C-terminal signaling domain (CTAR1). Additionally, knockdown of LMP1 in the recombinantly infected NPC cell line HK1 resulted in decreased induction of lytic proteins and infectious EBV titers. These findings are consistent with the hypothesis that in epithelial infections, regulation of LMP1 mechanisms may be a determinant of infection outcome and a potential risk factor for EBV persistence in preneoplastic cells. IMPORTANCE Latent membrane protein 1 (LMP1) is a constitutively active oncogenic signaling protein encoded by Epstein-Barr virus (EBV). Despite monoclonal infection in cases of nasopharyngeal carcinoma (NPC), it has been difficult to reconcile the heterogeneous LMP1 protein levels detected in tumor cells. The LMP1 protein is a pleiotropic signaling protein with oncogenic potential. Findings from this study are consistent with the hypothesis that LMP1 has a role distinct from that of oncogenesis that facilitates the viral life cycle by promoting an unstable but productive infection in differentiating epithelia.