A Cluster of Virus-Encoded MicroRNAs Accelerates Acute Systemic Epstein-Barr Virus Infection but Does Not Significantly Enhance Virus-Induced Oncogenesis In Vivo

Abstract

Over 90% of the adult human population is chronically infected with the Epstein-Barr virus (EBV), an oncogenic herpesvirus. EBV primarily infects naive human B cells and persists latently in memory B cells. Most individuals experience an asymptomaticinfection that is effectively controlled by the adaptive immune response. However, EBV-associated lymphomas can develop in immunocompromised individuals. These tumors typically express all nine EBV latent proteins (latency III). Latency III is also associated with the expression of three precursor microRNAs (miRNAs) located within the EBV BHRF1 gene locus. The role of these BHRF1 miRNAs was unclear until recent in vitro studies demonstrated that they cooperate to enhance virus-induced B cell transformation and decrease the antigenic load of virus-infected cells, indicating that the BHRF1 miRNA cluster may serve as a novel therapeutic target for the treatment of latency III EBV-associated malignancies. However, to date, it is not known if BHRF1 miRNAs enhance virus-induced oncogenesis and/or immune evasion of EBV in vivo. To understand the in vivo contribution of the BHRF1 miRNA cluster to EBV infection and EBV-associated tumorigenesis, we monitored EBV infection and assessed tumor formation in humanized mice exposed to wild-type virus and a viral mutant (⌬123) that lacks all three BHRF1 miRNAs. Our results demonstrate that while the BHRF1 miRNAs facilitate the development of acute systemic EBV infection, they do not enhance the overall oncogenic potential of EBV in vivo.E pstein-Barr Virus (EBV), a human gamma herpesvirus, is widespread in all populations; more than 90% of adults worldwide have antibodies to EBV, and infection with EBV persists for the life of its host (1). In healthy individuals, EBV is effectively controlled by the immune system and typically remains asymptomatic. Upon primary infection, virus-targeted B cells undergo a period of rapid proliferation until either CD8 ϩ T cells mount an efficient antiviral response (2) or infected B cells differentiate into a pool of latently infected memory-like B cells (3). When the immune system is suppressed, EBV can induce the development of certain lymphomas (4-6).EBV-associated malignancies that arise in immunodeficient individuals (i.e., posttransplant and AIDS patients) typically express all nine EBV latent proteins (latency III): six Epstein-Barr virus nuclear antigens (EBNA) and three latent membrane proteins (LMP) (1, 3). Although most EBV-associated malignancies respond poorly to chemotherapy, immunotherapy approaches that boost or supplement the patient's EBV-specific T cell response have been successful in treating some latency III tumors due to their high antigenic load (4). However, these strategies, which typically involve the adoptive transfer of autologous or allogeneic EBV-specific T cells stimulated in vitro, are often laborious and technically challenging (4). Therefore, new therapeutic targets and approaches are urgently needed for the treatment of EBV-associated malignancies.In addition to th...

Immune Evasion by Epstein-Barr Virus

“…Although our knowledge on the function of the EBV miRNAs is limited, knockout EBV strains have indicated a role for the BHRF1 miRNAs and, to a lesser extent, for the BART miRNAs in the early phase of B-cell transformation (Feederle et al 2011;Seto et al 2010;Vereide et al 2014). Experiments in humanized mice showed that the BHRF1 miRNAs were not crucial for infection, but knockout virus did show delayed kinetics compared to the wild-type virus (Wahl et al 2013). In a xenograft mouse model, expression of the BART miRNAs appeared to provide a tumor cell growth advantage (Qiu et al 2015).…”

MicroRNAs in large herpesvirus DNA genomes: recent advances

“…However, two recent in vivo studies using a humanized mouse model led to contradictory results. First, it was observed that BHRF1 miRNAs contribute to viral growth but did not play a role in EBV-mediated oncogenesis (74). Furthermore, a second study showed that although the BART miRNA cluster of the EBV M81 strain did not affect the ability of the virus to induce tumors, these viral miRNAs were able to repress B-cell tumorigenesis (75).…”

“…Indeed, while the absence of MuHV-4 miRNAs did not significantly affect the colonization and latency in immunocompetent mice, impairment of MuHV-4 miRNA expression reduced the virulence of MuHV-4 infection in immunocompromised IFN-γ-deficient mice, suggesting a role in promoting gammaherpesvirus-associated disease (96,97). In addition, research conducted on malignant catarrhal fever (MCF), a T-cell lymphoproliferative cattle disease induced by OvHV-2 or alcelaphine herpesvirus 1 (AlHV-1) revealed the expression of a very high number of miRNAs after propagation of LCLs and in vivo (73,74). However, the deletion from the AlHV-1 genome of 28 clustered-viral miRNAs did not alter the induction of MCF in the rabbit model (99).…”

Dynamic Epstein–Barr Virus Gene Expression on the Path to B-Cell Transformation

“…These LCLs were markedly deficient in S-phase progression indicating a persistent role for these miRNAs in B-cell proliferation. In very recent studies using humanized mouse models it was discovered that the presence of an intact BHRF1 miRNA locus was required for high-level systemic virus load, but both wild-type and BHRF1 mutant viruses had the same oncogenic potential (Wahl et al, 2013). …”