Comparison of Transcriptional Activation of Epstein-Barr Virus and Cellular Gene Expression by ZEBRA and Mutant Cellular AP-1 Protein

The ubiquitous gamma herpes virus, Epstein-Barr virus (EBV), has latent and lytic phases in its life cycle. The EBV ZEBRA protein activates the EBV lytic cycle from latency. Previous work has shown that the homologous cellular AP-1 proteins, c-Jun and c-Fos, with alanine-to-serine substitutions homologous to ZEBRA(S186) can assume some functions of EBV ZEBRA. These AP-1(A!S) mutants are capable of binding to methylated EBV DNA and activating expression of some EBV genes. Here, I compare the ability of ZEBRA and AP-1(A/S) to activate expression of both viral genes and cellular genes of the human host.;Previous work characterizing the ability of AP-1(A/S) to activate expression of EBV genes has been limited. In order to characterize the overall pattern of expression of viral genes by ZEBRA and AP-1(A/S), I performed an EBV gene array. I present findings that mutant c-Jun and c-Fos proteins selectively activate expression of EBV lytic genes, including a subgroup of viral late genes, in the absence of viral DNA replication. These findings indicate that newly synthesized viral DNA is not invariably required for viral late gene expression. While viral DNA replication may be obligatory for late gene expression that is driven by viral transcription factors, DNA replication does not limit the ability of cellular transcription factors to activate expression of some viral late genes. The c-Fos A 151 S mutation has been identified in a human cancer. This data provides proof of principle that mutant cellular transcription factors could cause aberrant regulation of viral lytic cycle gene expression and play important roles in EBV associated diseases. The ability of EBV to regulate cellular genes during the viral lytic cycle has been limited to two previous whole genome studies in lymphoid cells and studies of specific cellular genes. There have been no studies on the ability of AP-1(A/S) to activate cellular gene expression. In order to understand the ability of ZEBRA and AP-1(A/S) to activate cellular gene expression, I performed whole genome RNA-seq in epithelial cells infected with EBV expressing either ZEBRA or AP-1(A/S). I show that expression of either ZEBRA or AP-1(A/S) caused differential expression of cellular genes. A gene ontology term enrichment analysis identified several significantly enriched groups of genes; of particular interest were genes encoding secreted proteins. I identified five cellular genes for follow up that were upregulated in the RNA-seq data by ZEBRA and AP-1(A/S): CCL5, IL-8, IL-11, LOX, and NEFH. Neither of the previously published whole genome analyses analyzed protein expression of any genes identified as being upregulated at the RNA level during the viral lytic cycle. I could not detect changes in the protein expression of NEFH or IL-8 by ZEBRA even though changes in RNA expression could be detected. My findings show that while ZEBRA and AP-1(A/S) are capable of regulating cellular gene expression, the changes in RNA expression are not reflective of protein expression.