Epstein-Barr virus infection promotes epithelial cell growth by attenuating differentiation-dependent exit from cell cycle

Epstein-Barr virus (EBV) is a human herpesvirus that is associated with lymphomas as well as nasopharyngeal and gastric carcinomas. Although carcinomas account for almost 90% of EBV associated cancers, progress in examining EBV's role in their pathogenesis has been limited by difficulty in establishing latent infection in non-transformed epithelial cells. Recently, EBV infection of hTERT-immortalized normal oral keratinocytes (NOKs) has emerged as a model that recapitulates aspects of EBV infection in vivo, such as differentiation-associated viral replication. Using uninfected NOKs and NOKs infected with the Akata strain of EBV (NOKs-Akata), we examined changes in gene expression due to EBV infection and differentiation. Latent EBV infection produced very few significant gene expression changes in undifferentiated NOKs, but significantly reduced the extent of differentiation-induced gene expression changes. Gene set enrichment analysis revealed that differentiation-induced downregulation of cell cycle and metabolism pathways was markedly attenuated in NOKs-Akata relative to uninfected NOKs. We also observed that pathways induced by differentiation were less upregulated in NOKs-Akata. Consistent with our RNA-seq results, we observed decreased differentiation markers and increased suprabasal MCM7 expression in NOKs-Akata versus NOKs when grown in raft cultures. These effects were also observed in NOKs infected with a replication defective EBV mutant (Akata?RZ), implicating mechanisms other than lytic gene induced host-shutoff. Our results help define the mechanisms by which EBV infection alters keratinocyte differentiation and provide a basis for understanding the role of EBV in epithelial cancers.