CCAAT/enhancer binding proteins play a role in oriLyt-dependent genome replication during MHV-68 de novo infection

Murine gammaherpesvirus 68 (MHV-68), a member of the gammaherpesvirus family, replicates robustly in permissive cell lines and is able to infect laboratory mice. MHV-68 has emerged as a model for studying the basic aspects of viral replication and host-virus interactions of its human counterparts. Herpesvirus genome replication is mediated through a cis-element in the viral genome called the origin of lytic replication (oriLyt). A family of transcription factors, CCAAT/enhancer binding proteins (C/EBPs), assists in oriLyt-mediated DNA replication during gammaherpesvirus reactivation. In this study, we examined the role of C/EBPs in gammaherpesvirus DNA replication during de novo infection, using MHV-68 as a model. We found that C/EBP α and β bind to the CCAAT boxes in the MHV-68 oriLyt core region both in vitro and in vivo, as demonstrated by electrophoretic mobility shift assay and chromatin immunoprecipitation assay. A dominant negative form of C/EBPs significantly impaired the lytic replication efficiency of MHV-68 on both the plasmid and genome levels in a replication assay, indicating that functional C/EBPs are required for maximal MHV-68 genome DNA replication. Collectively, our data demonstrate that C/EBPs interact with the oriLyt core region and play an important role in MHV-68 lytic DNA replication during de novo infection.