Interaction of HCMV UL84 with C/EBPα transcription factor binding sites within oriLyt is essential for lytic DNA replication

AbstractHuman cytomegalovirus (HCMV) lytic DNA replication is initiated at the cis-acting oriLyt region and requires six core replication proteins along with UL84 and IE2. Although UL84 is thought to be the replication initiator protein, little is known about its interaction with oriLyt. We have now performed chromatin immunoprecipitation assays (ChIP) using antibodies specific to UL84, IE2, UL44, CCAAT/enhancer binding protein (C/EBPα) and PCR primers that span the entire oriLyt region to reveal an evaluation of specific protein binding across oriLyt. UL84 interacted with several regions of oriLyt that contain C/EBPα transcription factor binding sites. Mutation of either of one of C/EBPα (92,526 or 92,535) sites inactivated oriLyt and resulted in the loss of binding of UL84. These data reveal the regions of interaction within oriLyt for several key replication proteins and show that the interaction between UL84 and C/EBPα sites within oriLyt is essential for lytic DNA replication