oriLyt, the lytic origin of DNA replication of Epstein-Barr virus (EBV), ensures viral DNA amplification during the productive or lytic phase of the virus' life cycle. To understand the contribution of cis- and transacting elements involved in DNA replication of oriLyt, a detailed mutational analysis was undertaken which defined BZLF1, a viral transcriptional activator, as an essential replication factor. The BZLF1 protein belongs to the extended fos/jun family of transcription factors and binds to specific BZLF1 binding motifs within oriLyt, as well as to consensus AP-1 sites. Recombinant, chimeric transcription factors identified the transcriptional activation domain of BZLF1 as being necessary to mediate DNA replication, a function which could not be substituted by any other transcription factor tested, including jun, E2, myc or VP16.