HHV-8 ORIGIN-DEPENDENT DNA REPLICATION

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Kaposi's sarcoma herpesvirus (KSHV) or human herpesvirus type 8 (HHV-8) as it will be referred to in this proposal, is the likely cause of Kaposi's sarcoma (KS). Much of the data collected to date involves epidemiological evidence demonstrating that HHV-8 DNA sequences are present in a variety of lymphoid-derived cancers. For example, HHV-8 is implicated in multicentric Castleman's disease and body cavity-based large cell lymphomas. Despite intense research, little is known about HHV-8 lytic replication. Inspection of the HHV-8 nucleotide sequence does identify the candidate replication gene homologs to helicase- primase, polymerase and its processivity factor, and a single- stranded DNA binding protein. However none of these homologs have been shown by functional assay to be required for lytic replication. Now that the entire HHV-8 genome has been sequenced and putative ORFs have been identified, the next step is to identify the HHV-8 lytic origin of replication (oriLyt) and to elucidate those viral factors required for lytic DNA replication. To this end, a putative lytic origin of replication has been identified using a transient assay in BC3 cells. This proposal will: i) Characterize the putative lytic origin of replication for HHV-8, ii) identify the HHV-8 genes required for origin-dependent DNA replication, and iii) determine which genes participate directly in DNA replication and which play an ancillary role and characterize the HHV-8 initiator protein.