title: A sequence within the varicella-zoster virus (VZV) OriS is a negative regulator of DNA replication and is bound by a protein complex containing the VZV ORF29 protein.
abstract: The architecture of the varicella-zoster virus (VZV) origin of DNA replication (OriS) differs significantly from that of the herpes simplex virus (HSV) DNA replication origin. Novel aspects of the VZV OriS include a GA-rich region, three binding sites for the VZV origin-binding protein (OBP) all on the same strand and oriented in the same direction, and a partial OBP binding site of unknown function. We have designated this partial binding site Box D and have investigated the role it plays in DNA replication and flanking gene expression. This has been done with a model system using a replication-competent plasmid containing OriS and a replication- and transcription-competent dual-luciferase reporter plasmid containing both the OriS and the intergenic region between VZV open reading frames (ORFs) 62 and 63. We have found that (i) Box D is a negative regulator of DNA replication independent of flanking gene expression, (ii) the mutation of Box D results in a decrease in flanking gene expression, thus a sequence within the VZV OriS affects transcription, which is in contrast to results reported for HSV-1, (iii) there is a specific Box D complex formed with infected cell extracts in electrophoretic mobility shift assay experiments, (iv) supershift assays show that this complex contains the VZV ORF29 single-strand DNA-binding protein, and (v) the formation of this complex is dependent on the presence of CGC motifs in Box D and its downstream flanking region. These findings show that the VZV ORF29 protein, while required for DNA replication, also plays a novel role in the suppression of that process.
authors.affiliation: Department of Microbiology and Immunology and the Witebsky Center for Microbial Pathogenesis and Immunology, University at Buffalo, Buffalo, New York 14214, USA. mikhalil@buffalo.edu
authors.forename: Mohamed I
authors.initials: MI
authors.lastname: Khalil
authors.forename: Ann
authors.initials: A
authors.lastname: Arvin
authors.forename: Jeremy
authors.initials: J
authors.lastname: Jones
authors.forename: William T
authors.initials: WT
authors.lastname: Ruyechan
mesh_terms: D002467:Cell Nucleus / Q000378:metabolism / Q000821:virology; D004261:DNA Replication; D004279:DNA, Viral / Q000235:genetics; D024202:Electrophoretic Mobility Shift Assay; D017930:Genes, Reporter; D014645:Herpesvirus 3, Human / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D017874:Immediate-Early Proteins / Q000235:genetics; D008545:Melanoma / Q000235:genetics* / Q000473:pathology / Q000821:virology; D016366:Open Reading Frames / Q000235:genetics*; D010957:Plasmids; D018741:Replication Origin / Q000235:genetics*; D015534:Trans-Activators / Q000235:genetics; D014158:Transcription, Genetic; D014407:Tumor Cells, Cultured; D014759:Viral Envelope Proteins / Q000235:genetics; D014764:Viral Proteins / Q000235:genetics* / Q000378:metabolism*; D017735:Virus Latency; D014779:Virus Replication
chemical_list: D004279:DNA, Viral; C074221:IE62 protein, Human herpesvirus 3; D017874:Immediate-Early Proteins; D015534:Trans-Activators; D014759:Viral Envelope Proteins; D014764:Viral Proteins; C074551:immediate early protein 63, Human herpesvirus 3