Nucleosome assembly proteins bind to Epstein-Barr virus nuclear antigen 1 and affect its functions in DNA replication and transcriptional activation.
The EBNA1 protein of Epstein-Barr virus (EBV) plays several important roles in EBV latent infection, including activating DNA replication from the latent origin of replication (oriP) and activating the transcription of other latency genes within the EBV chromatin. These functions require EBNA1 binding to the DS and FR elements within oriP, respectively, although how these interactions activate these processes is not clear. We previously identified interactions of EBNA1 with the related nucleosome assembly proteins NAP1 and TAF-I, known to affect the replication and transcription of other chromatinized templates. We have further investigated these interactions, showing that EBNA1 binds directly to NAP1 and to the beta isoform of TAF-I (also called SET) and that these interactions greatly increase the solubility of EBNA1 in vitro. These interactions were confirmed in EBV-infected cells, and chromatin immunoprecipitation with these cells showed that NAP1 and TAF-I both localized with EBNA1 to the FR element, while only TAF-I was detected with EBNA1 at the DS element. In keeping with these observations, alteration of the NAP1 or TAF-Ibeta level by RNA interference and overexpression inhibited transcriptional activation by EBNA1 in FR reporter assays. In addition, EBNA1-mediated DNA replication was stimulated when TAF-I (but not NAP1) was downregulated and was inhibited by TAF-Ibeta overexpression. The results indicate that the interaction of EBNA1 with NAP1 and TAF-I is important for transcriptional activation and that EBNA1 recruits TAF-I to the DS element, where it negatively regulates DNA replication.
Wang
Shan
S
Department of Molecular Genetics, University of Toronto, 1 Kings College Circle, Toronto, Canada M5S 1A8.
Frappier
Lori
L
D015153:Blotting, Western; D002460:Cell Line; D004261:DNA Replication / Q000502:physiology*; D019309:Epstein-Barr Virus Nuclear Antigens / Q000502:physiology*; D004854:Herpesvirus 4, Human / Q000502:physiology*; D006801:Humans; D009687:Nuclear Proteins / Q000502:physiology; D009707:Nucleosomes / Q000502:physiology*; D011485:Protein Binding; D015533:Transcriptional Activation / Q000502:physiology*
D019309:Epstein-Barr Virus Nuclear Antigens; D009687:Nuclear Proteins; D009707:Nucleosomes; C417029:EBV-encoded nuclear antigen 1