Fusions between Epstein-Barr viral nuclear antigen-1 of Epstein-Barr virus and the large T-antigen of simian virus 40 replicate their cognate origins.
Epstein-Barr viral nuclear antigen-1 (EBNA-1) is required for the stable replication of plasmids that contain oriP, the origin of DNA synthesis used during the latent phase of the Epstein-Barr virus life cycle. EBNA-1 acts post-synthetically through unknown mechanisms to facilitate the continued synthesis of oriP plasmids in ensuing S phases. In contrast to viral replicons such as that of SV40, DNA synthesis of oriP is restricted to a single round during each cell cycle. Large T-antigen of SV40 is a DNA helicase and activates the synthesis of SV40 DNA by recruiting cellular proteins required for DNA synthesis to the origin of SV40. Using fusion proteins of EBNA-1 and large T-antigen, we tested whether tethering large T-antigen to oriP is sufficient to initiate multiple rounds of DNA synthesis from oriP during each cell cycle. We report here that, although these fusion proteins retain the biological activities of both EBNA-1 and large T-antigen, their constituent proteins do not confer the properties of one on the other. Thus, it is not possible to subvert the cellular controls that restrict DNA synthesis from oriP to a single round per cell cycle. These results also provide insights into architectural constraints at oriP and at the SV40 ori.
Aiyar
A
A
McArdle Laboratory for Cancer Research, University of Wisconsin, Madison, Wisconsin 53706, USA.
Sugden
B
B
D000952:Antigens, Polyomavirus Transforming / Q000276:immunology*; D001426:Bacterial Proteins / Q000235:genetics; D002460:Cell Line; D004247:DNA / Q000378:metabolism; D004261:DNA Replication; D015245:Deoxyribonuclease BamHI; D019309:Epstein-Barr Virus Nuclear Antigens / Q000276:immunology*; D004854:Herpesvirus 4, Human / Q000276:immunology*; D006801:Humans; D011401:Promoter Regions, Genetic; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D013539:Simian virus 40 / Q000276:immunology*; D015533:Transcriptional Activation
D000952:Antigens, Polyomavirus Transforming; D001426:Bacterial Proteins; D019309:Epstein-Barr Virus Nuclear Antigens; D011993:Recombinant Fusion Proteins; D004247:DNA; C096463:BamHIC protein, Bacillus amyloliquefaciens; D015245:Deoxyribonuclease BamHI