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Sequence requirements of the Epstein-Barr virus latent origin of DNA replication.
J Virol. 1994 Mar; 68(3):1913-25.JV

Abstract

The Epstein-Barr virus (EBV) latent origin of DNA replication (oriP) is composed of two elements that contain binding sites for the sole viral gene product required for latent cycle replication, EBNA-1. One of these elements, region I, functions as an EBNA-1-dependent enhancer for RNA polymerase II-transcribed genes, may play a role in plasmid segregation, and is required for origin function in B cells latently infected with EBV. The second element, region II, contains or is very near the site of initiation of DNA replication. A genetic approach was taken to determine the contribution of the EBNA-1 binding sites in oriP to origin function. Although region I is required for the transient replication of plasmids bearing region II in EBV-infected B cells, a plasmid lacking region I but containing region II, was observed to replicate transiently in both D98/Raji and HeLa cells expressing EBNA-1. Thus, binding of EBNA-1 to region I is not absolutely required for the molecular events that lead to initiation of DNA replication at region II. Site-directed mutagenesis of the four EBNA-1-binding sites in region II, individually and in various combinations, demonstrated that only two EBNA-1-binding sites are required for region II function. The results obtained with these mutants, together with the analysis of the replicative ability of plasmids containing insertions between EBNA-1-binding sites, suggested that the spatial relationship of the two sites is critical. Mutants that contain only two EBNA-1-binding sites separated by 26 to 31 bp in region II were not maintained as plasmids over many cell generations and were greatly reduced in their ability to replicate transiently in D98/Raji cells. The EBNA-1-induced bending or untwisting of the DNA in EBNA-1-binding sites 1 and 4 in region II did not, however, demonstrate this spatial constraint. It may be concluded from these results that specific protein-protein interactions between EBNA-1 and/or between EBNA-1 and a cellular protein(s) are required for origin function.

Authors+Show Affiliations

Department of Microbiology, State University of New York, Stony Brook 11794-5222.No affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

8107251

Citation

Harrison, S, et al. "Sequence Requirements of the Epstein-Barr Virus Latent Origin of DNA Replication." Journal of Virology, vol. 68, no. 3, 1994, pp. 1913-25.
Harrison S, Fisenne K, Hearing J. Sequence requirements of the Epstein-Barr virus latent origin of DNA replication. J Virol. 1994;68(3):1913-25.
Harrison, S., Fisenne, K., & Hearing, J. (1994). Sequence requirements of the Epstein-Barr virus latent origin of DNA replication. Journal of Virology, 68(3), 1913-25.
Harrison S, Fisenne K, Hearing J. Sequence Requirements of the Epstein-Barr Virus Latent Origin of DNA Replication. J Virol. 1994;68(3):1913-25. PubMed PMID: 8107251.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Sequence requirements of the Epstein-Barr virus latent origin of DNA replication. AU - Harrison,S, AU - Fisenne,K, AU - Hearing,J, PY - 1994/3/1/pubmed PY - 1994/3/1/medline PY - 1994/3/1/entrez SP - 1913 EP - 25 JF - Journal of virology JO - J Virol VL - 68 IS - 3 N2 - The Epstein-Barr virus (EBV) latent origin of DNA replication (oriP) is composed of two elements that contain binding sites for the sole viral gene product required for latent cycle replication, EBNA-1. One of these elements, region I, functions as an EBNA-1-dependent enhancer for RNA polymerase II-transcribed genes, may play a role in plasmid segregation, and is required for origin function in B cells latently infected with EBV. The second element, region II, contains or is very near the site of initiation of DNA replication. A genetic approach was taken to determine the contribution of the EBNA-1 binding sites in oriP to origin function. Although region I is required for the transient replication of plasmids bearing region II in EBV-infected B cells, a plasmid lacking region I but containing region II, was observed to replicate transiently in both D98/Raji and HeLa cells expressing EBNA-1. Thus, binding of EBNA-1 to region I is not absolutely required for the molecular events that lead to initiation of DNA replication at region II. Site-directed mutagenesis of the four EBNA-1-binding sites in region II, individually and in various combinations, demonstrated that only two EBNA-1-binding sites are required for region II function. The results obtained with these mutants, together with the analysis of the replicative ability of plasmids containing insertions between EBNA-1-binding sites, suggested that the spatial relationship of the two sites is critical. Mutants that contain only two EBNA-1-binding sites separated by 26 to 31 bp in region II were not maintained as plasmids over many cell generations and were greatly reduced in their ability to replicate transiently in D98/Raji cells. The EBNA-1-induced bending or untwisting of the DNA in EBNA-1-binding sites 1 and 4 in region II did not, however, demonstrate this spatial constraint. It may be concluded from these results that specific protein-protein interactions between EBNA-1 and/or between EBNA-1 and a cellular protein(s) are required for origin function. SN - 0022-538X UR - https://www.unboundmedicine.com/medline/citation/8107251/Sequence_requirements_of_the_Epstein_Barr_virus_latent_origin_of_DNA_replication_ DB - PRIME DP - Unbound Medicine ER -