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Initiation of DNA replication within oriP is dispensable for stable replication of the latent Epstein-Barr virus chromosome after infection of established cell lines.
J Virol. 2000 Sep; 74(18):8563-74.JV

Abstract

The 165-kb circularized chromosome of Epstein-Barr virus (EBV) is replicated in latently infected cells once per cell cycle by host proteins during S phase. Replication initiates at multiple sites on latent EBV chromosomes, including within a 1.8-kb region called oriP, which can provide both replication and stabilization for recombinant plasmids in the presence of the EBV-encoded protein, EBNA-1. Replication initiates at or near the dyad symmetry component (DS) of oriP, which depends on multiple EBNA-1 binding sites for activity. To test the importance of the replication function of oriP, the DS was deleted from the viral genome. EBV mutants lacking the DS and carrying a selectable gene could establish latent infections in BL30 cells, in which circular, mutant viral chromosomes were stably maintained. Analysis of replication fork movement using two-dimensional gel electrophoresis showed that the deletion of the DS reduced the initiation events to an undetectable level within the oriP region so that this segment was replicated exclusively by forks entering the region from either direction. A significant slowing or stalling of replication forks that occurs normally at the approximate position of the DS was also eliminated by deletion of the DS. The results confirm the DS as both a replication origin and a place where replication forks pause. Since the replication function of oriP is dispensable at least in certain cell lines, the essential role of EBNA-1 for infection of these cell lines is likely to be that of stabilizing the EBV chromosome by associating with the 30-bp repeats of oriP. The results also imply that in established cell lines, the EBV chromosome can be efficiently replicated entirely from origins that are activated by cellular factors. Presumably, initiation of replication at the DS, mediated by EBNA-1, is important for the natural life cycle of EBV, perhaps in establishing latent infections of normal B cells.

Authors+Show Affiliations

Department of Cell Biology, Albert Einstein College of Medicine, Bronx, New York 10461, USA.No affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, U.S. Gov't, P.H.S.

Language

eng

PubMed ID

10954558

Citation

Norio, P, et al. "Initiation of DNA Replication Within oriP Is Dispensable for Stable Replication of the Latent Epstein-Barr Virus Chromosome After Infection of Established Cell Lines." Journal of Virology, vol. 74, no. 18, 2000, pp. 8563-74.
Norio P, Schildkraut CL, Yates JL. Initiation of DNA replication within oriP is dispensable for stable replication of the latent Epstein-Barr virus chromosome after infection of established cell lines. J Virol. 2000;74(18):8563-74.
Norio, P., Schildkraut, C. L., & Yates, J. L. (2000). Initiation of DNA replication within oriP is dispensable for stable replication of the latent Epstein-Barr virus chromosome after infection of established cell lines. Journal of Virology, 74(18), 8563-74.
Norio P, Schildkraut CL, Yates JL. Initiation of DNA Replication Within oriP Is Dispensable for Stable Replication of the Latent Epstein-Barr Virus Chromosome After Infection of Established Cell Lines. J Virol. 2000;74(18):8563-74. PubMed PMID: 10954558.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Initiation of DNA replication within oriP is dispensable for stable replication of the latent Epstein-Barr virus chromosome after infection of established cell lines. AU - Norio,P, AU - Schildkraut,C L, AU - Yates,J L, PY - 2000/8/23/pubmed PY - 2000/9/30/medline PY - 2000/8/23/entrez SP - 8563 EP - 74 JF - Journal of virology JO - J Virol VL - 74 IS - 18 N2 - The 165-kb circularized chromosome of Epstein-Barr virus (EBV) is replicated in latently infected cells once per cell cycle by host proteins during S phase. Replication initiates at multiple sites on latent EBV chromosomes, including within a 1.8-kb region called oriP, which can provide both replication and stabilization for recombinant plasmids in the presence of the EBV-encoded protein, EBNA-1. Replication initiates at or near the dyad symmetry component (DS) of oriP, which depends on multiple EBNA-1 binding sites for activity. To test the importance of the replication function of oriP, the DS was deleted from the viral genome. EBV mutants lacking the DS and carrying a selectable gene could establish latent infections in BL30 cells, in which circular, mutant viral chromosomes were stably maintained. Analysis of replication fork movement using two-dimensional gel electrophoresis showed that the deletion of the DS reduced the initiation events to an undetectable level within the oriP region so that this segment was replicated exclusively by forks entering the region from either direction. A significant slowing or stalling of replication forks that occurs normally at the approximate position of the DS was also eliminated by deletion of the DS. The results confirm the DS as both a replication origin and a place where replication forks pause. Since the replication function of oriP is dispensable at least in certain cell lines, the essential role of EBNA-1 for infection of these cell lines is likely to be that of stabilizing the EBV chromosome by associating with the 30-bp repeats of oriP. The results also imply that in established cell lines, the EBV chromosome can be efficiently replicated entirely from origins that are activated by cellular factors. Presumably, initiation of replication at the DS, mediated by EBNA-1, is important for the natural life cycle of EBV, perhaps in establishing latent infections of normal B cells. SN - 0022-538X UR - https://www.unboundmedicine.com/medline/citation/10954558/Initiation_of_DNA_replication_within_oriP_is_dispensable_for_stable_replication_of_the_latent_Epstein_Barr_virus_chromosome_after_infection_of_established_cell_lines_ L2 - http://jvi.asm.org/cgi/pmidlookup?view=long&pmid=10954558 DB - PRIME DP - Unbound Medicine ER -