Mapping EBNA-1 Domains Involved in Binding to Metaphase Chromosomes

Abstract

The Epstein-Barr virus (EBV) genome can persist in dividing human B cells as multicopy circular episomes. Viral episomes replicate in synchrony with host cell DNA and are maintained at a relatively constant copy number for a long time. Only two viral elements, the replication origin OriP and the EBNA-1 protein, are required for the persistence of viral genomes during latency. EBNA-1 activates OriP during the S phase and may also contribute to the partition and/or retention of viral genomes during mitosis. Indeed, EBNA-1 has been shown to interact with mitotic chromatin. Moreover, viral genomes are noncovalently associated with metaphase chromosomes. This suggests that EBNA-1 may facilitate the anchorage of viral genomes on cellular chromosomes, thus ensuring proper partition and retention. In the present paper, we have investigated the chromosome-binding activity of EBV EBNA-1, herpesvirus papio (HVP) EBNA-1, and various derivatives of EBV EBNA-1, fused to a variant of the green fluorescent protein. The results show that binding to metaphase chromosomes is a common property of EBV and HVP EBNA-1. Further studies indicated that at least three independent domains (CBS-1, -2, and -3) mediate EBNA-1 binding to metaphase chromosomes. In agreement with the anchorage model, two of these domains mapped to a region that has been previously demonstrated to be required for the long-term persistence of OriP-containing plasmids.

The DNA segregation mechanism of Epstein–Barr virus nuclear antigen 1

“…A recent study by Marechal et al (1999) examined the contribution of EBNA1 sequences to mitotic chromosome attachment by fusing small fragments of EBNA1 to green fluorescent protein.…”

Nonviral in vivo gene transfer in the mucopolysaccharidosis I murine model

“…To improve nonviral transference systems, plasmids based on the Epstein-Barr virus (EBV) have been tested. These vectors contain viral elements responsible for the independent replication of the plasmid in the nucleus of the target cell, minimizing the effects of its dilution after cellular division and therefore providing a stable expression of the gene for longer times and also resistance to antibiotics (Herweijer and Wolff 2003;Marechal et al 1999). In this work, we tested the efficiency of a nonviral gene transfer system in a preclinical protocol for MPS I, using an EBV-based plasmid in a murine knockout model for the IDUA gene.…”

Epstein‐Barr virus infection and human malignancies

“…This is not surprising as EBNA1 is crucial for persistence of the viral DNA within the infected cell, and recombinant viruses devoid of EBNA1 lose their ability to immortalize B-cells . EBNA1 binds to several viral DNA domains and in particular to the 20 tandem direct 30 bp repeats present in the ori P latent origin of replication as well as to human chromatin (Rawlins et al 1985;Marechal et al 1999). EBNA1 is therefore thought to tether the viral episomes to the chromatin via ori P, ensuring their transmission to the cell progeny.…”