We sequenced the rhesus lymphocryptovirus (LCV) genome in order to determine its genetic similarity to Epstein-Barr virus (EBV). The rhesus LCV encodes a repertoire identical to that of EBV, with 80 open reading frames, including cellular interleukin-10, bcl-2, and colony-stimulating factor 1 receptor homologues and an equivalent set of viral glycoproteins. The highly conserved rhesus LCV gene repertoire provides a unique animal model for the study of EBV pathogenesis.Epstein-Barr virus (EBV)-related herpesviruses in the same gamma-1, or lymphocryptovirus (LCV), genera are known to naturally infect both Old and New World nonhuman primates, and the biology of these nonhuman LCVs appears indistinguishable from that of EBV (reviewed in reference 35). The potential utility of using Old World LCV as an animal model system was demonstrated by the ability to experimentally infect naive rhesus macaques with rhesus LCVs and reproduce many aspects of acute and persistent EBV infection in humans (20).Previous studies revealed that Old World LCV genomes are organized in a colinear fashion with EBV and that EBV DNA cross-reacts with viral DNA from simian LCVs (11,12). Rhesus LCV homologues for most of the EBV latent infection genes have been described (reviewed in reference 35). In virtually every aspect, these rhesus LCV latent infection genes are functionally interchangeable with the EBV genes despite modest degrees of homology (27 to 50% amino acid homology). However, the gene repertoire from the rhesus LCV, or any gamma-1 herpesvirus besides EBV, has not been completely characterized, particularly those genes encoding cellular homologues and viral glycoproteins that are highly relevant for studies in an animal model system. The development of a rhesus LCV genetic system to generate mutant viruses for use in experimental infections and study of molecular pathogenesis in vivo also requires a thorough understanding of the rhesus LCV genome and its sequence as a starting point.Primary sequence and genome structure of rhesus LCV. Six overlapping cosmid and two plasmid viral DNA clones were isolated from the rhesus LCV-infected B-cell line LCL8664 (Fig. 1A). A shotgun cloning and sequencing strategy was used to derive contiguous sequences from these eight viral DNA clones. The complete rhesus LCV sequence was assembled with a sevenfold average redundancy from 1,500 overlapping sequences of 300 to 800 nucleotides.The rhesus LCV genome contains internal (IR1 to IR4) and terminal repeats (TR) as in EBV (Fig. 1B). The major internal repeat, IR1, contains 5.7 copies of a 3,072-bp motif that is 61.5% homologous to the 3,072-bp BamHI W fragment of the EBV IR1. The rhesus LCV TR consists of a 933-bp motif versus a 538-bp motif in the EBV TR, and there is no significant sequence homology besides a similarly high GC content (75%). Based on 5.7 copies in the major internal repeat and 4 TR copies, the rhesus LCV genome has 171,096 nucleotides (versus 172,231 bp in B95-8 EBV with 11.3 IR1 copies and 4 TR copies), with an overall GC content...

No abstract

The Epstein-Barr virus (EBV) nuclear antigen 3B (EBNA-3B) is considered nonessential for EBV-mediated B-cell growth transformation in vitro based on three virus isolates with EBNA-3B mutations. Two of these isolates could potentially express truncated EBNA-3B products, and, similarly, we now show that the third isolate, IB4, has a point mutation and in-frame deletion of 263 amino acids. In order to test whether a virus with EBNA-3B completely deleted can immortalize B-cell growth, we first cloned the EBV genome as a bacterial artificial chromosome (BAC) and showed that the BAC-derived virus was B-cell immortalization competent. Deletion of the entire EBNA-3B open reading frame from the EBV BAC had no adverse impact on growth of EBV-immortalized B cells, providing formal proof that EBNA-3B is not essential for EBV-mediated B-cell growth transformation in vitro.