All RhCMV variants infected rhesus and human fibroblasts with comparable efficiency, showing that this rhesus UL128-UL131 locus is specifically required for infection of human epithelial cells. == Fig. Human cytomegalovirus (HCMV), a -herpesvirus, is usually carried as a latent contamination by the majority of the world’s populace (1). Main contamination and reactivation from latency are generally asymptomatic, but HCMV can cause morbidity and mortality in hosts with compromised or poorly developed immune systems, such as transplant recipients, AIDS patients, neonates, and the developing fetus. The mechanisms of HCMV pathogenesis are incompletely defined, in part because the computer virus is usually species-specific and cannot be utilized for experimental contamination of animal models (1). As an alternative, rodent CMV models have been used with considerable success (2), but these viruses diverge substantially from HCMV in terms of their gene content (3). Rhesus cytomegalovirus (RhCMV) has emerged as a model that is more closely related to the human computer virus (4,5). The pathogenesis of RhCMV contamination in rhesus macaques is similar to HCMV contamination of humans (6,7), the gene content of RhCMV is usually closely related to HCMV (8,9), and the phenotypes of trans-Zeatin RhCMV mutants during replication in cultured fibroblasts are similar to the corresponding HCMV mutants (10). HCMV infects a wide range of cell types in the host, including fibroblasts, endothelial cells, and epithelial cells (11). Epithelial cells play an important role in HCMV pathogenesis. The computer virus typically enters a new host via mucosal epithelial cells, it replicates in epithelial cells in several organs during the main and prolonged stages of contamination, and it is secreted from glandular epithelial cells into bodily fluids (12). The products of the UL128, UL130, and UL131 ORFs are major determinants of HCMV replication trans-Zeatin in epithelial and other cell types. Attenuated HCMV laboratory strains contain mutations in the UL128-UL131 locus and fail to replicate efficiently in many cell types (1320). In HCMV virions, pUL128-pUL131 form a glycoprotein complex with gH and gL (13,19,21), which mediates fusion of the virion envelope with the plasma membrane of endothelial and epithelial cells (17,18,22). Antibody Rabbit Polyclonal to GANP to pUL128, pUL130 (19), or pUL131 (13) can neutralize HCMV contamination of epithelial cells but not fibroblasts. In the absence of a functional UL128-UL131 complex, HCMV trans-Zeatin enters endothelial cells, and presumably epithelial cells, by endocytosis, albeit at low efficiency (22). RhCMV encodes a locus homologous to HCMV UL128-UL131 (9,23). However, RhCMV-loxP(r) (24), RhCMV derived from an infectious BAC clone of the viral genome, and its parent, strain 68-1, are missing UL128 and the rh157.5 ORF (8,9), subsequently identified as the second exon of UL130 (23). We reported that BAC-derived 68-1 infects epithelial cells inefficiently, and hypothesized that this defect results from a defective UL128-UL131 locus (10). We repaired the locus in the RhCMV BAC clone, and the variants replicate much more efficiently than the parental computer virus in rhesus epithelial cells. This demonstrates that RhCMV contains a UL128131 locus that controls computer virus host cell range, as has been explained for the human computer virus, and it raises the possibility that the repaired computer virus will exhibit altered pathogenesis in rhesus macaques when compared with the well analyzed 68-1 strain. In addition, the repaired variants grow efficiently in human epithelial and endothelial cells, causing us to speculate that RhCMV-like viruses might infect humans. == Results == == UL128-UL131 Locus Promotes Efficient RhCMV Replication in Rhesus Epithelial Cells. == Computer virus recovered from your trans-Zeatin BAC clone of RhCMV strain 68-1, RhCMV-loxP(r) (24), infects rhesus epithelial cells with low efficiency and spreads slowly, but produces large amounts of extracellular computer virus over extended periods of time (10). Sequence analyses of the RhCMV strain 180.92 and other RhCMV isolates have shown that BAC-derived 68-1 and its parent, 68-1, are missing UL128 and the second exon of UL130 (8,9,23). However, strain trans-Zeatin 180.92 has a large deletion in the ULbregion of the genome, and it has not been cloned as a BAC (9). To analyze the importance of the RhCMV UL128-UL131 locus for replication in epithelial cells, we repaired the locus in the 68-1 BAC clone using the genome of strain 180.92 as template. The producing BAC, pBRh68-1.1, has a wild-type UL128-UL131.
