The Genes Encoding the gCIII Complex of Human Cytomegalovirus Exist in Highly Diverse Combinations in Clinical Isolates

Rasmussen, Lucy; Geissler, Aimee; Cowan, Catherine M.; Chase, Amanda J.; Winters, Mark A.

doi:10.1128/jvi.76.21.10841-10848.2002

Cited by 92 publications

(125 citation statements)

References 53 publications

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“…Also among this group are genes UL146 (27,35,36,(66)(67)(68) and UL74 (gO) (27,35,(69)(70)(71), which are well characterized because of their utility in genotyping clinical isolates. The selection pressures responsible for generating such degrees of divergence are not fully understood, but their origins, and perhaps the era in which they have operated, appears to be ancient (66).…”

Section: Discussionmentioning

confidence: 99%

Reconstruction of the complete human cytomegalovirus genome in a BAC reveals RL13 to be a potent inhibitor of replication

Stanton¹,

Baluchova²,

Dargan³

et al. 2010

J. Clin. Invest.

244

375

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Human cytomegalovirus (HCMV) in clinical material cannot replicate efficiently in vitro until it has adapted by mutation. Consequently, wild-type HCMV differ fundamentally from the passaged strains used for research. To generate a genetically intact source of HCMV, we cloned strain Merlin into a self-excising BAC. The Merlin BAC clone had mutations in the RL13 gene and UL128 locus that were acquired during limited replication in vitro prior to cloning. The complete wild-type HCMV gene complement was reconstructed by reference to the original clinical sample. Characterization of viruses generated from repaired BACs revealed that RL13 efficiently repressed HCMV replication in multiple cell types; moreover, RL13 mutants rapidly and reproducibly emerged in transfectants. Virus also acquired mutations in genes UL128, UL130, or UL131A, which inhibited virus growth specifically in fibroblast cells in wild-type form. We further report that RL13 encodes a highly glycosylated virion envelope protein and thus has the potential to modulate tropism. To overcome rapid emergence of mutations in genetically intact HCMV, we developed a system in which RL13 and UL131A were conditionally repressed during virus propagation. This technological advance now permits studies to be undertaken with a clonal, characterized HCMV strain containing the complete wild-type gene complement and promises to enhance the clinical relevance of fundamental research on HCMV.

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Section: Discussionmentioning

confidence: 99%

Reconstruction of the complete human cytomegalovirus genome in a BAC reveals RL13 to be a potent inhibitor of replication

Stanton¹,

Baluchova²,

Dargan³

et al. 2010

J. Clin. Invest.

244

375

View full text Add to dashboard Cite

show abstract

“…DNA sequences of the UL74 genes of clinical HCMV isolates and HCMV BAC clones have been reported previously along with GenBank accession numbers (35,45,48). Alignment and phylogenetic analyses were performed using the default parameters of the online MAFFT (version 7) tool offered by the Computational Biology Research Center (CBRC), National Institute of Advanced Industrial Science and Technology (AIST), Japan.…”

Section: Methodsmentioning

confidence: 99%

“…Rasmussen et al performed a phylogenetic analysis of the predicted gO sequences of more than 40 clinical isolates of HCMV and found eight groups, or families (35). Within each group, the gO amino acid sequences were 98 to 100% identical, whereas differences between groups ranged from about 10 to 30%.…”

mentioning

confidence: 99%

Comparative Analysis of gO Isoforms Reveals that Strains of Human Cytomegalovirus Differ in the Ratio of gH/gL/gO and gH/gL/UL128-131 in the Virion Envelope

Zhou

Wechsler

et al. 2013

J Virol

142

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Herpesvirus glycoprotein complex gH/gL provides a core entry function through interactions with the fusion protein gB and can also influence tropism through receptor interactions. The Epstein-Barr virus gH/gL and gH/gL/gp42 serve both functions for entry into epithelial and B cells, respectively. Human cytomegalovirus (HCMV) gH/gL can be bound by the UL128-131 proteins or gO. The phenotypes of gO and UL128-131 mutants suggest that gO-gH/gL interactions are necessary for the core entry function on all cell types, whereas the binding of UL128-131 to gH/gL likely relates to a distinct receptor-binding function for entry into some specific cell types (e.g., epithelial) but not others (e.g., fibroblasts and neurons). There are at least eight isoforms of gO that differ by 10 to 30% of amino acids, and previous analysis of two HCMV strains suggested that some isoforms of gO function like chaperones, disassociating during assembly to leave unbound gH/gL in the virion envelope, while others remain bound to gH/gL. For the current report, we analyzed the gH/gL complexes present in the virion envelope of several HCMV strains, each of which encodes a distinct gO isoform. Results indicate that all strains of HCMV contain stable gH/gL/gO trimers and gH/gL/UL128-131 pentamers and little, if any, unbound gH/gL. TR, TB40/e, AD169, and PH virions contained vastly more gH/gL/gO than gH/gL/UL128-131, whereas Merlin virions contained mostly gH/gL/UL128-131, despite abundant unbound gO remaining in the infected cells. Suppression of UL128-131 expression during Merlin replication dramatically shifted the ratio toward gH/gL/gO. These data suggest that Merlin gO is less efficient than other gO isoforms at competing with UL128-131 for binding to gH/gL. Thus, gO diversity may influence the pathogenesis of HCMV through effects on the assembly of the core versus tropism gH/gL complexes.

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“…It seemed possible that the adaptation of AD169 to longterm passage in fibroblasts might also involve alterations in gO. HCMV gO is unusually variable (15 to 25% amino acid differences) among different HCMV strains compared with other viral genes (13,34,35,37,46). In recent studies, Jiang et al (26) described a gO-null mutant derived from the HCMV strain TB40/E, a strain that can infect endothelial cells following extensive passage on these cells.…”

mentioning

confidence: 99%

A Human Cytomegalovirus gO-Null Mutant Fails To Incorporate gH/gL into the Virion Envelope and Is Unable To Enter Fibroblasts and Epithelial and Endothelial Cells

Wille¹,

Knoche

Nelson

et al. 2010

J Virol

142

189

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Human cytomegalovirus (HCMV) depends upon a five-protein complex, gH/gL/UL128-131, to enter epithelial and endothelial cells. A separate HCMV gH/gL-containing complex, gH/gL/gO, has been described. Our prevailing model is that gH/gL/UL128-131 is required for entry into biologically important epithelial and endothelial cells and that gH/gL/gO is required for infection of fibroblasts. Genes encoding UL128-131 are rapidly mutated during laboratory propagation of HCMV on fibroblasts, apparently related to selective pressure for the fibroblast entry pathway. Arguing against this model in the accompanying paper by B. J. Ryckman et al. (J. Virol., 84:2597-2609, 2010), we describe evidence that clinical HCMV strain TR expresses a gO molecule that acts to promote endoplasmic reticulum (ER) export of gH/gL and that gO is not stably incorporated into the virus envelope. This was different from results involving fibroblast-adapted HCMV strain AD169, which incorporates gO into the virion envelope. Here, we constructed a TR gO-null mutant, TR⌬gO, that replicated to low titers, spread poorly among fibroblasts, but produced normal quantities of extracellular virus particles. TR⌬gO particles released from fibroblasts failed to infect fibroblasts and epithelial and endothelial cells, but the chemical fusogen polyethylene glycol (PEG) could partially overcome defects in infection. Therefore, TR⌬gO is defective for entry into all three cell types. Defects in entry were explained by observations showing that TR⌬gO incorporated about 5% of the quantities of gH/gL in extracellular virus particles compared with that in wild-type virions. Although TR⌬gO particles could not enter cells, cell-to-cell spread involving epithelial and endothelial cells was increased relative to TR, apparently resulting from increased quantities of gH/gL/UL128-131 in virions. Together, our data suggest that TR gO acts as a chaperone to promote ER export and the incorporation of gH/gL complexes into the HCMV envelope. Moreover, these data suggest that it is gH/gL, and not gH/gL/gO, that is present in virions and is required for infection of fibroblasts and epithelial and endothelial cells. Our observations that both gH/gL and gH/gL/UL128-131 are required for entry into epithelial/endothelial cells differ from models for other beta-and gammaherpesviruses that use one of two different gH/gL complexes to enter different cells.

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The Genes Encoding the gCIII Complex of Human Cytomegalovirus Exist in Highly Diverse Combinations in Clinical Isolates

Cited by 92 publications

References 53 publications

Reconstruction of the complete human cytomegalovirus genome in a BAC reveals RL13 to be a potent inhibitor of replication

Reconstruction of the complete human cytomegalovirus genome in a BAC reveals RL13 to be a potent inhibitor of replication

Comparative Analysis of gO Isoforms Reveals that Strains of Human Cytomegalovirus Differ in the Ratio of gH/gL/gO and gH/gL/UL128-131 in the Virion Envelope

A Human Cytomegalovirus gO-Null Mutant Fails To Incorporate gH/gL into the Virion Envelope and Is Unable To Enter Fibroblasts and Epithelial and Endothelial Cells

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