Mutational Mapping of pUL131A of Human Cytomegalovirus Emphasizes Its Central Role for Endothelial Cell Tropism

Schuessler, Andrea; Sampaio, Kerstin Laib; Straschewski, Sarah; Sinzger, Christian

doi:10.1128/jvi.05354-11

Cited by 19 publications

(15 citation statements)

References 41 publications

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“…This was a surprising result and stands in stark contrast to the reports that similar CCTA mutagenesis of the UL128-131 proteins greatly disrupted gH/gL/UL128-131 (54)(55)(56). Given that gH/gL/UL128-131 likely binds cell type-specific receptors to facilitate infection (21,22), these observations suggest that the receptor-binding surfaces lie on UL128-131 themselves and gH/gL may serve as a scaffold or platform for presentation of the UL128-131 proteins.…”

Section: Discussioncontrasting

confidence: 88%

Scanning Mutagenesis of Human Cytomegalovirus Glycoprotein gH/gL

Schultz

Lanchy

Ellerbeck

et al. 2016

J Virol

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The core, conserved function of the herpesvirus gH/gL is to promote gB-mediated membrane fusion during entry, although the mechanism is poorly understood. The human cytomegalovirus (HCMV) gH/gL can exist as either the gH/gL/gO trimer or the gH/gL/UL128/UL130/UL131 (gH/gL/UL128-131) pentamer. One model suggests that gH/gL/gO provides the core fusion role during entry into all cells within the broad tropism of HCMV, whereas gH/gL/UL128-131 acts at an earlier stage, by a distinct receptor-binding mechanism to enhance infection of select cell types, such as epithelial cells, endothelial cells, and monocytes/ macrophages. To further study the distinct functions of these complexes, mutants with individual charged cluster-to-alanine (CCTA) mutations of gH and gL were combined to generate a library of 80 mutant gH/gL heterodimers. The majority of the mutant gH/gL complexes were unable to facilitate gB-mediated membrane fusion in transient-expression cell-cell fusion experiments. In contrast, these mutants supported the formation of gH/gL/UL128-131 complexes that could block HCMV infection in receptor interference experiments. These results suggest that receptor interactions with gH/gL/UL128-131 involve surfaces contained on the UL128-131 proteins but not on gH/gL. gH/gL/UL128-131 receptor interference could be blocked with anti-gH antibodies, suggesting that interference is a cell surface phenomenon and that anti-gH antibodies can block gH/gL/UL128-131 in a manner that is distinct from that for gH/gL/gO. IMPORTANCEInterest in the gH/gL complexes of HCMV (especially gH/gL/UL128-131) as vaccine targets has far outpaced our understanding of the mechanism by which they facilitate entry and contribute to broad cellular tropism. For Epstein-Barr virus (EBV), gH/gL and gH/gL/gp42 are both capable of promoting gB fusion for entry into epithelial cells and B cells, respectively. In contrast, HCMV gH/gL/gO appears to be the sole fusion cofactor that promotes gB fusion activity, whereas gH/gL/UL128-131 expands cell tropism through a distinct yet unknown mechanism. This study suggests that the surfaces of HCMV gH/gL are critical for promoting gB fusion but are dispensable for gH/gL/UL128-131 receptor interaction. This underscores the importance of gH/gL/gO in HCMV entry into all cell types and reaffirms the complex as a candidate target for vaccine development. The two functionally distinct forms of gH/gL present in HCMV make for a useful model with which to study the fundamental mechanisms by which herpesvirus gH/gL regulates gB fusion. H uman cytomegalovirus (HCMV), an exemplar of the betaherpesvirus subfamily, is endemic in human populations and causes lifelong persistent infections (1-3). Primary infection of healthy individuals is usually subclinical and asymptomatic; however, in immunocompromised hosts, such as those infected with HIV or transplant recipients on antirejection therapies, primary infection or reactivation can have serious complications. Furthermore, maternal transmission of HCMV to the developing fetus ...

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Section: Discussioncontrasting

confidence: 88%

Scanning Mutagenesis of Human Cytomegalovirus Glycoprotein gH/gL

Schultz

Lanchy

Ellerbeck

et al. 2016

J Virol

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“…This method, which has been used in studies of other HCMV proteins and in other systems (Schuessler et al, 2010; Schuessler et al, 2008; Schuessler et al, 2012), capitalizes on the propensity for charged clusters of amino acids to be surface-exposed. Therefore, charged clusters are likely to be mediators of protein-protein interactions and to tolerate mutations without drastically impacting the overall protein structure.…”

Section: Resultsmentioning

confidence: 99%

Essential role of protein kinase R antagonism by TRS1 in human cytomegalovirus replication

2016

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Human cytomegalovirus (HCMV) lacking TRS1 and IRS1 (HCMV[ΔI/ΔT]) cannot replicate in cell culture. Although both proteins can block the protein kinase R (PKR) pathway, they have multiple other activities and binding partners. It remains unknown which functions are essential for HCMV replication. To investigate this issue, we first identified a TRS1 mutant that is unable to bind to PKR. Like HCMV[ΔI/ΔT], a recombinant HCMV containing this mutant (HCMV[TRS1-Mut 1]) did not replicate in wild-type cells. However, HCMV[ΔI/ΔT] did replicate in cells in which PKR expression was reduced by RNA interference. Moreover, HCMV[ΔI/ΔT] and HCMV[TRS1-Mut 1] replicated to similar levels as virus containing wild-type TRS1 in cell lines in which PKR expression was knocked out by CRISPR/Cas9-mediated genome editing. These results demonstrate that the sole essential function of TRS1 is to antagonize PKR and that its other activities do not substantially enhance HCMV replication, at least in cultured human fibroblasts.

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“…One essential virion component expressed in clinical CMV strains, is the pentameric complex of gH/gL/pUL128-pUL131A. Wild type CMV strains including Merlin, TB40E, and TR have been shown to express substantial amounts and differential ratios of gH/gL/pUL128-pUL131A pentameric complexes in the virion envelope 28,38,71 . This pentameric protein complex is necessary for entry into epithelial cells, endothelial cells, monocytes, and dendritic cells [27][28][29]31,72,73 .…”

Section: Discussionmentioning

confidence: 99%

Differential Expression of PDGF Receptor-α in Human Placental Trophoblasts Leads to Different Entry Pathways by Human Cytomegalovirus Strains

Naing

Hamilton

Zuylen

et al. 2020

Sci Rep

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Human cytomegalovirus (CMV) is the leading non-genetic cause of fetal malformation in developed countries. CMV placental infection is a prerequisite for materno-fetal transmission of virus, and fetal infection. We investigated the roles of the viral pentameric complex gH/gL/pUL128-pUL131A, and cellular platelet-derived growth factor receptor-α (PDGFRα) for CMV infection in first trimester extravillous-derived (SGHPL-4) and villous-derived (HTR-8/SVneo) trophoblast cells. Infection with four CMV clinical and laboratory strains (Merlin, TB40E, Towne, AD169), and Merlin deletion mutants of UL128-, UL130-, and UL131A-genes, showed a cell type-dependent requirement of the viral pentameric complex for infection of trophoblast cells. The viral pentameric complex was essential for infection of villous trophoblasts, but non-essential for extravillous trophoblasts. Blocking of PDGFRα in extravillous trophoblasts, which naturally express PDGFRα, inhibited entry of pentameric complex-deficient CMV strains, but not the entry of pentameric positive CMV strains. Transient expression of PDGFRα in villous trophoblasts, which are naturally deficient in PDGFRα, promoted the entry of CMV strains lacking gH/ gL/pUL128-pUL131A, but had no effect on entry of pentameric positive CMV strains. These results suggest PDGFRα is an important cell receptor for entry of CMV mutant strains lacking gH/gL/pUL128-pUL131A complexes in some placental cells, suggesting these entry pathways could be potential antiviral targets.

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Mutational Mapping of pUL131A of Human Cytomegalovirus Emphasizes Its Central Role for Endothelial Cell Tropism

Cited by 19 publications

References 41 publications

Scanning Mutagenesis of Human Cytomegalovirus Glycoprotein gH/gL

Scanning Mutagenesis of Human Cytomegalovirus Glycoprotein gH/gL

Essential role of protein kinase R antagonism by TRS1 in human cytomegalovirus replication

Differential Expression of PDGF Receptor-α in Human Placental Trophoblasts Leads to Different Entry Pathways by Human Cytomegalovirus Strains

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