Viral Glycoprotein Complex Formation, Essential Function and Immunogenicity in the Guinea Pig Model for Cytomegalovirus

Coleman, Stewart; Hornig, Julia; Maddux, Sarah; Choi, K. Yeon; McGregor, Alistair

doi:10.1371/journal.pone.0135567

Cited by 21 publications

(135 citation statements)

References 80 publications

(133 reference statements)

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“…The gH/gL complex has been identified as a potentially important target antigen, including in a congenital GPCMV infection model. In this study, a novel antibody therapy strategy was found to be effective in reducing the incidence of congenital CMV infection (36,58). Consequently, an effective immune response to both gB and gH/gL is likely an important factor for a successful vaccine against congenital CMV.…”

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confidence: 88%

“…Manipulation of an infectious GPCMV BAC has allowed the preliminary study of some viral genes (1,(30)(31)(32)(33)(34)(35)(36). Analysis of the viral genome (15,29) indicated that GPCMV encodes homologs to the HCMV glycoproteins (gB, gH, gL, gM, gN, and gO) in genes colinear with the HCMV genome (designated GP55, GP75, GP115, GP100, GP73, and GP74, respectively).…”

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confidence: 99%

“…In HCMV, these complexes are important neutralizing antibody targets and vaccine candidates (40)(41)(42)(43)(44). We recently demonstrated that GPCMV forms functionally similar glycoprotein complexes, and these complexes are essential for infection of fibroblast cells as well as serving as important target antigens (36). Additionally, GPCMV forms a homologous pentameric complex (gH/gL/UL128-131) that is necessary for epithelial tropism in HCMV (45) and GPCMV (103) as well as other cell types (46).…”

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confidence: 99%

“…Plaques were stained with 10% Giemsa stain or visualized by fluorescence microscopy. High-titer virus stocks were generated as previously described (36). Pathogenic wild-type GPCMV used in congenital CMV challenge studies was serially maintained as salivary gland stocks from infected guinea pigs.…”

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confidence: 99%

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A Novel Non-Replication-Competent Cytomegalovirus Capsid Mutant Vaccine Strategy Is Effective in Reducing Congenital Infection

Choi

Root

McGregor

2016

J Virol

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Congenital cytomegalovirus (CMV) infection is a leading cause of mental retardation and deafness in newborns. The guinea pig is the only small animal model for congenital CMV infection. A novel CMV vaccine was investigated as an intervention strategy against congenital guinea pig cytomegalovirus (GPCMV) infection. In this disabled infectious single-cycle (DISC) vaccine strategy, a GPCMV mutant virus was used that lacked the ability to express an essential capsid gene (the UL85 homolog GP85) except when grown on a complementing cell line. In vaccinated animals, the GP85 mutant virus (GP85 DISC) induced an antibody response to important glycoprotein complexes considered neutralizing target antigens (gB, gH/gL/gO, and gM/gN). The vaccine also generated a T cell response to the pp65 homolog (GP83), determined via a newly established guinea pig gamma interferon enzyme-linked immunosorbent spot assay. In a congenital infection protection study, GP85 DISC-vaccinated animals and a nonvaccinated control group were challenged during pregnancy with wild-type GPCMV (10 5 PFU). The pregnant animals carried the pups to term, and viral loads in target organs of pups were analyzed. Based on live pup births in the vaccinated and control groups (94.1% versus 63.6%), the vaccine was successful in reducing mortality (P ‫؍‬ 0.0002). Additionally, pups from the vaccinated group had reduced CMV transmission, with 23.5% infected target organs versus 75.9% in the control group. Overall, these preliminary studies indicate that a DISC CMV vaccine strategy has the ability to induce an immune response similar to that of natural virus infection but has the increased safety of a non-replication-competent virus, which makes this approach attractive as a CMV vaccine strategy. IMPORTANCECongenital CMV infection is a leading cause of mental retardation and deafness in newborns. An effective vaccine against CMV remains an elusive goal despite over 50 years of CMV research. The guinea pig, with a placenta structure similar to that in humans, is the only small animal model for congenital CMV infection and recapitulates disease symptoms (e.g., deafness) in newborn pups. In this report, a novel vaccine strategy against congenital guinea pig cytomegalovirus (GPCMV) infection was developed, characterized, and tested for efficacy. This disabled infectious single-cycle (DISC) vaccine strategy induced a neutralizing antibody or a T cell response to important target antigens. In a congenital infection protection study, animals were protected against CMV in comparison to the nonvaccinated group (52% reduction of transmission). This novel vaccine was more effective than previously tested gB-based vaccines and most other strategies involving live virus vaccines. Overall, the DISC vaccine is a safe and promising approach against congenital CMV infection. H uman cytomegalovirus (HCMV), a betaherpesvirus, has evolved very closely with its human host. Virus infection in a healthy host is normally asymptomatic but leads to a lifelong infection. In contrast,...

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confidence: 88%

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confidence: 99%

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confidence: 99%

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confidence: 99%

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A Novel Non-Replication-Competent Cytomegalovirus Capsid Mutant Vaccine Strategy Is Effective in Reducing Congenital Infection

Choi

Root

McGregor

2016

J Virol

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“…In human herpesvirus 6 (HHV-6) and HHV-7, the homologous protein also forms a complex with gH and gL (27,28), but to date no function has been assigned to this protein (29). Similarly to HCMV gO, murine and guinea pig cytomegalovirus (CMV) gO binds to gH/gL, and its deletion leads to a severely reduced cell-free infectivity (30,31), indicating that the function of gO is conserved among the various CMVs. An essential role of the gH/gL/gO complex for pathogenesis of murine CMV (MCMV) was recently demonstrated in a mouse model: a gO-deficient MCMV mutant was unable to establish infection in mice (32), indicating that gO plays a pivotal role for host-to-host transmission of CMV.…”

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Importance of Highly Conserved Peptide Sites of Human Cytomegalovirus gO for Formation of the gH/gL/gO Complex

Stegmann¹,

Abdellatif

Sampaio³

et al. 2017

J Virol

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The glycoprotein O (gO) is betaherpesvirus specific. Together with the viral glycoproteins H and L, gO forms a covalent trimeric complex that is part of the viral envelope. This trimer is crucial for cell-free infectivity of human cytomegalovirus (HCMV) but dispensable for cell-associated spread. We hypothesized that the amino acids that are conserved among gOs of different cytomegaloviruses are important for the formation of the trimeric complex and hence for efficient virus spread. In a mutational approach, nine peptide sites, containing all 13 highly conserved amino acids, were analyzed in the context of HCMV strain TB40-BAC4 with regard to infection efficiency and formation of the gH/gL/gO complex. Mutation of amino acids (aa) 181 to 186 or aa 193 to 198 resulted in the loss of the trimer and a complete small-plaque phenotype, whereas mutation of aa 108 or aa 249 to 254 caused an intermediate phenotype. While individual mutations of the five conserved cysteines had little impact, their relevance was revealed in a combined mutation, which abrogated both complex formation and cell-free infectivity. C343 was unique, as it was sufficient and necessary for covalent binding of gO to gH/gL. Remarkably, however, C218 together with C167 rescued infectivity in the absence of detectable covalent complex formation. We conclude that all highly conserved amino acids contribute to the function of gO to some extent but that aa 181 to 198 and cysteines 343, 218, and 167 are particularly relevant. Surprisingly, covalent binding of gO to gH/gL is required neither for its incorporation into virions nor for proper function in cell-free infection.IMPORTANCE Like all herpesviruses, the widespread human pathogen HCMV depends on glycoproteins gB, gH, and gL for entry into target cells. Additionally, gH and gL have to bind gO in a trimeric complex for efficient cell-free infection. Homologs of gO are shared by all cytomegaloviruses, with 13 amino acids being highly conserved. In a mutational approach we analyzed these amino acids to elucidate their role in the function of gO. All conserved amino acids contributed either to formation of the trimeric complex or to cell-free infection. Notably, these two phenotypes were not inevitably linked as the mutation of a charged cluster in the center of gO abrogated cell-free infection while trimeric complexes were still being formed. Cysteine 343 was essential for covalent binding of gO to gH/gL; however, noncovalent complex formation in the absence of cysteine 343 also allowed for cell-free infectivity.

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Betaherpesvirus Virion Assembly and Egress

Anderson

Pellett

2018

Advances in Experimental Medicine and Biology

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Virions are the vehicle for cell-to-cell and host-to-host transmission of viruses. Virions need to be assembled reliably and efficiently, be released from infected cells, survive in the extracellular environment during transmission, recognize and then trigger entry of appropriate target cells, and disassemble in an orderly manner during initiation of a new infection. The betaherpesvirus subfamily includes four human herpesviruses (human cytomegalovirus and human herpesviruses 6A, 6B, and 7), as well as viruses that are the basis of important animal models of infection and immunity. Similar to other herpesviruses, betaherpesvirus virions consist of four main parts (in order from the inside): the genome, capsid, tegument, and envelope. Betaherpesvirus genomes are dsDNA and range in length from ~145 to 240 kb. Virion capsids (or nucleocapsids) are geometrically well-defined vessels that contain one copy of the dsDNA viral genome. The tegument is a collection of several thousand protein and RNA molecules packed into the space between the envelope and the capsid for delivery and immediate activity upon cellular entry at the initiation of an infection. Betaherpesvirus envelopes consist of lipid bilayers studded with virus-encoded glycoproteins; they protect the virion during transmission and mediate virion entry during initiation of new infections. Here, we summarize the mechanisms of betaherpesvirus virion assembly, including how infection modifies, reprograms, hijacks, and otherwise manipulates cellular processes and pathways to produce virion components, assemble the parts into infectious virions, and then transport the nascent virions to the extracellular environment for transmission.

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Viral Glycoprotein Complex Formation, Essential Function and Immunogenicity in the Guinea Pig Model for Cytomegalovirus

Cited by 21 publications

References 80 publications

A Novel Non-Replication-Competent Cytomegalovirus Capsid Mutant Vaccine Strategy Is Effective in Reducing Congenital Infection

A Novel Non-Replication-Competent Cytomegalovirus Capsid Mutant Vaccine Strategy Is Effective in Reducing Congenital Infection

Importance of Highly Conserved Peptide Sites of Human Cytomegalovirus gO for Formation of the gH/gL/gO Complex

Betaherpesvirus Virion Assembly and Egress

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