The Tegument Protein pp65 of Human Cytomegalovirus Acts as an Optional Scaffold Protein That Optimizes Protein Uploading into Viral Particles

Reyda, Sabine; Navarro, Pedro J.; Gebauer, Wolfgang; Saur, Michael; Krauter, Steffi; Büscher, Nicole; Plachter, Bodo

doi:10.1128/jvi.01415-14

Cited by 29 publications

(45 citation statements)

References 67 publications

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“…11,67,71,72 Immunofluorescence analysis in normal HCMV infected fibroblasts showed that pp65 was not present in all viral ACs (data not shown), a fact which is in agreement with previous studies reporting that pp65 serves as an optional scaffold protein for virion formation. 73 The same was also true for pUL97 (data not shown). Moreover, in RhoB-knockdown HCMV infected cells, residual RhoB could be detected at the AC by immunofluorescence (data not shown).…”

Section: Discussionsupporting

confidence: 53%

RhoB is a component of the human cytomegalovirus assembly complex and is required for efficient viral production

et al. 2015

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Human Cytomegalovirus (HCMV), an ubiquitous b-herpesvirus, is a significant pathogen that causes medically severe diseases in immunocompromised individuals and in congenitally infected neonates. RhoB belongs to the family of Rho GTPases, which regulates diverse cellular processes. Rho proteins are implicated in the entry and egress from the host cell of mainly a-and g-herpesviruses, whereas b-herpesviruses are the least studied in this regard. Here, we studied the role of RhoB GTPase during HCMV lytic infection. Microscopy analysis, both in fixed and live infected cells showed that RhoB was translocated to the assembly complex/compartment (AC) of HCMV, a cytoplasmic zone in infected cells where many viral structural proteins are known to accumulate and assembly of new virions takes place. Furthermore, RhoB was localized at the AC even when the expression of the late HCMV AC proteins was inhibited. At the very late stages of infection, cellular projections were formed containing RhoB and HCMV virions, potentially contributing to the successful viral spread. Interestingly, the knockdown of RhoB in HCMV-infected cells resulted in a significant reduction of the virus titer and could also affect the accumulation of AC viral proteins at this subcellular compartment. RhoB knockdown also affected actin fibers' structure. Actin reorganization was observed at late stages of infection originating from the viral AC and surrounding the cellular projections, implying a potential interplay between RhoB and actin during HCMV assembly and egress. In conclusion, our results demonstrate for the first time that RhoB is a constituent of the viral AC and is required for HCMV productive infection.

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Section: Discussionsupporting

confidence: 53%

RhoB is a component of the human cytomegalovirus assembly complex and is required for efficient viral production

et al. 2015

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“…8C), which is reminiscent of the phenotype observed in HSV-1 VP22-null mutants (70,74). We speculate that KSHV ORF52 reinforces the interaction network between virion proteins, much as HSV-1 VP22 and HCMV pp65 do (62,72). The shared and unique structural features and functions of ORF52 homologues in KSHV, MHV-68, RRV, and EBV allude to the common, yet distinct strategies employed by these viruses to maintain persistent infection of their human host.…”

Section: Discussionmentioning

confidence: 74%

“…Similar interactions between tegument proteins of betaherpesviruses have also been reported. The most abundant tegument protein of HCMV, pp65 (pUL83), interacts with pUL25 and pUL43 (60,61), and the loss of pp65 impairs the incorporation of other tegument proteins into virions (62). Interactions among gammaherpesvirus-specific tegument proteins may also exist (38,48,52,63,64).…”

Section: Discussionmentioning

confidence: 99%

Kaposi's Sarcoma-Associated Herpesvirus Inhibitor of cGAS (KicGAS), Encoded by ORF52, Is an Abundant Tegument Protein and Is Required for Production of Infectious Progeny Viruses

Avey

et al. 2016

J Virol

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Although Kaposi's sarcoma-associated herpesvirus (KSHV) ORF52 (also known as KSHV inhibitor of cGAS [KicGAS]) has been detected in purified virions, the roles of this protein during KSHV replication have not been characterized. Using specific monoclonal antibodies, we revealed that ORF52 displays true late gene expression kinetics and confirmed its cytoplasmic localization in both transfected and KSHV-infected cells. We demonstrated that ORF52 comigrates with other known virion proteins following sucrose gradient centrifugation. We also determined that ORF52 resides inside the viral envelope and remains partially associated with capsid when extracellular virions are treated with various detergents and/or salts. There results indicate that ORF52 is a tegument protein abundantly present in extracellular virions. To characterize the roles of ORF52 in the KSHV life cycle, we engineered a recombinant KSHV ORF52-null mutant virus and found that loss of ORF52 results in reduced virion production and a further defect in infectivity. Upon analysis of the virion composition of ORF52-null viral particles, we observed a decrease in the incorporation of ORF45, as well as other tegument proteins, suggesting that ORF52 is important for the packaging of other virion proteins. In summary, our results indicate that, in addition to its immune evasion function, KSHV ORF52 is required for the optimal production of infectious virions, likely due to its roles in virion assembly as a tegument protein. IMPORTANCEThe tegument proteins of herpesviruses, including Kaposi's sarcoma-associated herpesvirus (KSHV), play key roles in the viral life cycle. Each of the three subfamilies of herpesviruses (alpha, beta, and gamma) encode unique tegument proteins with specialized functions. We recently found that one such gammaherpesvirus-specific protein, ORF52, has an important role in immune evasion during KSHV primary infection, through inhibition of the host cytosolic DNA sensing pathway. In this report, we further characterize ORF52 as a tegument protein with vital roles during KSHV lytic replication. We found that ORF52 is important for the production of infectious viral particles, likely through its role in virus assembly, a critical process for KSHV replication and pathogenesis. More comprehensive investigation of the functions of tegument proteins and their roles in viral replication may reveal novel targets for therapeutic interventions against KSHV-associated diseases. K aposi's sarcoma-associated herpesvirus (KSHV), also known as human herpesvirus 8 (HHV-8), is the etiologic agent of Kaposi's sarcoma (KS) (1) and, also, two lymphoproliferative disorders, primary effusion lymphoma (PEL) (2) and multicentric Castleman disease (MCD) (3). KSHV belongs to the Rhadinovirus genus in the Gammaherpesvirinae subfamily and is related to rhesus rhadinovirus (RRV), herpesvirus saimiri (HVS), and murine gammaherpesvirus 68 (MHV-68). The closest relative of KSHV among the known human herpesviruses is Epstein-Barr virus (EBV), which belongs to t...

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“…3A–3C). All cell types expressed both the major immediate early and the pp65 late protein by immunofluorescent staining [25]. Pericytes also expressed the pp28 late antigen (insert Fig.…”

Section: Resultsmentioning

confidence: 99%

Placental pericytes and cytomegalovirus infectivity: Implications for HCMV placental pathology and congenital disease

Aronoff

Correa

Rogers

et al. 2017

American J Rep Immunol

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Problem Placental pericytes are essential for placental microvascular function, stability, and integrity. Mechanisms of HCMV pathogenesis incorporating placental pericytes are unknown. Method of Study HCMV infected placental tissue was stained by dual-labeled immunohistochemistry. Primary placental pericytes, cytotrophoblasts, and villous fibroblasts were exposed to HCMV; infectivity was analyzed by microscopy and immunofluorescence. Cytokine expression was examined by Luminex assay. A HCMV-GFP recombinant virus was used to examine replication kinetics. Results Immunohistochemistry showed HCMV in trophoblast and the villous core with T-cell and macrophage infiltration. SBCMV-infected pericytes showed dysregulation of proinflammatory and angiogenic cytokines when compared to control cells. A tri-cell model of the villous floor showed a unique expression profile. Finally, we show pericytes infected in vivo with HCMV in placental tissue from a congenitally infected child. Conclusions Placental pericytes support HCMV replication, inducing proinflammatory and angiogenic cytokines that likely contribute to viral dissemination, placenta inflammation, and dysregulation of placental angiogenesis.

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The Tegument Protein pp65 of Human Cytomegalovirus Acts as an Optional Scaffold Protein That Optimizes Protein Uploading into Viral Particles

Cited by 29 publications

References 67 publications

RhoB is a component of the human cytomegalovirus assembly complex and is required for efficient viral production

RhoB is a component of the human cytomegalovirus assembly complex and is required for efficient viral production

Kaposi's Sarcoma-Associated Herpesvirus Inhibitor of cGAS (KicGAS), Encoded by ORF52, Is an Abundant Tegument Protein and Is Required for Production of Infectious Progeny Viruses

Placental pericytes and cytomegalovirus infectivity: Implications for HCMV placental pathology and congenital disease

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