Brefeldin A inhibits expression of DNA packaging proteins and nucleocapsid formation of human cytomegalovirus

Kregler, Oliver; Schilf, Rita; Lander, Angelika; Bannert, Norbert; Bogner, Elke

doi:10.1016/j.febslet.2009.03.011

Cited by 5 publications

(3 citation statements)

References 36 publications

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“…The sum of vDNA in and vDNA rep is the total cell-associated vDNA (vDNA tot ), which we can experimentally measure. For the purpose of this empirical model, we set free vDNA tot to be in excess compared to genomes packaged into particles destined to leave the infected cell ( 32 – 37 ). To develop the empirical model, we experimentally quantified HCMV vDNA tot using different input MOIs (in infectious units per cell) over 96 h ( Fig.…”

Section: Resultsmentioning

confidence: 99%

“…3 ). This assumption is justified because mature C-type capsids represent a small fraction of the total capsid types in the nucleus, with viral terminase activity requiring an excess of vDNA templates for packaging ( 32 – 37 ). Next, the mechanisms proposed in our model of the late lytic replication cycle represent the lumping of many, potentially unmeasurable, smaller subprocesses.…”

Section: Discussionmentioning

confidence: 99%

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Computational modeling of protracted HCMV replication using genome substrates and protein temporal profiles

Monti

Mokry

Schumacher

et al. 2022

Proc. Natl. Acad. Sci. U.S.A.

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Human cytomegalovirus (HCMV) is a major cause of illness in immunocompromised individuals. The HCMV lytic cycle contributes to the clinical manifestations of infection. The lytic cycle occurs over ∼96 h in diverse cell types and consists of viral DNA (vDNA) genome replication and temporally distinct expression of hundreds of viral proteins. Given its complexity, understanding this elaborate system can be facilitated by the introduction of mechanistic computational modeling of temporal relationships. Therefore, we developed a multiplicity of infection (MOI)-dependent mechanistic computational model that simulates vDNA kinetics and late lytic replication based on in-house experimental data. The predictive capabilities were established by comparison to post hoc experimental data. Computational analysis of combinatorial regulatory mechanisms suggests increasing rates of protein degradation in association with increasing vDNA levels. The model framework also allows expansion to account for additional mechanisms regulating the processes. Simulating vDNA kinetics and the late lytic cycle for a wide range of MOIs yielded several unique observations. These include the presence of saturation behavior at high MOIs, inefficient replication at low MOIs, and a precise range of MOIs in which virus is maximized within a cell type, being 0.382 IU to 0.688 IU per fibroblast. The predicted saturation kinetics at high MOIs are likely related to the physical limitations of cellular machinery, while inefficient replication at low MOIs may indicate a minimum input material required to facilitate infection. In summary, we have developed and demonstrated the utility of a data-driven and expandable computational model simulating lytic HCMV infection.

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Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Computational modeling of protracted HCMV replication using genome substrates and protein temporal profiles

Monti

Mokry

Schumacher

et al. 2022

Proc. Natl. Acad. Sci. U.S.A.

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“…Brefeldin A disrupts clathrin-coated vesicle formation from Golgi, whereas it has no effect on clathrin-coated vesicles formed elsewhere in the cell. Clathrin has been found to be required for late events in the HCMV life cycle (45,46) as treatment with brefeldin A inhibits envelopment. In addition, clathrin is found in highly purified extracellular virions (37).…”

Section: Targeted Proteomics Of Virion Maturation Processesmentioning

confidence: 99%

A Targeted Spatial-Temporal Proteomics Approach Implicates Multiple Cellular Trafficking Pathways in Human Cytomegalovirus Virion Maturation

Moorman¹,

Sharon-Friling²,

Shenk

et al. 2010

Molecular & Cellular Proteomics

114

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The assembly of infectious virus particles is a complex event. For human cytomegalovirus (HCMV) this process requires the coordinated expression and localization of at least 60 viral proteins that comprise the infectious virion. To gain insight into the mechanisms controlling this process, we identified protein binding partners for two viral proteins, pUL99 (also termed pp28) and pUL32 (pp150), which are essential for HCMV virion assembly. We utilized HCMV strains expressing pUL99 or pUL32 carboxyl-terminal green fluorescent protein fusion proteins from their native location in the HCMV genome. Based on the presence of ubiquitin in the pUL99 immunoisolation, we discovered that this viral protein colocalizes with components of the cellular endosomal sorting complex required for transport (ESCRT) pathway during the initial stages of virion assembly. We identified the nucleocapsid and a large number of tegument proteins as pUL32 binding partners, suggesting that events controlling trafficking of this viral protein in the cytoplasm regulate nucleocapsid/tegument maturation. The finding that pUL32, but not pUL99, associates with clathrin led to the discovery that the two viral proteins traffic via distinct pathways during the early stages of virion assembly. Additional investigation revealed that the majority of the major viral glycoprotein gB initially resides in a third compartment. Analysis of the trafficking of these three viral proteins throughout a time course of virion assembly allowed us to visualize their merger into a single large cytoplasmic structure during the late stages of viral assembly. We propose a model of HCMV virion maturation in which multiple components of the virion traffic independently of one another before merging. Molecular & Cellular Proteomics 9:851-860, 2010.

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Sequestration of human cytomegalovirus by human renal and mammary epithelial cells

Andrei

Kummert²,

Donner

et al. 2014

Virology

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Urine and breast milk represent the main routes of human cytomegalovirus (HCMV) transmission but the contribution of renal and mammary epithelial cells to viral excretion remains unclear. We observed that kidney and mammary epithelial cells were permissive to HCMV infection and expressed immediate early, early and late antigens within 72 h of infection. During the first 24 h after infection, high titers of infectious virus were measured associated to the cells and in culture supernatants, independently of de novo synthesis of virus progeny. This phenomenon was not observed in HCMV-infected fibroblasts and suggested the sequestration and the release of HCMV by epithelial cells. This hypothesis was supported by confocal and electron microscopy analyses. The sequestration and progressive release of HCMV by kidney and mammary epithelial cells may play an important role in the excretion of the virus in urine and breast milk and may thereby contribute to HCMV transmission.

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Brefeldin A inhibits expression of DNA packaging proteins and nucleocapsid formation of human cytomegalovirus

Cited by 5 publications

References 36 publications

Computational modeling of protracted HCMV replication using genome substrates and protein temporal profiles

Computational modeling of protracted HCMV replication using genome substrates and protein temporal profiles

A Targeted Spatial-Temporal Proteomics Approach Implicates Multiple Cellular Trafficking Pathways in Human Cytomegalovirus Virion Maturation

Sequestration of human cytomegalovirus by human renal and mammary epithelial cells

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