Varicella-Zoster Virus Infection Induces Autophagy in both Cultured Cells and Human Skin Vesicles

ABSTRACT When grown in cultured cells, varicella-zoster virus (VZV) forms many aberrant light particles and produces low titers. Various studies have explored the reasons for such a phenotype and have pointed to impaired expression of specific late genes and at lysosomal targeting of egressing virions as possible causes. In the studies presented here, we report that the autophagic degradation pathway was induced at late time points after VZV infection of cultured cells, as documented by immunoblot analysis of the cellular proteins LC3B and p62/SQSTM1, along with electron microscopy analysis, which demonstrated the presence of both early autophagosomes and late autophagic compartments. Autophagy was induced in infected cells even in the presence of phosphonoacetic acid, an inhibitor of viral late gene expression, thus suggesting that accumulation of immediate-early and early viral gene products might be the major stimulus for its induction. We also showed that the autophagic response was not dependent on a specific cell substrate, virus strain, or type of inoculum. Finally, using immunofluorescence imaging, we demonstrated autophagosome-specific staining in human zoster vesicles but not in normal skin. Thus, our results document that this innate immune response pathway is a component of the VZV infectious cycle in both cultured cells and the human skin vesicle, the final site of virion formation in the infected human host.

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Cellular Stress Response to Varicella-Zoster Virus Infection of Human Skin Includes Highly Elevated Interleukin-6 Expression

Open Forum Infectious Diseases ◽

10.1093/ofid/ofy118 ◽

2018 ◽

Vol 5 (6) ◽

Cited By ~ 6

Author(s):

Keith W Jarosinski ◽

John E Carpenter ◽

Erin M Buckingham ◽

Wallen Jackson ◽

Kevin Knudtson ◽

...

Keyword(s):

Stress Response ◽

Varicella Zoster Virus ◽

Human Skin ◽

Cellular Stress ◽

Cellular Stress Response ◽

Varicella Zoster Virus Infection ◽

Varicella Zoster ◽

Protein Levels ◽

Skin Explants ◽

Infected Skin

Abstract Background The infectious cycle of varicella-zoster virus (VZV) after reactivation from the dorsal root ganglia includes replication and assembly of complete enveloped virions in the human skin to cause the characteristic herpes zoster (shingles). Methods To pursue studies of innate immunity to VZV infection, we have adapted a fetal skin organ culture model to a human neonatal foreskin explant model. Results Abundant expression of VZV IE62, gE, and gC was visualized by confocal microscopy while numerous enveloped virions were observed by electron microscopy in infected skin organ cultures. Microarray experiments demonstrated that the patterns of upregulated transcripts differed between VZV-infected cells and VZV-infected skin explants. One result stood out, namely a >30-fold elevated interleukin (IL)-6 level in the infected skin explant that was not present in the infected monolayer culture. The IL-6 results in the polyermase chain reaction (PCR) assay were reproduced by quantitative PCR testing with newly designed primers. To determine if increased transcription was accompanied by increased IL-6 expression, we quantitated the levels of IL-6 protein in the explant media at increasing intervals after infection. We found a statistically significant increase in IL-6 protein levels secreted into the media from VZV-infected skin explants as compared with mock-infected explants. Conclusions The cellular stress response to VZV infection in neonatal skin explants included highly elevated levels of IL-6 transcription and expression. This skin organ model could be adapted to other viruses with a skin tropism, such as herpes simplex virus.

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Varicella-Zoster Virus Infection of Human Foreskin Fibroblast Cells Results in Atypical Cyclin Expression and Cyclin-Dependent Kinase Activity

Journal of Virology ◽

10.1128/jvi.00163-06 ◽

2006 ◽

Vol 80 (11) ◽

pp. 5577-5587 ◽

Cited By ~ 17

Author(s):

Stacey A. Leisenfelder ◽

Jennifer F. Moffat

Keyword(s):

Cell Cycle ◽

Varicella Zoster Virus ◽

Cultured Cells ◽

Cyclin B1 ◽

Human Infection ◽

Dermal Fibroblasts ◽

Cyclin D3 ◽

Varicella Zoster Virus Infection ◽

Varicella Zoster ◽

Protein Levels

ABSTRACT In its course of human infection, varicella-zoster virus (VZV) infects rarely dividing cells such as dermal fibroblasts, differentiated keratinocytes, mature T cells, and neurons, none of which are actively synthesizing DNA; however, VZV is able to productively infect them and use their machinery to replicate the viral genome. We hypothesized that VZV alters the intracellular environment to favor viral replication by dysregulating cell cycle proteins and kinases. Cyclin-dependent kinases (CDKs) and cyclins displayed a highly unusual profile in VZV-infected confluent fibroblasts: total amounts of CDK1, CDK2, cyclin B1, cyclin D3, and cyclin A protein increased, and kinase activities of CDK2, CDK4, and cyclin B1 were strongly and simultaneously induced. Cyclins B1 and D3 increased as early as 24 h after infection, concurrent with VZV protein synthesis. Confocal microscopy indicated that cyclin D3 overexpression was limited to areas of IE62 production, whereas cyclin B1 expression was irregular across the VZV plaque. Downstream substrates of CDKs, including pRb, p107, and GM130, did not show phosphorylation by immunoblotting, and p21 and p27 protein levels were increased following infection. Finally, although the complement of cyclin expression and high CDK activity indicated a progression through the S and G2 phases of the cell cycle, DNA staining and flow cytometry indicated a possible G1/S blockade in infected cells. These data support earlier studies showing that pharmacological CDK inhibitors can inhibit VZV replication in cultured cells.

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Varicella Zoster Virus in the Nervous System

F1000Research ◽

10.12688/f1000research.7153.1 ◽

2015 ◽

Vol 4 ◽

pp. 1356 ◽

Cited By ~ 19

Author(s):

Don Gilden ◽

Maria Nagel ◽

Randall Cohrs ◽

Ravi Mahalingam ◽

Nicholas Baird

Keyword(s):

Nervous System ◽

Varicella Zoster Virus ◽

Clinical Laboratory ◽

Viral Gene Expression ◽

Neurological Complications ◽

Viral Gene ◽

Cell Mediated Immunity ◽

Specific Cell ◽

Varicella Zoster ◽

Cranial Nerve Palsies

Varicella zoster virus (VZV) is a ubiquitous, exclusively human alphaherpesvirus. Primary infection usually results in varicella (chickenpox), after which VZV becomes latent in ganglionic neurons along the entire neuraxis. As VZV-specific cell-mediated immunity declines in elderly and immunocompromised individuals, VZV reactivates and causes herpes zoster (shingles), frequently complicated by postherpetic neuralgia. VZV reactivation also produces multiple serious neurological and ocular diseases, such as cranial nerve palsies, meningoencephalitis, myelopathy, and VZV vasculopathy, including giant cell arteritis, with or without associated rash. Herein, we review the clinical, laboratory, imaging, and pathological features of neurological complications of VZV reactivation as well as diagnostic tests to verify VZV infection of the nervous system. Updates on the physical state of VZV DNA and viral gene expression in latently infected ganglia, neuronal, and primate models to study varicella pathogenesis and immunity are presented along with innovations in the immunization of elderly individuals to prevent VZV reactivation.

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Differential Requirement for Cell Fusion and Virion Formation in the Pathogenesis of Varicella-Zoster Virus Infection in Skin and T Cells

Journal of Virology ◽

10.1128/jvi.78.23.13293-13305.2004 ◽

2004 ◽

Vol 78 (23) ◽

pp. 13293-13305 ◽

Cited By ~ 29

Author(s):

Jaya Besser ◽

Minako Ikoma ◽

Konstanze Fabel ◽

Marvin H. Sommer ◽

Leigh Zerboni ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

Varicella Zoster Virus ◽

Cell Fusion ◽

Cultured Cells ◽

Varicella Zoster Virus Infection ◽

Virion Assembly ◽

Varicella Zoster ◽

Human T Cell ◽

Virion Formation

ABSTRACT The protein product of varicella-zoster virus (VZV) ORF47 is a serine/threonine protein kinase and tegument component. Evaluation of two recombinants of the Oka strain, rOka47ΔC, with a C-terminal truncation of ORF47, and rOka47D-N, with a point mutation in the conserved kinase motif, showed that ORF47 kinase function was necessary for optimal VZV replication in human skin xenografts in SCID mice but not in cultured cells. We now demonstrate that rOka47ΔC and rOka47D-N mutants do not infect human T-cell xenografts. Differences in the growth of kinase-defective ORF47 mutants allowed an examination of requirements for VZV pathogenesis in skin and T cells in vivo. Although virion assembly was reduced and no virion transport to cell surfaces was observed, epidermal cell fusion persisted, and VZV polykaryocytes were generated by rOka47ΔC and rOka47D-N in skin. Virion assembly was also impaired in vitro, but VZV-induced cell fusion continued to cause syncytia in cultured cells infected with rOka47ΔC or rOka47D-N. Intracellular trafficking of envelope glycoprotein E and the ORF47 and IE62 proteins, components of the tegument, was aberrant without ORF47 kinase activity. In summary, normal VZV virion assembly appears to require ORF47 kinase function. Cell fusion was induced by ORF47 mutants in skin, and cell-cell spread occurred even though virion formation was deficient. VZV-infected T cells do not undergo cell fusion, and impaired virion assembly by ORF47 mutants was associated with a complete elimination of T-cell infectivity. These observations suggest a differential requirement for cell fusion and virion formation in the pathogenesis of VZV infection in skin and T cells.

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Productive Varicella-Zoster Virus Infection of Cultured Intact Human Ganglia

Journal of Virology ◽

10.1128/jvi.02793-06 ◽

2007 ◽

Vol 81 (12) ◽

pp. 6752-6756 ◽

Cited By ~ 28

Author(s):

Kavitha Gowrishankar ◽

Barry Slobedman ◽

Anthony L. Cunningham ◽

Monica Miranda-Saksena ◽

Ross A. Boadle ◽

...

Keyword(s):

Electron Microscopy ◽

Varicella Zoster Virus ◽

Dorsal Root ◽

Cultured Cells ◽

Varicella Zoster Virus Infection ◽

Sensory Ganglia ◽

Varicella Zoster ◽

Viral Antigens ◽

Transmission Electron ◽

Species Specific

ABSTRACT Varicella-zoster virus (VZV) is a species-specific herpesvirus which infects sensory ganglia. We have developed a model of infection of human intact explant dorsal root ganglia (DRG). Following exposure of DRG to VZV, viral antigens were detected in neurons and nonneuronal cells. Enveloped virions were visualized by transmission electron microscopy in neurons and nonneuronal cells and within the extracellular space. Moreover, rather than remaining highly cell associated during infection of cultured cells, such as fibroblasts, cell-free VZV was released from infected DRG. This model enables VZV infection of ganglionic cells to be studied in the context of intact DRG.

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Discordant varicella-zoster virus glycoprotein C expression and localization between cultured cells and human skin vesicles

Virology ◽

10.1016/j.virol.2008.09.031 ◽

2008 ◽

Vol 382 (2) ◽

pp. 171-181 ◽

Cited By ~ 20

Author(s):

Johnathan Storlie ◽

John E. Carpenter ◽

Wallen Jackson ◽

Charles Grose

Keyword(s):

Varicella Zoster Virus ◽

Human Skin ◽

Cultured Cells ◽

Varicella Zoster ◽

Glycoprotein C ◽

Virus Glycoprotein

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Transcriptomal analysis of varicella-zoster virus infection using long oligonucleotide-based microarrays

Journal of General Virology ◽

10.1099/vir.0.80946-0 ◽

2005 ◽

Vol 86 (10) ◽

pp. 2673-2684 ◽

Cited By ~ 35

Author(s):

Peter G. E. Kennedy ◽

Esther Grinfeld ◽

Marie Craigon ◽

Klemens Vierlinger ◽

Douglas Roy ◽

...

Keyword(s):

Gene Expression ◽

Varicella Zoster Virus ◽

Gene Transcription ◽

Spinal Ganglia ◽

Viral Gene ◽

Varicella Zoster Virus Infection ◽

Global Pattern ◽

Varicella Zoster ◽

Viral Gene Transcription ◽

Diagnostic Potential

Varicella-zoster virus (VZV) is a human herpes virus that causes varicella as a primary infection and herpes zoster following reactivation of the virus from a latent state in trigeminal and spinal ganglia. In order to study the global pattern of VZV gene transcription, VZV microarrays using 75-base oligomers to 71 VZV open reading frames (ORFs) were designed and validated. The long-oligonucleotide approach maximizes the stringency of detection and polarity of gene expression. To optimize sensitivity, microarrays were hybridized to target RNA and the extent of hybridization measured using resonance light scattering. Microarray data were normalized to a subset of invariant ranked host-encoded positive-control genes and the data subjected to robust formal statistical analysis. The programme of viral gene expression was determined for VZV (Dumas strain)-infected MeWo cells and SVG cells (an immortalized human astrocyte cell line) 72 h post-infection. Marked quantitative and qualitative differences in the viral transcriptome were observed between the two different cell types using the Dumas laboratory-adapted strain. Oligonucleotide-based VZV arrays have considerable promise as a valuable tool in the analysis of viral gene transcription during both lytic and latent infections, and the observed heterogeneity in the global pattern of viral gene transcription may also have diagnostic potential.

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Ileocolic Intussusception as the Presenting Symptom of Primary Enteric Varicella-Zoster Virus Infection in a 7-Month-Old Infant

The Journal of Infectious Diseases ◽

10.1093/infdis/jiaa148 ◽

2020 ◽

Vol 222 (2) ◽

pp. 305-308

Author(s):

Jonathan D Windster ◽

Werner J D Ouwendijk ◽

Cornelius E J Sloots ◽

Georges M G M Verjans ◽

Robert M Verdijk

Keyword(s):

Varicella Zoster Virus ◽

Ganglion Cells ◽

Varicella Zoster Virus Infection ◽

Specific Cell ◽

Wild Type ◽

Ileocolic Intussusception ◽

Varicella Zoster ◽

Resected Intestine ◽

Immunocompetent Infant

Abstract Ileocolic intussusception is the invagination of ileum into the colon. In a subset of patients, the disease is caused by mesenteric lymphadenopathy in response to (viral) infection. We present a case of an ileocolic intussusception necessitating surgery in a 7-month-old immunocompetent infant with concurrent primary wild-type varicella-zoster virus (VZV) infection, in whom chickenpox rash developed 2 days after surgery. Detailed in situ analyses of resected intestine for specific cell type markers and VZV RNA demonstrated VZV-infected lymphocytes and neurons in the gut wall and in ganglion cells of the myenteric plexus.

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