scholarly journals Varicella-Zoster Virus ORF47 Protein Kinase, Which Is Required for Replication in Human T Cells, and ORF66 Protein Kinase, Which Is Expressed during Latency, Are Dispensable for Establishment of Latency

2003 ◽  
Vol 77 (20) ◽  
pp. 11180-11185 ◽  
Author(s):  
Hitoshi Sato ◽  
Lesley Pesnicak ◽  
Jeffrey I. Cohen
Keyword(s):  
T Cells ◽  
Protein Kinase ◽  
Varicella Zoster Virus ◽  
Latent Infection ◽  
Viral Proteins ◽  
Parental Virus ◽  
Reading Frame ◽  
Varicella Zoster ◽  
Human T Cells ◽  
Simplex Virus

ABSTRACT Varicella-zoster virus (VZV) results in a lifelong latent infection in human sensory and cranial nerve ganglia after primary infection. VZV open reading frame 47 (ORF47) and ORF66 encode protein kinases that phosphorylate several viral proteins, including VZV glycoprotein gE and ORF32, ORF62, and ORF63 proteins. Here we show that the ORF47 protein kinase also phosphorylates gI. While ORF47 is essential for virus replication in human T cells and skin, we found the gene to be dispensable for establishment of latent infection in dorsal root ganglia of rodents. ORF66 protein is expressed during latency. Rodents infected with VZV unable to express ORF66 developed latent infection at a rate similar to that for the parental virus. ORF63 transcripts, a hallmark of VZV latency, were also detected in similar numbers of animals infected with the ORF47 and ORF66 mutants and with the parental virus. VZV mutants unable to express four of the six genes that do not have herpes simplex virus (HSV) homologs (ORFs 1, 13, 32, 57) were also unimpaired for establishment of latency. While a truncated HSV VP16 mutant was previously reported to be unable to establish latency in a mouse model, we found that VZV with a deletion of ORF10, the homolog of HSV VP16, was dispensable for establishment of latency. Thus, seven genes, including one expressed during latency, are dispensable for establishing latent VZV infection.

scholarly journals Varicella-Zoster Virus (VZV) ORF17 Protein Induces RNA Cleavage and Is Critical for Replication of VZV at 37oC but Not 33oC

2002 ◽  
Vol 76 (21) ◽  
pp. 11012-11023 ◽  
Author(s):  
Hitoshi Sato ◽  
Lawrence D. Callanan ◽  
Lesley Pesnicak ◽  
Tammy Krogmann ◽  
Jeffrey I. Cohen
Keyword(s):  
Varicella Zoster Virus ◽  
Deletion Mutant ◽  
Host Protein ◽  
Parental Virus ◽  
Rna Cleavage ◽  
Reading Frame ◽  
Varicella Zoster ◽  
Cotton Rats ◽  
Simplex Virus

ABSTRACT Varicella-zoster virus (VZV) open reading frame 17 (ORF17) is homologous to herpes simplex virus (HSV) UL41, which encodes the viral host shutoff protein (vhs). HSV vhs induces degradation of mRNA and rapid shutoff of host protein synthesis. An antibody to ORF17 protein detected a 46-kDa protein in VZV-infected cells. While HSV vhs is located in virions, VZV ORF17 protein was not detectable in virions. ORF17 protein induced RNA cleavage, but to a substantially lesser extent than HSV-1 vhs. Expression of ORF17 protein did not inhibit expression from a β-galactosidase reporter plasmid, while HSV type 1 vhs abolished reporter expression. Two VZV ORF17 deletion mutants were constructed to examine the role of ORF17 in virus replication. While the ORF17 VZV mutants grew to peak titers that were similar to those of the parental virus at 33°C, the ORF17 mutants grew to 20- to 35-fold-lower titers than parental virus at 37°C. ORF62 protein was distributed in a different pattern in the nuclei and cytoplasm of cells infected with an ORF17 deletion mutant at 37°C compared to 33°C. Inoculation of cotton rats with the ORF17 deletion mutant resulted in a level of latent infection similar to that produced by inoculation with the parental virus. The importance of ORF17 protein for viral replication at 37°C but not at 33°C suggests that this protein may facilitate the growth of virus in certain tissues in vivo.

scholarly journals Dissecting the Molecular Mechanisms of the Tropism of Varicella-Zoster Virus for Human T Cells

2016 ◽  
Vol 90 (7) ◽  
pp. 3284-3287 ◽  
Author(s):  
Nandini Sen ◽  
Ann M. Arvin
Keyword(s):  
T Cells ◽  
T Cell ◽  
Varicella Zoster Virus ◽  
Molecular Mechanisms ◽  
Cell Mass ◽  
Surface Proteins ◽  
Reading Frame ◽  
Varicella Zoster ◽  
Human T Cells ◽  
Skin Homing

Studies of varicella-zoster virus (VZV) tropism for T cells support their role in viral transport to the skin during primary infection. Multiparametric single-cell mass cytometry demonstrates that, instead of preferentially infecting skin-homing T cells, VZV alters cell signaling and remodels surface proteins to enhance T cell skin trafficking. Viral proteins dispensable in skin, such as that encoded by open reading frame 66, are necessary in T cells. Interference with VZV T cell tropism may offer novel strategies for drug and vaccine design.

scholarly journals Varicella-Zoster Virus (VZV) ORF32 Encodes a Phosphoprotein That Is Posttranslationally Modified by the VZV ORF47 Protein Kinase

1998 ◽  
Vol 72 (10) ◽  
pp. 8083-8088 ◽  
Author(s):  
Sanjay M. Reddy ◽  
Edward Cox ◽  
Ilya Iofin ◽  
Weily Soong ◽  
Jeffrey I. Cohen
Keyword(s):  
Protein Kinase ◽  
Varicella Zoster Virus ◽  
Protein Kinases ◽  
Gene Products ◽  
Osteosarcoma Cells ◽  
Reading Frame ◽  
Varicella Zoster ◽  
Intestine Alkaline Phosphatase ◽  
Infected Cells ◽  
Simplex Virus

ABSTRACT Varicella-zoster virus (VZV) encodes five gene products that do not have homologs in herpes simplex virus. One of these genes, VZV open reading frame 32 (ORF32), is predicted to encode a protein of 16 kDa. VZV ORF32 protein was shown to be phosphorylated and located in the cytosol of virus-infected cells. Antibody to ORF32 protein immunoprecipitated 16- and 18-kDa phosphoproteins from VZV-infected cells. Since VZV encodes two protein kinases that might phosphorylate ORF32 protein, immunoprecipitations were performed with cells infected with VZV mutants unable to express either of the viral protein kinases. Cells infected with VZV unable to express the ORF66 protein kinase contained both the 16- and 18-kDa ORF32 phosphoproteins; however, cells infected with the VZV ORF47 protein kinase mutant showed only the 16-kDa ORF32 phosphoprotein. Treatment of [35S]methionine-labeled proteins with calf intestine alkaline phosphatase resulted in a decrease in size of the ORF32 proteins from 16 and 18 kDa to 15 and 17 kDa, respectively. VZV unable to express ORF32 protein replicated in human melanoma cells to titers similar to those seen with parental virus; however, VZV unable to express ORF32 was impaired for replication in U20S osteosarcoma cells. Thus, VZV ORF32 protein is posttranslationally modified by the ORF47 protein kinase. Since the VZV ORF47 protein kinase has recently been shown to be critical for replication in human fetal skin and lymphocytes, its ability to modify the ORF32 protein suggests that the latter protein may have a role for VZV replication in human tissues.

scholarly journals Varicella-Zoster Virus Open Reading Frame 66 Protein Kinase Is Required for Efficient Viral Growth in Primary Human Corneal Stromal Fibroblast Cells

2008 ◽  
Vol 82 (15) ◽  
pp. 7653-7665 ◽  
Author(s):  
Angela Erazo ◽  
Michael B. Yee ◽  
Nikolaus Osterrieder ◽  
Paul R. Kinchington
Keyword(s):  
T Cells ◽  
Protein Kinase ◽  
Varicella Zoster Virus ◽  
Nuclear Import ◽  
Fluorescent Protein ◽  
Regulatory Protein ◽  
Open Reading Frame ◽  
Stromal Fibroblast ◽  
Reading Frame ◽  
Varicella Zoster

ABSTRACT Varicella-zoster virus (VZV) open reading frame 66 (ORF66) encodes a serine/threonine protein kinase that is not required for VZV growth in most cell types but is needed for efficient growth in T cells. The ORF66 kinase affects nuclear import and virion packaging of IE62, the major regulatory protein, and is known to regulate apoptosis in T cells. Here, we further examined the importance of ORF66 using VZV recombinants expressing green fluorescent protein (GFP)-tagged functional and kinase-negative ORF66 proteins. VZV virions with truncated or kinase-inactivated ORF66 protein were marginally reduced for growth and progeny yields in MRC-5 fibroblasts but were severely growth and replication impaired in low-passage primary human corneal stromal fibroblasts (PCF). To determine if the growth impairment was due to ORF66 kinase regulation of IE62 nuclear import, recombinant VZVs that expressed IE62 with alanine residues at S686, the suspected target by which ORF66 kinase blocks IE62 nuclear import, were made. IE62 S686A expressed by the VZV recombinant remained nuclear throughout infection and was not packaged into virions. However, the mutant virus still replicated efficiently in PCF cells. We also show that inactivation of the ORF66 kinase resulted in only marginally increased levels of apoptosis in PCF cells, which could not fully account for the cell-specific growth requirement of ORF66 kinase. Thus, the unique short region VZV kinase has important cell-type-specific functions that are separate from those affecting IE62 and apoptosis.

scholarly journals Varicella-Zoster Virus Open Reading Frame 21, Which Is Expressed during Latency, Is Essential for Virus Replication but Dispensable for Establishment of Latency

2003 ◽  
Vol 77 (2) ◽  
pp. 1211-1218 ◽  
Author(s):  
Dongxiang Xia ◽  
Shamala Srinivas ◽  
Hitoshi Sato ◽  
Lesley Pesnicak ◽  
Stephen E. Straus ◽  
...  
Keyword(s):  
Varicella Zoster Virus ◽  
Deletion Mutant ◽  
Latent Infection ◽  
Open Reading Frame ◽  
Protein Insertion ◽  
Reading Frame ◽  
Varicella Zoster ◽  
Latently Infected ◽  
Simplex Virus ◽  
Hsv 1

ABSTRACT Varicella-zoster virus (VZV) open reading frame 21 (ORF21) is one of at least five VZV genes expressed in latently infected human and rodent ganglia. To determine whether ORF21 is required for latent and lytic infection, we deleted 99% of ORF21 from the viral genome. The ORF21 deletion mutant virus could be propagated only in a cell line expressing the ORF21 protein. Insertion of the herpes simplex virus type 1 (HSV-1) homolog of VZV ORF21, HSV-1 UL37, into the ORF21 deletion mutant failed to complement the mutant for growth in cell culture. Inoculation of cotton rats with the ORF21 deletion virus resulted in latent infection in numbers of animals similar to those infected after inoculation with the parental virus. The mean numbers of latent VZV genomes were similar in animals infected with parental and ORF21 deletion viruses. Transcription of ORF63, another latency-associated gene, was detected in ganglia from similar numbers of animals infected with the mutant and parental viruses. Thus, ORF21 is the first VZV gene expressed during latency that has been shown to be dispensable for the establishment of latent infection.

scholarly journals Varicella-Zoster Virus Open Reading Frame 2 Encodes a Membrane Phosphoprotein That Is Dispensable for Viral Replication and for Establishment of Latency

2002 ◽  
Vol 76 (7) ◽  
pp. 3575-3578 ◽  
Author(s):  
Hitoshi Sato ◽  
Lesley Pesnicak ◽  
Jeffrey I. Cohen
Keyword(s):  
Varicella Zoster Virus ◽  
Deletion Mutant ◽  
Latent Infection ◽  
Open Reading Frame ◽  
Reading Frame ◽  
Herpesvirus Type ◽  
Varicella Zoster ◽  
Latently Infected ◽  
Simplex Virus ◽  
Open Reading Frame 2

ABSTRACT Varicella-zoster virus (VZV) encodes six genes that do not have homologs in herpes simplex virus. One of these genes, VZV open reading frame 2 (ORF2), was expressed as a 31-kDa phosphoprotein in the membranes of infected cells. Unlike equine and bovine herpesvirus type 1 ORF2 homologs that are associated with virions, VZV virions contained no detectable ORF2 protein. The ORF2 deletion mutant established a latent infection in cotton rats at a frequency and with a number of VZV genomes similar to that of the parental virus. ORF63 transcripts, a hallmark of latent infection, were present in ganglia latently infected with both the ORF2 deletion mutant and parental VZV. Thus, ORF2 is the first VZV gene shown to be dispensable for establishment of latent infection in an animal model.

scholarly journals Microarray Analysis of Host Cell Gene Transcription in Response to Varicella-Zoster Virus Infection of Human T Cells and Fibroblasts In Vitro and SCIDhu Skin Xenografts In Vivo

2003 ◽  
Vol 77 (2) ◽  
pp. 1268-1280 ◽  
Author(s):  
Jeremy O. Jones ◽  
Ann M. Arvin
Keyword(s):  
T Cells ◽  
Gene Regulation ◽  
Varicella Zoster Virus ◽  
Host Cell ◽  
Gene Transcription ◽  
Cell Types ◽  
Cellular Gene ◽  
Varicella Zoster ◽  
Human T Cells

ABSTRACT During primary infection, varicella-zoster virus (VZV) is spread via lymphocytes to skin, where it induces a rash and establishes latency in sensory ganglia. A live, attenuated varicella vaccine (vOka) was generated by using the VZV Oka strain (pOka), but the molecular basis for vOka attenuation remains unknown. Little is known concerning the effects of wild-type or attenuated VZV on cellular gene regulation in the host cells that are critical for pathogenesis. In this study, transcriptional profiles of primary human T cells and fibroblasts infected with VZV in cell culture were determined by using 40,000-spot human cDNA microarrays. Cellular gene transcription in human skin xenografts in SCID mice that were infected with VZV in vivo was also evaluated. The profiles of cellular gene transcripts that were induced or inhibited in infected human foreskin fibroblasts (HFFs), T cells, and skin in response to pOka and vOka infection were similar. However, significant alterations in cellular gene regulation were observed among the three differentiated human cell types that were examined, suggesting specific differences in the biological consequences of VZV infection related to the target cell. Changes in cellular gene transcription detected by microarray analysis were confirmed for selected genes by quantitative real-time reverse transcription-PCR analysis of VZV-infected cells. Interestingly, the transcription of caspase 8 was found to be decreased in infected T cells but not in HFFs or skin, which may signify a tissue-specific antiapoptosis mechanism. The use of microarrays to demonstrate differences in effects on host cell genes in primary, biologically relevant cell types provides background information for experiments to link these various response phenotypes with mechanisms of VZV pathogenesis that are important for the natural course of human infection.

scholarly journals Nuclear Accumulation of IE62, the Varicella-Zoster Virus (VZV) Major Transcriptional Regulatory Protein, Is Inhibited by Phosphorylation Mediated by the VZV Open Reading Frame 66 Protein Kinase

2000 ◽  
Vol 74 (5) ◽  
pp. 2265-2277 ◽  
Author(s):  
Paul R. Kinchington ◽  
Karen Fite ◽  
Stephanie E. Turse
Keyword(s):  
Protein Kinase ◽  
Varicella Zoster Virus ◽  
Nuclear Localization ◽  
Kinase Activity ◽  
Open Reading Frame ◽  
Nuclear Accumulation ◽  
Protein Kinase Activity ◽  
Reading Frame ◽  
Varicella Zoster ◽  
In Cells

ABSTRACT IE62, the major transcriptional activator protein encoded by varicella-zoster virus (VZV), locates to the nucleus when expressed in transfected cells. We show here that cytoplasmic forms of IE62 accumulate in transfected and VZV-infected cells as the result of the protein kinase activity associated with VZV open reading frame 66 (ORF66). Expression of the ORF66 protein kinase but not the VZV ORF47 protein kinase impaired the ability of coexpressed IE62 to transactivate promoter-reporter constructs. IE62 that was coexpressed with the ORF66 protein accumulated predominantly in the cytoplasm, whereas the normal nuclear localization of other proteins was not affected by the ORF66 protein. In cells infected with VZV, IE62 accumulated in the cytoplasm at late times of infection, whereas in cells infected with a VZV recombinant unable to express ORF66 protein (ROka66S), IE62 was completely nuclear. Point mutations introduced into the predicted serine/threonine catalytic domain and ATP binding domain of ORF66 abrogated its ability to influence IE62 nuclear localization, indicating that the protein kinase activity was required. The region of IE62 that was targeted by ORF66 was mapped to amino acids 602 to 733. IE62 peptides containing this region were specifically phosphorylated in cells coexpressing the ORF66 protein kinase and in cells infected with wild-type VZV but were not phosphorylated in cells infected with ROka66S. We conclude that the ORF66 protein kinase phosphorylates IE62 to induce its cytoplasmic accumulation, most likely by inhibiting IE62 nuclear import.

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