scholarly journals Physical and Functional Interactions between the Herpes Simplex Virus UL15 and UL28 DNA Cleavage and Packaging Proteins

1999 ◽  
Vol 73 (2) ◽  
pp. 1704-1707 ◽  
Author(s):  
Kim M. Koslowski ◽  
Patti R. Shaver ◽  
James T. Casey ◽  
Todd Wilson ◽  
Gregory Yamanaka ◽  
...  
Keyword(s):  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Dna Cleavage ◽  
Gradient Centrifugation ◽  
Viral Proteins ◽  
Physical Interaction ◽  
Cell Nuclei ◽  
Infected Cells ◽  
Simplex Virus ◽  
Dna Affinity Chromatography

ABSTRACT Herpes simplex virus (HSV) DNA is cleaved from concatemers and packaged into capsids in infected cell nuclei. This process requires seven viral proteins, including UL15 and UL28. UL15 expressed alone displays a nuclear localization, while UL28 remains cytoplasmic. Coexpression with UL15 enables UL28 to enter nuclei, suggesting an interaction between the two proteins. Additionally, UL28 copurified with UL15 from HSV-infected cells after ion-exchange and DNA affinity chromatography, and the complex sedimented as a 1:1 heterodimer upon sucrose gradient centrifugation. These findings are evidence of a physical interaction of UL15 and UL28 and a functional role for UL15 in directing UL28 to the nucleus.

scholarly journals Effects of Major Capsid Proteins, Capsid Assembly, and DNA Cleavage/Packaging on the pUL17/pUL25 Complex of Herpes Simplex Virus 1

2009 ◽  
Vol 83 (24) ◽  
pp. 12725-12737 ◽  
Author(s):  
Luella Scholtes ◽  
Joel D. Baines
Keyword(s):  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Nuclear Localization ◽  
Conformational Changes ◽  
Viral Proteins ◽  
Dna Packaging ◽  
Capsid Proteins ◽  
Herpes Simplex Virus 1 ◽  
Infected Cells ◽  
Simplex Virus

ABSTRACT The UL17 and UL25 proteins (pUL17 and pUL25, respectively) of herpes simplex virus 1 are located at the external surface of capsids and are essential for DNA packaging and DNA retention in the capsid, respectively. The current studies were undertaken to determine whether DNA packaging or capsid assembly affected the pUL17/pUL25 interaction. We found that pUL17 and pUL25 coimmunoprecipitated from cells infected with wild-type virus, whereas the major capsid protein VP5 (encoded by the UL19 gene) did not coimmunoprecipitate with these proteins under stringent conditions. In addition, pUL17 (i) coimmunoprecipitated with pUL25 in the absence of other viral proteins, (ii) coimmunoprecipitated with pUL25 from lysates of infected cells in the presence or absence of VP5, (iii) did not coimmunoprecipitate efficiently with pUL25 in the absence of the triplex protein VP23 (encoded by the UL18 gene), (iv) required pUL25 for proper solubilization and localization within the viral replication compartment, (v) was essential for the sole nuclear localization of pUL25, and (vi) required capsid proteins VP5 and VP23 for nuclear localization and normal levels of immunoreactivity in an indirect immunofluorescence assay. Proper localization of pUL25 in infected cell nuclei required pUL17, pUL32, and the major capsid proteins VP5 and VP23, but not the DNA packaging protein pUL15. The data suggest that VP23 or triplexes augment the pUL17/pUL25 interaction and that VP23 and VP5 induce conformational changes in pUL17 and pUL25, exposing epitopes that are otherwise partially masked in infected cells. These conformational changes can occur in the absence of DNA packaging. The data indicate that the pUL17/pUL25 complex requires multiple viral proteins and functions for proper localization and biochemical behavior in the infected cell.

scholarly journals Herpes Simplex Virus Entry Mediator Associates in Infected Cells in a Complex with Viral Proteins gD and at Least gH

2005 ◽  
Vol 79 (7) ◽  
pp. 4540-4544 ◽  
Author(s):  
Pilar Perez-Romero ◽  
Aleida Perez ◽  
Althea Capul ◽  
Rebecca Montgomery ◽  
A. Oveta Fuller
Keyword(s):  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Protein Interactions ◽  
Virus Entry ◽  
Viral Proteins ◽  
Receptor Protein ◽  
Infected Cells ◽  
Complex Forms ◽  
Simplex Virus ◽  
In Cells

ABSTRACT We examined herpes simplex virus (HSV)-infected human HEp-2 cells or porcine cells that express herpes virus entry mediator (HVEM) for virus and receptor protein interactions. Antibody to HVEM, or its viral ligand gD, coimmunoprecipitated several similar proteins. A prominent 110-kDa protein that coprecipitated was identified as gH. The HVEM/gD/gH complex was detected with mild or stringent cell lysis conditions. It did not form in cells infected with HSV-1(KOS)Rid1 virus or with null virus lacking gD, gH, or gL. Thus, in cells a complex forms through physical associations of HVEM, gD, and at least gH.

scholarly journals Modified VP22 Localizes to the Cell Nucleus during Synchronized Herpes Simplex Virus Type 1 Infection

1999 ◽  
Vol 73 (8) ◽  
pp. 6769-6781 ◽  
Author(s):  
Lisa E. Pomeranz ◽  
John A. Blaho
Keyword(s):  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Sodium Dodecyl ◽  
Nuclear Fraction ◽  
Relative Distribution ◽  
Cell Nuclei ◽  
Cell Extracts ◽  
Infected Cells ◽  
Simplex Virus ◽  
Hsv 1

ABSTRACT The UL49 gene product (VP22) of herpes simplex virus types 1 and 2 (HSV-1 and HSV-2) is a virion phosphoprotein which accumulates inside infected cells at late stages of infection. We previously (J. A. Blaho, C. Mitchell, and B. Roizman, J. Biol. Chem. 269:17401–17410, 1994) discovered that the form of VP22 packaged into infectious virions differed from VP22 extracted from infected-cell nuclei in that the virion-associated form had a higher electrophoretic mobility in denaturing gels. Based on these results, we proposed that VP22 in virions was “undermodified” in some way. The goal of this study is to document the biological and biochemical properties of VP22 throughout the entire course of a productive HSV-1 infection. We now report the following. (i) VP22 found in infected cells is distributed in at least three distinct subcellular localizations, which we define as cytoplasmic, diffuse, and nuclear, as measured by indirect immunofluorescence. (ii) Using a synchronized infection system, we determined that VP22 exists predominantly in the cytoplasm early in infection and accumulates in the nucleus late in infection. (iii) While cytoplasmic VP22 colocalizes with the HSV-1 glycoprotein D early in infection, the nuclear form of VP22 is not restricted to replication compartments which accumulate ICP4. (iv) VP22 migrates as at least three unique electrophoretic species in denaturing sodium dodecyl sulfate-DATD-polyacrylamide gels. VP22a, VP22b, and VP22c have high, intermediate, and low mobility, respectively. (v) The relative distribution of the various forms of VP22 derived from infected whole-cell extracts varies during the course of infection such that low-mobility species predominate at early times and high-mobility forms accumulate later. (vi) The highest-mobility forms of VP22 partition with the cytoplasmic fraction of infected cells, while the lowest-mobility forms are associated with the nuclear fraction. (vii) Finally, full-length VP22 which partitions in the nucleus incorporates radiolabel from [32P]orthophosphate whereas cytoplasmic VP22 does not. Based on these results, we conclude that modification of VP22 coincides with its appearance in the nucleus during the course of productive HSV-1 infection.

scholarly journals Tryptophan Residues in the Portal Protein of Herpes Simplex Virus 1 Critical to the Interaction with Scaffold Proteins and Incorporation of the Portal into Capsids

2009 ◽  
Vol 83 (22) ◽  
pp. 11726-11733 ◽  
Author(s):  
Kui Yang ◽  
Joel D. Baines
Keyword(s):  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Dna Cleavage ◽  
Scaffold Proteins ◽  
Herpes Simplex Virus 1 ◽  
Hydrophobic Domain ◽  
Portal Protein ◽  
Tryptophan Residues ◽  
Infected Cells ◽  
Simplex Virus

ABSTRACT Incorporation of the herpes simplex virus 1 (HSV-1) portal vertex into the capsid requires interaction with a 12-amino-acid hydrophobic domain within capsid scaffold proteins. The goal of this work was to identify domains and residues in the UL6-encoded portal protein pUL6 critical to the interaction with scaffold proteins. We show that whereas the wild-type portal and scaffold proteins readily coimmunoprecipitated with one another in the absence of other viral proteins, truncation beyond the first 18 or last 36 amino acids of the portal protein precluded this coimmunoprecipitation. The coimmunoprecipitation was also precluded by mutation of conserved tryptophan (W) residues to alanine (A) at positions 27, 90, 127, 163, 241, 262, 532, and 596 of UL6. All of these W-to-A mutations precluded the rescue of a viral deletion mutant lacking UL6, except W163A, which supported replication poorly, and W596A, which fully rescued replication. A recombinant virus bearing the W596A mutation replicated and packaged DNA normally, and scaffold proteins readily coimmunoprecipitated with portal protein from lysates of infected cells. Thus, viral functions compensated for the W596A mutation's detrimental effects on the portal-scaffold interaction seen during transient expression of portal and scaffold proteins. In contrast, the W27A mutation precluded portal-scaffold interactions in infected cell lysates, reduced the solubility of pUL6, decreased incorporation of the portal into capsids, and abrogated viral-DNA cleavage and packaging.

scholarly journals DNA Cleavage and Packaging Proteins Encoded by Genes UL28, UL15, and UL33 of Herpes Simplex Virus Type 1 Form a Complex in Infected Cells

2002 ◽  
Vol 76 (10) ◽  
pp. 4785-4791 ◽  
Author(s):  
Philippa M. Beard ◽  
Naomi S. Taus ◽  
Joel D. Baines
Keyword(s):  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Virus Type ◽  
Herpes Simplex Virus Type ◽  
Dna Cleavage ◽  
Rabbit Polyclonal Antibody ◽  
Infected Cells ◽  
Simplex Virus ◽  
Hsv 1

ABSTRACT Previous studies have indicated that the UL6, UL15, UL17, UL28, UL32, and UL33 genes are required for the cleavage and packaging of herpes simplex viral DNA. To identify proteins that interact with the UL28-encoded DNA binding protein of herpes simplex virus type 1 (HSV-1), a previously undescribed rabbit polyclonal antibody directed against the UL28 protein fused to glutathione S-transferase was used to immunopurify UL28 and the proteins with which it associated. It was found that the antibody specifically coimmunoprecipitated proteins encoded by the genes UL28, UL15, and UL33 from lysates of both HEp-2 cells infected with HSV-1(F) and insect cells infected with recombinant baculoviruses expressing these three proteins. In reciprocal reactions, antibodies directed against the UL15- or UL33-encoded proteins also coimmunoprecipitated the UL28 protein. The coimmunoprecipitation of the three proteins from HSV-infected cells confirms earlier reports of an association between the UL28 and UL15 proteins and represents the first evidence of the involvement of the UL33 protein in this complex.

scholarly journals Role of Cellular Phosphatase cdc25C in Herpes Simplex Virus 1 Replication

2008 ◽  
Vol 82 (9) ◽  
pp. 4527-4532 ◽  
Author(s):  
Benjamin A. Smith-Donald ◽  
Lizette O. Durand ◽  
Bernard Roizman
Keyword(s):  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Cyclin B1 ◽  
Wild Type Virus ◽  
Physical Interaction ◽  
Wild Type ◽  
Herpes Simplex Virus 1 ◽  
Infected Cells ◽  
Simplex Virus

ABSTRACT Earlier studies have shown that in herpes simplex virus 1-infected cells, ICP22 upregulates the accumulation of a subset of γ2 proteins exemplified by the products of the UL38, UL41, and US11 genes. The ICP22-dependent process involves degradation of cyclins A and B1, the stabilization and activation of cdc2, physical interaction of activated cdc2 with the UL42 DNA synthesis processivity factor, and recruitment and phosphorylation of topoisomerase IIα by the cdc2/UL42 complex. Activation of cdc2, the first step in the process, is a key function of the mitotic phosphatase cdc25C. To define the role of cdc25C, we probed some features of the ICP22-dependent pathway of upregulation of γ2 genes in cdc25C−/− cells and in cdc25C+/+ cells derived from sibling mice. We report that cyclin B1 turned over in cdc25C+/+ or cdc25C−/− cells at the same rate, that cdc2 increased in amount, and that US11 and UL38 proteins and infectious virus accumulated in smaller amounts than in wild-type infected cells. The reduction in UL38 protein accumulation and virus was greater in cdc25C−/− cells infected with virus lacking ICP22 than in cells infected with wild-type virus. We conclude that cdc25C phosphatase plays a role in viral replication and that this role extends beyond its function of activating cdc2 for initiation of the ICP22-dependent cascade for upregulation of γ2 gene expression.

The arginine requirement for nucleocapsid maturation in Herpes simplex virus development

1970 ◽  
Vol 16 (9) ◽  
pp. 851-854 ◽  
Author(s):  
Sylvia Stein ◽  
Paul Todd ◽  
Judith Mahoney
Keyword(s):  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Gradient Centrifugation ◽  
Sucrose Density Gradient ◽  
Sucrose Density Gradient Centrifugation ◽  
Virus Growth ◽  
Normal Medium ◽  
Arginine Deprivation ◽  
Infected Cells ◽  
Simplex Virus

Naked and enveloped Herpes simplex virus (HSV) particles from infected HEp-2 cells were separated by sucrose density-gradient centrifugation. Infected cells deprived of arginine during the latter half of the virus growth cycle were found capable of synthesizing about the same total number of naked particles as were infected cells maintained in normal medium throughout the cycle. The formation of full capsids in arginine-deprived cells was greatly depressed. The number of enveloped virus particles in the arginine-deprived culture was one-half that of the normal culture, but those full capsids which were enveloped in both cultures were equally infectious.Evidently late arginine deprivation affects the maturation of HSV nucleocapsids, rather than the synthesis of capsid protein, and the enveloping process appears to be affected by arginine deprivation as well.

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