Full trans -activation mediated by the immediate-early protein of equine herpesvirus 1 requires a consensus TATA box, but not its cognate binding sequence

ABSTRACT The sole immediate-early (IE) gene of equine herpesvirus 1 encodes a 1,487-amino-acid (aa) regulatory phosphoprotein that independently activates expression of early viral genes. Coimmunoprecipitation assays demonstrated that the IE protein physically interacts with the general transcription factor TFIIB. Using a variety of protein-binding assays that employed a panel of IE truncation and deletion mutants expressed as in vitro-synthesized or glutathione S-transferase fusion proteins, we mapped a TFIIB-binding domain to aa 407 to 757 of the IE protein. IE mutants carrying internal deletions of aa 426 to 578 and 621 to 757 were partially defective for TFIIB binding, indicating that aa 407 to 757 may harbor more than one TFIIB-binding domain. The interaction between the IE protein and TFIIB is of physiological importance, as evidenced by transient-cotransfection assays. Partial deletion of the TFIIB-binding domain within the IE protein inhibited its ability to activate expression of the viral thymidine kinase gene, a representative early promoter, and of the IR5 gene, a representative late promoter, by greater than 20 and 50%, respectively. These results indicate that the interaction of the IE protein with TFIIB is necessary for its full transactivation function and that the IE-TFIIB interaction may be part of the mechanism by which the IE protein activates transcription.

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The EICP27 protein of equine herpesvirus 1 is recruited to viral promoters by its interaction with the immediate-early protein

Virology ◽

10.1016/j.virol.2004.12.014 ◽

2005 ◽

Vol 333 (1) ◽

pp. 74-87 ◽

Cited By ~ 14

Author(s):

Randy A. Albrecht ◽

Seong K. Kim ◽

Dennis J. O'Callaghan

Keyword(s):

Immediate Early ◽

Equine Herpesvirus ◽

Equine Herpesvirus 1 ◽

Early Protein ◽

Viral Promoters ◽

Herpesvirus 1

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Interferon Gamma Inhibits Equine Herpesvirus 1 Replication in a Cell Line-Dependent Manner

Pathogens ◽

10.3390/pathogens10040484 ◽

2021 ◽

Vol 10 (4) ◽

pp. 484

Author(s):

Seong K. Kim ◽

Akhalesh K. Shakya ◽

Dennis J. O’Callaghan

Keyword(s):

Cell Line ◽

Immediate Early ◽

M Cells ◽

Dependent Manner ◽

Equine Herpesvirus ◽

Equine Herpesvirus 1 ◽

Early Protein ◽

A Cell ◽

Ifn Γ ◽

Herpesvirus 1

The sole equine herpesvirus 1 (EHV-1) immediate-early protein (IEP) is essential for viral replication by transactivating viral immediate-early (IE), early (E), and late (L) genes. Here, we report that treatment of mouse MH-S, equine NBL6, and human MRC-5 cells with 20 ng/mL of IFN-γ reduced EHV-1 yield by 1122-, 631-, and 10,000-fold, respectively. However, IFN-γ reduced virus yield by only 2–4-fold in mouse MLE12, mouse L-M, and human MeWo cells compared to those of untreated cells. In luciferase assays with the promoter of the EHV-1 early regulatory EICP0 gene, IFN-γ abrogated trans-activation activity of the IEP by 96% in MH-S cells, but only by 21% in L-M cells. Similar results were obtained in assays with the early regulatory UL5 and IR4 promoter reporter plasmids. IFN-γ treatment reduced IEP protein expression by greater than 99% in MH-S cells, but only by 43% in L-M cells. The expression of IEP and UL5P suppressed by IFN-γ was restored by JAK inhibitor treatment, indicating that the inhibition of EHV-1 replication is mediated by JAK/STAT1 signaling. These results suggest that IFN-γ blocks EHV-1 replication by inhibiting the production of the IEP in a cell line-dependent manner. Affymetrix microarray analyses of IFN-γ-treated MH-S and L-M cells revealed that five antiviral ISGs (MX1, SAMHD1, IFIT2, NAMPT, TREX1, and DDX60) were upregulated 3.2–18.1-fold only in MH-S cells.

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Peptide transport activity of the transporter associated with antigen processing (TAP) is inhibited by an early protein of equine herpesvirus-1

Journal of General Virology ◽

10.1099/vir.0.19563-0 ◽

2004 ◽

Vol 85 (2) ◽

pp. 349-353 ◽

Cited By ~ 35

Author(s):

Aruna P. N. Ambagala ◽

Raju S. Gopinath ◽

S. Srikumaran

Keyword(s):

Antigen Processing ◽

Protein S ◽

Class I ◽

Peptide Transport ◽

Equine Herpesvirus ◽

Equine Herpesvirus 1 ◽

Early Protein ◽

Infected Cells ◽

Herpesvirus 1

Equine herpesvirus-1 (EHV-1) downregulates surface expression of major histocompatibility complex (MHC) class I molecules on infected cells. The objective of this study was to investigate whether EHV-1 interferes with peptide translocation by the transporter associated with antigen processing (TAP) and to identify the proteins responsible. Using an in vitro transport assay, we showed that EHV-1 inhibited transport of peptides by TAP as early as 2 h post-infection (p.i). Complete shutdown of peptide transport was observed by 8 h p.i. Furthermore, pulse–chase experiments revealed that maturation of class I molecules in the endoplasmic reticulum (ER) was delayed in EHV-1-infected cells, which may be due to reduced availability of peptides in the ER as a result of TAP inhibition. Metabolic inhibition studies indicated that an early protein(s) of EHV-1 is responsible for this effect.

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Equine Herpesvirus 1 Gene 12 Can Substitute for vmw65 in the Growth of Herpes Simplex Virus (HSV) Type 1, Allowing the Generation of Optimized Cell Lines for the Propagation of HSV Vectors with Multiple Immediate-Early Gene Defects

Journal of Virology ◽

10.1128/jvi.73.9.7399-7409.1999 ◽

1999 ◽

Vol 73 (9) ◽

pp. 7399-7409 ◽

Cited By ~ 15

Author(s):

S. K. Thomas ◽

C. E. Lilley ◽

D. S. Latchman ◽

R. S. Coffin

Keyword(s):

Gene Expression ◽

Herpes Simplex Virus ◽

Herpes Simplex ◽

Cell Lines ◽

Gene Deletion ◽

Immediate Early ◽

Equine Herpesvirus ◽

Equine Herpesvirus 1 ◽

Herpesvirus 1 ◽

Simplex Virus

ABSTRACT Herpes simplex virus (HSV) has often been suggested for development as a vector, particularly for the nervous system. Considerable evidence has shown that for use of HSV as a vector, immediate-early (IE) gene expression must be minimized or abolished, otherwise such vectors are likely to be highly cytotoxic. Mutations of vmw65 which abolish IE promoter transactivating activity may also be included to reduce IE gene expression generally. However, when vmw65 mutations are combined with an IE gene deletion, such viruses are hard to propagate, even on cells which otherwise complement the IE gene deletion effectively. We have found that vmw65 mutants can be effectively grown on cell lines expressing equine herpesvirus 1 gene 12, a non-HSV homologue of vmw65 with little sequence similarity to its HSV counterpart. This prevents repair by homologous recombination of vmw65 mutations in the virus, which would occur if mutations were complemented by vmw65 itself. The gene 12 protein is not packaged into HSV virions, which is important if viruses grown on such cells are to be used as vectors. These results not only further strengthen the evidence for direct functional homology between and similar modes of action of the two proteins but have allowed the generation of gene 12-containing cell lines in which ICP4 and ICP27 expression is induced by virus infection (probably by ICP0) and which give efficient growth of viruses deficient in ICP27, ICP4, and vmw65 (the viruses also have ICP34.5/ORFP deleted). Efficient growth of such viruses has not previously been possible. As these viruses are highly deficient in IE gene expression generally, such virus-cell line combinations may provide an alternative to HSV vectors with deletions of all four of the regulatory IE genes which, for optimal growth, require cell lines containing all four IE genes but which are hard to generate due to the intrinsic cytotoxicity of each of the proteins.

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