Evolution of the herpes thymidine kinase: identification and comparison of the equine herpesvirus 1 thymidine kinase gene reveals similarity to a cell-encoded thymidylate kinase

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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Yeast gene CDC8 encodes thymidylate kinase and is complemented by herpes thymidine kinase gene TK.

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.81.18.5821 ◽

1984 ◽

Vol 81 (18) ◽

pp. 5821-5825 ◽

Cited By ~ 47

Author(s):

R. A. Sclafani ◽

W. L. Fangman

Keyword(s):

Thymidine Kinase ◽

Thymidine Kinase Gene ◽

Yeast Gene ◽

Kinase Gene ◽

Thymidylate Kinase

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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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