ORF36 protein kinase of Kaposi's sarcoma herpesvirus activates the c-Jun N-terminal kinase signaling pathway

M. Sabry Hamza, Richard A. Reyes, Yoshihiro Izumiya, Ronald Wisdom, Hsing Jien Kung, Paul A. Luciw

研究成果: 雜誌貢獻文章

52 引文 (Scopus)

摘要

α, β, and γ-Herpesviruses encode putative viral protein kinases. The herpes simplex virus UL13, varicellazoster virus ORF47, and Epstein-Barr virus BGLF4 genes all show protein kinase domains in their protein sequences. Mutational analysis of these herpesviruses demonstrated that the viral kinase is important for optimal virus growth. Previous studies have shown that ORF36 of Kaposi's sarcoma herpesvirus (KSHV) has protein kinase activity and is autophosphorylated on serine. The gene for ORF36 is expressed during lytic growth of the virus and has been classified as a late gene. Inspection of the ORF36 sequence indicated potential motifs that could be involved in activation of cellular transcription factors. To analyze the function of ORF36, the cDNA for this viral gene was tagged with the FLAG epitope and inserted into an expression vector for mammalian cells. Transfection experiments in 293T and SLK cells demonstrated that expression of ORF36 resulted in phosphorylation of the c-Jun N-terminal kinase. Autophosphorylation of ORF36 is important for JNK activation because a mutation in the predicted catalytic domain of ORF36 blocked its ability to phosphorylate JNK. Western blot analysis, using phosphospecific antibodies, revealed that mitogen-activated kinases MEK4 and MKK7 were phosphorylated by ORF36 but not by the kinase-negative mutant. Binding experiments in transfected cells also demonstrated that both the wild type and kinase-negative mutant of ORF36 form a complex with JNK, MKK4, and MKK7. In addition, using a tetracycline-inducible Rta BCBL-1 cell line (TREx BCBL1-Rta), JNK was phosphorylated during lytic replication, and inhibition of JNK activation blocked late viral gene expression but not early viral gene expression. In summary, these studies demonstrate that KSHV ORF36 activates the JNK pathway; thus this cell signaling pathway may function in the KSHV life cycle by regulating viral and/or cellular transcription.
原文英語
頁(從 - 到)38325-38330
頁數6
期刊Journal of Biological Chemistry
279
發行號37
DOIs
出版狀態已發佈 - 九月 10 2004
對外發佈Yes

指紋

JNK Mitogen-Activated Protein Kinases
Herpesviridae
Viruses
Viral Genes
Phosphotransferases
Genes
Kaposi's Sarcoma
Protein Kinases
Chemical activation
Gene expression
Phospho-Specific Antibodies
Gene Expression
Cells
MAP Kinase Signaling System
HEK293 Cells
Cell signaling
Viral Proteins
Simplexvirus
Growth
Tetracycline

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

引用此文

ORF36 protein kinase of Kaposi's sarcoma herpesvirus activates the c-Jun N-terminal kinase signaling pathway. / Hamza, M. Sabry; Reyes, Richard A.; Izumiya, Yoshihiro; Wisdom, Ronald; Kung, Hsing Jien; Luciw, Paul A.

於: Journal of Biological Chemistry, 卷 279, 編號 37, 10.09.2004, p. 38325-38330.

研究成果: 雜誌貢獻文章

Hamza, M. Sabry ; Reyes, Richard A. ; Izumiya, Yoshihiro ; Wisdom, Ronald ; Kung, Hsing Jien ; Luciw, Paul A. / ORF36 protein kinase of Kaposi's sarcoma herpesvirus activates the c-Jun N-terminal kinase signaling pathway. 於: Journal of Biological Chemistry. 2004 ; 卷 279, 編號 37. 頁 38325-38330.
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abstract = "α, β, and γ-Herpesviruses encode putative viral protein kinases. The herpes simplex virus UL13, varicellazoster virus ORF47, and Epstein-Barr virus BGLF4 genes all show protein kinase domains in their protein sequences. Mutational analysis of these herpesviruses demonstrated that the viral kinase is important for optimal virus growth. Previous studies have shown that ORF36 of Kaposi's sarcoma herpesvirus (KSHV) has protein kinase activity and is autophosphorylated on serine. The gene for ORF36 is expressed during lytic growth of the virus and has been classified as a late gene. Inspection of the ORF36 sequence indicated potential motifs that could be involved in activation of cellular transcription factors. To analyze the function of ORF36, the cDNA for this viral gene was tagged with the FLAG epitope and inserted into an expression vector for mammalian cells. Transfection experiments in 293T and SLK cells demonstrated that expression of ORF36 resulted in phosphorylation of the c-Jun N-terminal kinase. Autophosphorylation of ORF36 is important for JNK activation because a mutation in the predicted catalytic domain of ORF36 blocked its ability to phosphorylate JNK. Western blot analysis, using phosphospecific antibodies, revealed that mitogen-activated kinases MEK4 and MKK7 were phosphorylated by ORF36 but not by the kinase-negative mutant. Binding experiments in transfected cells also demonstrated that both the wild type and kinase-negative mutant of ORF36 form a complex with JNK, MKK4, and MKK7. In addition, using a tetracycline-inducible Rta BCBL-1 cell line (TREx BCBL1-Rta), JNK was phosphorylated during lytic replication, and inhibition of JNK activation blocked late viral gene expression but not early viral gene expression. In summary, these studies demonstrate that KSHV ORF36 activates the JNK pathway; thus this cell signaling pathway may function in the KSHV life cycle by regulating viral and/or cellular transcription.",
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T1 - ORF36 protein kinase of Kaposi's sarcoma herpesvirus activates the c-Jun N-terminal kinase signaling pathway

AU - Hamza, M. Sabry

AU - Reyes, Richard A.

AU - Izumiya, Yoshihiro

AU - Wisdom, Ronald

AU - Kung, Hsing Jien

AU - Luciw, Paul A.

PY - 2004/9/10

Y1 - 2004/9/10

N2 - α, β, and γ-Herpesviruses encode putative viral protein kinases. The herpes simplex virus UL13, varicellazoster virus ORF47, and Epstein-Barr virus BGLF4 genes all show protein kinase domains in their protein sequences. Mutational analysis of these herpesviruses demonstrated that the viral kinase is important for optimal virus growth. Previous studies have shown that ORF36 of Kaposi's sarcoma herpesvirus (KSHV) has protein kinase activity and is autophosphorylated on serine. The gene for ORF36 is expressed during lytic growth of the virus and has been classified as a late gene. Inspection of the ORF36 sequence indicated potential motifs that could be involved in activation of cellular transcription factors. To analyze the function of ORF36, the cDNA for this viral gene was tagged with the FLAG epitope and inserted into an expression vector for mammalian cells. Transfection experiments in 293T and SLK cells demonstrated that expression of ORF36 resulted in phosphorylation of the c-Jun N-terminal kinase. Autophosphorylation of ORF36 is important for JNK activation because a mutation in the predicted catalytic domain of ORF36 blocked its ability to phosphorylate JNK. Western blot analysis, using phosphospecific antibodies, revealed that mitogen-activated kinases MEK4 and MKK7 were phosphorylated by ORF36 but not by the kinase-negative mutant. Binding experiments in transfected cells also demonstrated that both the wild type and kinase-negative mutant of ORF36 form a complex with JNK, MKK4, and MKK7. In addition, using a tetracycline-inducible Rta BCBL-1 cell line (TREx BCBL1-Rta), JNK was phosphorylated during lytic replication, and inhibition of JNK activation blocked late viral gene expression but not early viral gene expression. In summary, these studies demonstrate that KSHV ORF36 activates the JNK pathway; thus this cell signaling pathway may function in the KSHV life cycle by regulating viral and/or cellular transcription.

AB - α, β, and γ-Herpesviruses encode putative viral protein kinases. The herpes simplex virus UL13, varicellazoster virus ORF47, and Epstein-Barr virus BGLF4 genes all show protein kinase domains in their protein sequences. Mutational analysis of these herpesviruses demonstrated that the viral kinase is important for optimal virus growth. Previous studies have shown that ORF36 of Kaposi's sarcoma herpesvirus (KSHV) has protein kinase activity and is autophosphorylated on serine. The gene for ORF36 is expressed during lytic growth of the virus and has been classified as a late gene. Inspection of the ORF36 sequence indicated potential motifs that could be involved in activation of cellular transcription factors. To analyze the function of ORF36, the cDNA for this viral gene was tagged with the FLAG epitope and inserted into an expression vector for mammalian cells. Transfection experiments in 293T and SLK cells demonstrated that expression of ORF36 resulted in phosphorylation of the c-Jun N-terminal kinase. Autophosphorylation of ORF36 is important for JNK activation because a mutation in the predicted catalytic domain of ORF36 blocked its ability to phosphorylate JNK. Western blot analysis, using phosphospecific antibodies, revealed that mitogen-activated kinases MEK4 and MKK7 were phosphorylated by ORF36 but not by the kinase-negative mutant. Binding experiments in transfected cells also demonstrated that both the wild type and kinase-negative mutant of ORF36 form a complex with JNK, MKK4, and MKK7. In addition, using a tetracycline-inducible Rta BCBL-1 cell line (TREx BCBL1-Rta), JNK was phosphorylated during lytic replication, and inhibition of JNK activation blocked late viral gene expression but not early viral gene expression. In summary, these studies demonstrate that KSHV ORF36 activates the JNK pathway; thus this cell signaling pathway may function in the KSHV life cycle by regulating viral and/or cellular transcription.

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