The IE2 regulatory protein of human cytomegalovirus induces expression of the human transforming growth factor β1 gene through an Egr-1 binding site

Young Do Yoo, Chuang Jiun Chiou, Kyeong Sook Choi, Youngsuk Yi, Susan Michelson, Sunyoung Kim, Gary Selwyn Hayward, Seong Jin Kim

Research output: Contribution to journalArticle

Abstract

Increases in transforming growth factor β1 (TGF-β1) mRNA and biological activity in the early phase of human cytomegalovirus (CMV) infection in fibroblasts are paralleled by increased TGF-β1-chloramphenicol acetyltransferase (CAT) reporter gene activity. To determine how CMV infection transactivates the TGF-β1 promoter, we examined the effects of the cotransfected IE2 regulatory protein of human CMV on 5'-deleted TGF-β1 promoter-CAT reporter genes in transient DNA transfection assays. Two upstream TGF-β1 promoter regions each containing an Egr-1 consensus site were shown to be important for IE2-induced transactivation in a cell type that displayed greatly reduced nonspecific activity. Furthermore, transfer of an Egr-1 site from between positions -125 and -98, but not point mutant versions of this site, to a heterologous promoter also conveyed IE2 responsiveness. Addition of an IE2 expression vector or use of the U373 A45 astrocytoma cell line expressing IE2 also produced synergistic stimulation of GAL4-Egr-1-mediated activation of a target promoter containing GAL4 binding sites. The 80-kDa IE2 protein present in A45 cells proved to selectively bind to glutathione S-transferase (GST)-Egr-1 heads. The results of in vitro protein binding assays also revealed that an intact in vitro-translated IE2 protein bound directly to the GST-Egr-1 fusion protein through the zinc finger domain of the Egr-1 protein and that this binding activity was abolished by deletion of parts of the zinc finger DNA-binding domain. Similarly, the Egr-1 protein was found to associate preferentially with a small region within the C-terminal half of the IE2 protein adjacent to the DNA-binding and dimerization domains that are important for both transactivation and downregulation. We conclude from these observations that IE2 may regulate transcription of the TGF-β1 gene as well as other potential cellular targets by virtue of its ability to interact with the Egr-1 DNA- binding protein.

Original languageEnglish (US)
Pages (from-to)7062-7070
Number of pages9
JournalJournal of Virology
Volume70
Issue number10
StatePublished - Oct 1996

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Human herpesvirus 5
transforming growth factors
regulatory proteins
Transforming Growth Factors
Cytomegalovirus
binding sites
Binding Sites
promoter regions
chloramphenicol acetyltransferase
Genes
Proteins
Chloramphenicol O-Acetyltransferase
genes
proteins
zinc finger motif
Zinc Fingers
Cytomegalovirus Infections
transcriptional activation
Glutathione Transferase
Reporter Genes

ASJC Scopus subject areas

  • Immunology

Cite this

Yoo, Y. D., Chiou, C. J., Choi, K. S., Yi, Y., Michelson, S., Kim, S., ... Kim, S. J. (1996). The IE2 regulatory protein of human cytomegalovirus induces expression of the human transforming growth factor β1 gene through an Egr-1 binding site. Journal of Virology, 70(10), 7062-7070.

The IE2 regulatory protein of human cytomegalovirus induces expression of the human transforming growth factor β1 gene through an Egr-1 binding site. / Yoo, Young Do; Chiou, Chuang Jiun; Choi, Kyeong Sook; Yi, Youngsuk; Michelson, Susan; Kim, Sunyoung; Hayward, Gary Selwyn; Kim, Seong Jin.

In: Journal of Virology, Vol. 70, No. 10, 10.1996, p. 7062-7070.

Research output: Contribution to journalArticle

Yoo, Young Do ; Chiou, Chuang Jiun ; Choi, Kyeong Sook ; Yi, Youngsuk ; Michelson, Susan ; Kim, Sunyoung ; Hayward, Gary Selwyn ; Kim, Seong Jin. / The IE2 regulatory protein of human cytomegalovirus induces expression of the human transforming growth factor β1 gene through an Egr-1 binding site. In: Journal of Virology. 1996 ; Vol. 70, No. 10. pp. 7062-7070.
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abstract = "Increases in transforming growth factor β1 (TGF-β1) mRNA and biological activity in the early phase of human cytomegalovirus (CMV) infection in fibroblasts are paralleled by increased TGF-β1-chloramphenicol acetyltransferase (CAT) reporter gene activity. To determine how CMV infection transactivates the TGF-β1 promoter, we examined the effects of the cotransfected IE2 regulatory protein of human CMV on 5'-deleted TGF-β1 promoter-CAT reporter genes in transient DNA transfection assays. Two upstream TGF-β1 promoter regions each containing an Egr-1 consensus site were shown to be important for IE2-induced transactivation in a cell type that displayed greatly reduced nonspecific activity. Furthermore, transfer of an Egr-1 site from between positions -125 and -98, but not point mutant versions of this site, to a heterologous promoter also conveyed IE2 responsiveness. Addition of an IE2 expression vector or use of the U373 A45 astrocytoma cell line expressing IE2 also produced synergistic stimulation of GAL4-Egr-1-mediated activation of a target promoter containing GAL4 binding sites. The 80-kDa IE2 protein present in A45 cells proved to selectively bind to glutathione S-transferase (GST)-Egr-1 heads. The results of in vitro protein binding assays also revealed that an intact in vitro-translated IE2 protein bound directly to the GST-Egr-1 fusion protein through the zinc finger domain of the Egr-1 protein and that this binding activity was abolished by deletion of parts of the zinc finger DNA-binding domain. Similarly, the Egr-1 protein was found to associate preferentially with a small region within the C-terminal half of the IE2 protein adjacent to the DNA-binding and dimerization domains that are important for both transactivation and downregulation. We conclude from these observations that IE2 may regulate transcription of the TGF-β1 gene as well as other potential cellular targets by virtue of its ability to interact with the Egr-1 DNA- binding protein.",
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