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PMID: 7666507 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't

Functional interaction between the human cytomegalovirus 86-kilodalton IE2 protein and the cellular transcription factor CREB.

Journal of virology ·Vol. 69 ·No. 10 ·1995-10-00 ·Pages 6030-7

Lang D, Gebert S, Arlt H, Stamminger T

Abstract

The 86-kDa IE2 protein (IE86) of human cytomegalovirus (HCMV) has been described as a promiscuous transactivator of viral, as well as cellular, gene expression. Investigation of the mechanism used by IE86 to activate gene expression from the early UL112/113 promoter of HCMV revealed the existence of three binding sites for IE86 located between nucleotides -290 and -120 relative to the transcriptional start site (H. Arlt, D. Lang, S. Gebert, and T. Stamminger, J. Virol. 68:4117-4125, 1994). As shown previously, deletion of these target sites resulted in a reduction of IE86-mediated transactivation by approximately 70%. The remaining promoter, however, could still be stimulated about 40-fold, indicating the presence of an additional responsive element within these sequences. Here, we provide evidence that a binding site for the cellular transcription factor CREB can also act as a target for IE86 transactivation. By DNase I protection analysis, a binding sequence for CREB could be detected between nucleotides -78 and -56 within the respective promoter region. After in vitro mutagenesis of this CREB-binding site within the context of the entire UL112/113 promoter, a marked reduction in transactivation levels was evident. Moreover, when individual CREB-binding sites were positioned upstream of a minimal, TATA box-containing UL112/113 promoter, they were able to confer strong IE86 responsiveness, whereas a mutated sequence did not exert any effect. In far Western blot and pull-down experiments, a direct interaction of IE86 with the cellular transcription factor CREB could be observed. The in vivo relevance of this in vitro interaction was confirmed by using various GAL4 fusion proteins in the presence or absence of IE86 which revealed a strong activation only in the presence of both a GAL4-CREB fusion and IE86. This shows that at least one specific member of the ATF/CREB family of transcription factors is involved in mediating transactivation by the HCMV IE86 protein.

MeSH Terms
Base Sequence Consensus Sequence Cyclic AMP Response Element-Binding Protein/metabolism Cytomegalovirus/metabolism Gene Expression Regulation, Viral Globins/biosynthesis Histidine Humans Immediate-Early Proteins/metabolism Membrane Glycoproteins Molecular Sequence Data Mutagenesis, Site-Directed Oligodeoxyribonucleotides Plasmids Promoter Regions, Genetic RNA, Viral/analysis,biosynthesis Recombinant Fusion Proteins/biosynthesis,metabolism Sequence Deletion Sequence Tagged Sites Trans-Activators/metabolism Transcriptional Activation Viral Envelope Proteins Viral Proteins
Chemicals
Cyclic AMP Response Element-Binding Protein IE2 protein, Cytomegalovirus Immediate-Early Proteins Membrane Glycoproteins Oligodeoxyribonucleotides RNA, Viral Recombinant Fusion Proteins Trans-Activators UL115 protein, Human herpesvirus 5 Viral Envelope Proteins Viral Proteins glycoprotein H, Cytomegalovirus glycoprotein H, Human cytomegalovirus glycoprotein O, cytomegalovirus Histidine Globins
Authors & Affiliations
4 authors, click to expand affiliations / ORCID
Lang D
Institut für Klinische und Molekulare Virologie, Universität Erlangen-Nürnberg, Germany.
Gebert S
Arlt H
Stamminger T
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Article Info
Journal
Journal of virology
Abbr.
J Virol
ISSN
0022-538X
DOI
10.1128/JVI.69.10.6030-6037.1995
Published
1995-10-00
Pages
6030-7
Language
English
Region
United States
NLM ID
0113724
PMCID
PMC189499
Subset
IM
External Links
PubMed source DOI: 10.1128/JVI.69.10.6030-6037.1995 PMC Full Text View journal details
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