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PMID: 8164693 Published · ppublish English Comparative Study Journal Article

Transcriptional activation by herpes simplex virus type 1 VP16 in vitro and its inhibition by oligopeptides.

Molecular and cellular biology ·Vol. 14 ·No. 5 ·1994-05-00 ·Pages 3484-93

Wu TJ, Monokian G, Mark DF, Wobbe CR

Abstract

VP16 is a herpes simplex virus (HSV)-encoded transcriptional activator protein that is essential for efficient viral replication and as such may be a target for novel therapeutic agents directed against viral gene expression. We have reconstituted transcriptional activation by VP16 in an in vitro system that is dependent on DNA sequences from HSV immediate-early gene promoters and on protein-protein interactions between VP16 and Oct-1 that are required for VP16 activation in vivo. Activation increased synergistically with the number of TAATGARAT elements (the cis-acting element for VP16 activation in vivo) upstream of the core promoter, and mutations of this element that reduce Oct-1 or VP16 DNA binding reduced transactivation in vitro. A VP16 insertion mutant unable to interact with Oct-1 was inactive, but, surprisingly, a deletion mutant lacking the activation domain was approximately 65% as active as the full-length protein. The activation domains of Oct-1 were necessary for activation in reactions containing the VP16 deletion mutant, and they contributed significantly to activation by full-length VP16. Addition of a GA-rich element present in many HSV immediate-early gene enhancers synergistically stimulated VP16-activated transcription. Finally, oligopeptides that are derived from a region of VP16 thought to contact a cellular factor known as HCF (host cell factor) and that inhibit efficient VP16 binding to the TAATGARAT element also specifically inhibited VP16-activated, but not basal, transcription. Amino acid substitutions in one of these peptides identified three residues that are absolutely required for inhibition and presumably for interaction of VP16 with HCF.

MeSH Terms
Amino Acid Sequence Base Sequence Cell Nucleus/metabolism DNA Primers DNA, Viral/drug effects,metabolism DNA-Binding Proteins/metabolism Herpes Simplex Virus Protein Vmw65/metabolism Herpesvirus 1, Human/drug effects,genetics Host Cell Factor C1 Kinetics Molecular Sequence Data Octamer Transcription Factor-1 Oligodeoxyribonucleotides Oligopeptides/pharmacology Polymerase Chain Reaction Promoter Regions, Genetic Recombinant Fusion Proteins/metabolism Restriction Mapping Structure-Activity Relationship TATA Box Transcription Factors/metabolism Transcription, Genetic Transcriptional Activation/drug effects
Chemicals
DNA Primers DNA, Viral DNA-Binding Proteins Herpes Simplex Virus Protein Vmw65 Host Cell Factor C1 Octamer Transcription Factor-1 Oligodeoxyribonucleotides Oligopeptides Recombinant Fusion Proteins Transcription Factors
Authors & Affiliations
4 authors, click to expand affiliations / ORCID
Wu T J
Department of Microbial Chemotherapeutics and Molecular Genetics, Merck Research Laboratories, Rahway, New Jersey 07063.
Monokian G
Mark D F
Wobbe C R
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Article Info
Journal
Molecular and cellular biology
Abbr.
Mol Cell Biol
ISSN
0270-7306
DOI
10.1128/mcb.14.5.3484-3493.1994
Published
1994-05-00
Pages
3484-93
Language
English
Region
United States
NLM ID
8109087
PMCID
PMC358712
Subset
IM
External Links
PubMed source DOI: 10.1128/mcb.14.5.3484-3493.1994 PMC Full Text View journal details
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