title: Inducible transcriptional activation of the human immunodeficiency virus long terminal repeat by protein kinase inhibitors.
abstract: The protein kinase inhibitor 2-aminopurine (2-AP) greatly stimulated expression in human promonocytes-macrophages of plasmid constructs carrying various reporter genes (chloramphenicol acetyltransferase, lacZ, firefly luciferase [luc], and Salmonella typhimurium histidinol dehydrogenase [his]) driven by the human immunodeficiency virus type 1 (HIV-1) long terminal repeat. Adenine, adenosine, and caffeine were also effective inducers, but other purine or pyrimidine derivatives were ineffective. Experiments with mutant derivatives of the HIV-1 long terminal repeat revealed no specific eukaryotic promoter elements necessary for 2-AP induction but indicated the need for some minimum combination of such elements. Induction of HIV-1-directed gene expression appeared not to require action of the transcription factor NF-kappa B. The mechanism of induction was investigated by using the luc and his genes linked to the HIV-1 long terminal repeat. 2-AP induced marked, steady rises in mRNA accumulation from both transfected and chromosomally integrated HIV-1 constructs but no increases from an endogenous gene encoding gamma-actin or glucose 6-phosphate dehydrogenase. Thus, induction is selective and not an artifact induced by transfecting DNA into cells. In run-on transcription experiments, the rates of transcription initiation of both transfected and integrated copies of the his gene increased about sixfold in cells treated with 2-AP. Thus, while increased initiation accounted for a portion of 2-AP induction, it could not cause the far greater increase in steady-state mRNA levels. 2-AP induction did not change mRNA decay rates and differed from the phorbol ester (phorbol myristate acetate)-induced activation of the protein kinase C-NF-kappa B pathway in its time course and in its requirement for new protein synthesis. Gel retardation assays showed that unlike phorbol myristate acetate induction, 2-AP induction is enhancer independent. Whereas many previous studies have implicated the activation of various protein kinases in gene induction, we here describe a mechanism of gene activation that appears to involve protein kinase inhibition as a component of the induction response.
authors.affiliation: Department of Cell Biology, Albert Einstein College of Medicine, Bronx, New York 10461.
authors.forename: F L
authors.initials: FL
authors.lastname: Brown
authors.forename: E
authors.initials: E
authors.lastname: Tahaoglu
authors.forename: G J
authors.initials: GJ
authors.lastname: Graham
authors.forename: J J
authors.initials: JJ
authors.lastname: Maio
mesh_terms: D015075:2-Aminopurine / Q000494:pharmacology*; D001483:Base Sequence; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism; D003513:Cycloheximide / Q000494:pharmacology; D003609:Dactinomycin / Q000494:pharmacology; D015967:Gene Expression Regulation, Viral / Q000187:drug effects*; D005814:Genes, Viral; D016325:HIV Long Terminal Repeat / Q000235:genetics*; D015497:HIV-1 / Q000235:genetics*; D006801:Humans; D066298:In Vitro Techniques; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009838:Oligodeoxyribonucleotides / Q000737:chemistry; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000235:genetics; D013329:Structure-Activity Relationship; D014158:Transcription, Genetic; D015533:Transcriptional Activation; D014162:Transfection; D015678:Viral Structural Proteins / Q000235:genetics
chemical_list: D009838:Oligodeoxyribonucleotides; D012333:RNA, Messenger; D015678:Viral Structural Proteins; D003609:Dactinomycin; D015075:2-Aminopurine; D003513:Cycloheximide