title: Identification of a novel enhancer element mediating calcium-dependent induction of gene expression in response to either epidermal growth factor or activation of protein kinase C.
abstract: The VL30 family of defective murine retroviruses consists of 100 to 200 members, of which fewer than 5% appear to be transcriptionally active. A genomic clone of the transcriptionally active VL30 element RVL-3 was identified and sequenced. Genetic analysis indicated that a triple-repeat sequence within the RVL-3 long terminal repeat is capable of functioning as an inducible enhancer element responding to a variety of agonists. In Rat-1 fibroblasts, the ability of the RVL-3 enhancer to mediate induction of gene expression from a heterologous promoter in response to either epidermal growth factor (EGF) or phorbol ester treatment required coelevation of intracellular calcium. Two CArG boxes present in the triple-repeat sequence appeared to exert a negative effect on gene expression, as mutation of these sequences elevated the basal level of expression observed without altering the fold induction in response to either EGF or protein kinase C activation. In the presence of these CArG elements, mutation of AP-1-like sites adjacent to the CArG elements significantly inhibited the ability of either EGF or phorbol esters to induce gene expression. The effect of mutating these AP-1-like sites was relieved by simultaneous mutation of the CArG sites, indicating that interactions among these sites modulate RVL-3 expression. Mutational analysis and gel mobility shift experiments have identified a third sequence within the VL30 triple-repeat element that is required for the induction of gene expression and serves as a binding site for nuclear proteins. Sequence comparisons indicate that this enhancer element has not been described previously.
authors.affiliation: Centre de Biochimie, Université de Nice, 06034 France.
authors.forename: P
authors.initials: P
authors.lastname: Lenormand
authors.forename: D
authors.initials: D
authors.lastname: Pribnow
authors.forename: K D
authors.initials: KD
authors.lastname: Rodland
authors.forename: B E
authors.initials: BE
authors.lastname: Magun
mesh_terms: D000818:Animals; D001483:Base Sequence; D002118:Calcium / Q000502:physiology*; D004252:DNA Mutational Analysis; D004279:DNA, Viral / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism; D003673:Defective Viruses / Q000235:genetics; D004742:Enhancer Elements, Genetic; D004789:Enzyme Activation; D004815:Epidermal Growth Factor / Q000494:pharmacology*; D015967:Gene Expression Regulation, Viral / Q000187:drug effects; D066298:In Vitro Techniques; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000378:metabolism; D011401:Promoter Regions, Genetic; D011493:Protein Kinase C / Q000502:physiology*; D051381:Rats; D012091:Repetitive Sequences, Nucleic Acid; D012190:Retroviridae / Q000235:genetics*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014158:Transcription, Genetic
chemical_list: D004279:DNA, Viral; D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; D004815:Epidermal Growth Factor; D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate; D002118:Calcium