Delineating minimal protein domains and promoter elements for transcriptional activation by lentivirus Tat proteins.
Lentivirus Tat proteins comprise a novel class of RNA-binding transcriptional activators that are essential for viral replication. In this study, we performed a series of protein fusion experiments to delineate the minimal protein domains and promoter elements required for Tat action. We show that a 15-amino-acid region of equine infectious anemia virus (EIAV) Tat protein, when fused to the GAL4 or LexA DNA binding domain, can activate transcription in appropriate promoter contexts. In the natural human immunodeficiency virus type 1 long terminal repeat, activation by Tat is dependent on multiple binding sites for the cellular transcription factor SP1. We delineate a 114-amino-acid region of the SP1 glutamine-rich activation domain that when fused to the GAL4 DNA binding domain can support transcription activation by Tat. Using these Tat and SP1 derivatives, we show that Tat activation can be reconstructed on a completely synthetic promoter lacking all cis-acting elements unique to the human immunodeficiency virus long terminal repeat. Our results indicate that lentivirus Tat proteins have essential properties of typical cellular transcriptional activators and define useful reagents for studying the detailed mechanism of Tat action.
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Howard Hughes Medical Institute, Program in Molecular Medicine, University of Massachusetts Medical Center 01605.
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D000818:Animals; D006224:Cricetinae; D004268:DNA-Binding Proteins; D005656:Fungal Proteins / Q000378:metabolism; D015696:Gene Products, tat / Q000378:metabolism*; D016325:HIV Long Terminal Repeat; D015497:HIV-1 / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D004860:Infectious Anemia Virus, Equine / Q000235:genetics / Q000378:metabolism; D016086:Lentivirus / Q000235:genetics* / Q000378:metabolism; D011401:Promoter Regions, Genetic; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors / Q000378:metabolism; D015533:Transcriptional Activation; D054322:tat Gene Products, Human Immunodeficiency Virus
D004268:DNA-Binding Proteins; D005656:Fungal Proteins; C052130:GAL4 protein, S cerevisiae; D015696:Gene Products, tat; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors; D054322:tat Gene Products, Human Immunodeficiency Virus