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Orientation-specific cis complementation by bulge- and loop-mutated human immunodeficiency virus type 1 TAR RNAs.

M Braddock·R Powell·J Sutton·A J Kingsman·S M Kingsman
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Abstract

Tat activates human immunodeficiency type 1 gene expression by binding to TAR RNA. TAR comprises a partially base paired stem and hexanucleotide loop with a tripyrimidine bulge in the upper stem. In vitro, Tat binds to the bulge and upper stem, with no requirement for the loop. However, in vivo, loop sequences are critical for activation, implying that a loop binding cellular factor may be involved in the activation pathway. Given that activation appears to be a two-component system comprising a Tat-bulge interaction and a cellular factor-loop interaction, we considered that it might be possible to spatially separate the two components and retain activation. We have constructed a series of double TAR elements comprising various combinations of mutated TAR structures. Defective TARs with nucleotide substitutions in either the bulge or the loop complemented each other to give wild-type activation. However, the complementation was orientation specific, requiring the intact Tat binding site to reside on the 5'-proximal TAR. These data suggest that provided the wild-type orientation of the bulge and loop elements is retained, there is no requirement for them to coexist on the same TAR structure.

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Bacterial ProteinsBase SequenceBinding SitesChemoreceptor CellsEscherichia coli ProteinsGene Products, tatGenetic Complementation TestHIV Long Terminal RepeatHIV-1HeLa CellsHumansMembrane ProteinsModels, StructuralMolecular Sequence DataMutagenesis, InsertionalMutagenesis, Site-DirectedNucleic Acid ConformationPlasmidsPromoter Regions, GeneticRNA, ViralReceptors, Cell Surfacetat Gene Products, Human Immunodeficiency VirusJournal ArticleResearch Support, Non-U.S. Gov't

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M Braddock, R Powell, J Sutton, A J Kingsman, S M Kingsman. 1994. Orientation-specific cis complementation by bulge- and loop-mutated human immunodeficiency virus type 1 TAR RNAs.. https://doi.org/10.1128/jvi.68.12.8396-8400.1994

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