Braddock M, Powell R, Blanchard A D, Kingsman A J, Kingsman S M
Department of Biochemistry, Oxford University, England.
FASEB J. 1993 Jan;7(1):214-22. doi: 10.1096/fasebj.7.1.8422967.
Human immunodeficiency virus (HIV-1) gene expression is activated by the viral TAT protein that interacts with an RNA sequence, TAR, located at the 5' end of all viral mRNAs. TAT functions primarily as a transcriptional activator in mammalian cells. However, in Xenopus oocytes TAT functions primarily as a translational activator. TAR is an RNA structure comprising a partially base-paired stem, a tripyrimidine bulge in the upper stem, and an unpaired six-nucleotide loop. In vitro, TAT binds directly to the bulge with no requirement for the loop. In vivo, however, mutations in the loop abolish TAT activation of transcription and translation, implying a requirement for TAR-binding cellular factors. We now provide genetic evidence for the presence of two TAR-specific cellular factors in Xenopus oocytes. These factors display independent and mutually exclusive interactions with either the loop or the bulge region of TAR. Furthermore, by using in vivo RNA competition assays we show that the cellular factors regulate the accessibility of the TAT binding site. The fact that Xenopus oocytes contain factors that specifically interact with a human viral RNA sequence might indicate that the TAT/TAR interaction is subverting a conserved pathway in the cell.
人类免疫缺陷病毒（HIV-1）基因表达由病毒TAT蛋白激活，该蛋白与位于所有病毒mRNA 5'端的RNA序列TAR相互作用。TAT在哺乳动物细胞中主要作为转录激活因子发挥作用。然而，在非洲爪蟾卵母细胞中，TAT主要作为翻译激活因子发挥作用。TAR是一种RNA结构，由部分碱基配对的茎、上部茎中的三嘧啶凸起和一个未配对的六核苷酸环组成。在体外，TAT直接与凸起结合，不需要环。然而，在体内，环中的突变消除了TAT对转录和翻译的激活，这意味着需要与TAR结合的细胞因子。我们现在提供了非洲爪蟾卵母细胞中存在两种TAR特异性细胞因子的遗传学证据。这些因子与TAR的环或凸起区域表现出独立且相互排斥的相互作用。此外，通过体内RNA竞争试验，我们表明细胞因子调节TAT结合位点的可及性。非洲爪蟾卵母细胞含有与人类病毒RNA序列特异性相互作用的因子这一事实可能表明，TAT/TAR相互作用正在破坏细胞中的一条保守途径。
