Cai R, Carpick B, Chun R F, Jeang K T, Williams B R
Lerner Research Institute, Cleveland Clinic Foundation, 9500 Euclid Avenue, Cleveland, Ohio 44195, USA.
Arch Biochem Biophys. 2000 Jan 15;373(2):361-7. doi: 10.1006/abbi.1999.1583.
Replication of the human immunodeficiency virus type 1 (HIV-1) is inhibited by interferons (IFNs), in part through activity of the IFN-inducible protein kinase PKR. To escape this antiviral effect, HIV-1 has developed strategies for blocking PKR function. We have previously shown that the HIV-1 Tat protein can associate with PKR in vitro and in vivo and inhibit PKR activity. Here we present evidence that Tat can inhibit PKR activity by both RNA-dependent and RNA-independent mechanisms. Tat inhibited PKR activation by the non-RNA activator heparin, and also suppressed PKR basal level autophosphorylation in the absence of RNA. However, when Tat and dsRNA were preincubated, the amount of Tat required to inhibit PKR activation by dsRNA depended on the dsRNA concentration. In addition to its function in vitro, Tat can also reverse translation inhibition mediated by PKR in COS cells. The Tat amino acid sequence required for interaction with PKR was mapped to residues 40-58, overlapping the hydrophobic core and basic region of HIV-1 Tat. Alignment of amino acid sequences of Tat and eIF-2alpha indicates similarity between the Tat-PKR binding region and the residues around the eIF-2alpha phosphorylation site, suggesting that Tat and eIF-2alpha may bind to the same site on PKR.
1型人类免疫缺陷病毒（HIV-1）的复制受到干扰素（IFN）的抑制，部分原因是IFN诱导的蛋白激酶PKR的活性。为了逃避这种抗病毒作用，HIV-1已开发出阻断PKR功能的策略。我们之前已经表明，HIV-1 Tat蛋白在体外和体内都能与PKR结合并抑制PKR活性。在此我们提供证据表明，Tat可以通过RNA依赖性和RNA非依赖性机制抑制PKR活性。Tat抑制了非RNA激活剂肝素对PKR的激活作用，并且在没有RNA的情况下也抑制了PKR的基础水平自磷酸化。然而，当Tat与双链RNA（dsRNA）预孵育时，抑制dsRNA激活PKR所需的Tat量取决于dsRNA的浓度。除了其在体外的功能外，Tat还可以逆转COS细胞中由PKR介导的翻译抑制。与PKR相互作用所需的Tat氨基酸序列被定位到40-58位残基，与HIV-1 Tat的疏水核心和碱性区域重叠。Tat和真核起始因子2α（eIF-2α）的氨基酸序列比对表明Tat-PKR结合区域与eIF-2α磷酸化位点周围的残基相似，这表明Tat和eIF-2α可能结合到PKR上的同一位点。
