Fujii R, Okamoto M, Aratani S, Oishi T, Ohshima T, Taira K, Baba M, Fukamizu A, Nakajima T
Institute of Applied Biochemistry, University of Tsukuba, 1-1-1 Tennoudai, Tsukuba, Ibaraki 305-8577, Japan.
J Biol Chem. 2001 Feb 23;276(8):5445-51. doi: 10.1074/jbc.M006892200. Epub 2000 Nov 28.
RNA helicase A (RHA) has two double-stranded (ds) RNA-binding domains (dsRBD1 and dsRBD2). These domains are conserved with the cis-acting transactivation response element (TAR)-binding protein (TRBP) and dsRNA-activated protein kinase (PKR). TRBP and PKR are involved in the regulation of HIV-1 gene expression through their binding to TAR RNA. This study shows that RHA also plays an important role in TAR-mediated HIV-1 gene expression. Wild-type RHA preferably bound to TAR RNA in vitro and in vivo. Overexpression of wild type RHA strongly enhanced viral mRNA synthesis and virion production as well as HIV-1 long terminal repeat-directed reporter (luciferase) gene expression. Substitution of lysine for glutamate at residue 236 in dsRBD2 (RHA(K236E)) reduced its affinity for TAR RNA and impaired HIV-1 transcriptional activity. These results indicate that TAR RNA is a preferred target of RHA dsRBDs and that RHA enhances HIV-1 transcription in vivo in part through the TAR-binding of RHA.
RNA解旋酶A（RHA）有两个双链（ds）RNA结合结构域（dsRBD1和dsRBD2）。这些结构域与顺式作用反式激活应答元件（TAR）结合蛋白（TRBP）和dsRNA激活蛋白激酶（PKR）保守。TRBP和PKR通过与TAR RNA结合参与HIV-1基因表达的调控。本研究表明，RHA在TAR介导的HIV-1基因表达中也起重要作用。野生型RHA在体外和体内均优先结合TAR RNA。野生型RHA的过表达强烈增强病毒mRNA合成、病毒粒子产生以及HIV-1长末端重复序列指导的报告基因（荧光素酶）表达。在dsRBD2中第236位残基处将赖氨酸替换为谷氨酸（RHA(K236E)）降低了其对TAR RNA的亲和力并损害了HIV-1转录活性。这些结果表明，TAR RNA是RHA dsRBDs的优先靶标，并且RHA部分通过RHA与TAR的结合在体内增强HIV-1转录。
