Characterization of functional domains of equine infectious anemia virus Rev suggests a bipartite RNA-binding domain
Characterization of functional domains of equine infectious anemia virus Rev suggests a bipartite RNA-binding domain
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DOI：
10.1128/jvi.80.8.3844-3852.2006
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发表时间：
2006-04-01
影响因子：
5.4
通讯作者：
Carpenter, S
中科院分区：
文献类型：
--
作者：
Lee, JH;Murphy, SC;Carpenter, S
马传染性贫血病毒（EIAV） Rev是一种重要的调节蛋白，可促进编码结构蛋白和基因组RNA的病毒mRNA的表达，并调节双链RNA / Rev mRNA的选择性剪接。EIAV Rev在体内具有较高的遗传变异率，并且Rev基因型和表型的变化已被证明与临床疾病的变化一致。为了更好地理解遗传变异如何改变Rev表型，我们进行了缺失和突变分析来绘制功能域，并确定了对EIAV Rev活性至关重要的特定基序。所有功能域都包含在Rev的第二外显子内。Rev外显子2内结构域的总体组织包括核输出信号，RNA结合所需的大中心区域，非必需区域以及核定位和RNA结合所需的c端区域。绿色荧光蛋白-Rev突变体的亚细胞定位表明，Rev c端区域KRRRK基序内的基本残基是Rev靶向细胞核所必需的。Rev的两个独立区域是RNA结合所必需的：一个包含残基57至130的中心区域和一个跨越残基144至165的c端区域。在这些区域内有两个不同的短的富含精氨酸的基序，它们是RNA结合所必需的，包括位于中心区域的RRDRW基序和靠近C端的KRRRK基序。这些发现表明，EIAV Rev利用了一个双rna结合结构域。
Equine infectious anemia virus (EIAV) Rev is an essential regulatory protein that facilitates expression of viral mRNAs encoding structural proteins and genomic RNA and regulates alternative splicing of the bicistronic tat/rev mRNA. EIAV Rev is characterized by a high rate of genetic variation in vivo, and changes in Rev genotype and phenotype have been shown to coincide with changes in clinical disease. To better understand how genetic variation alters Rev phenotype, we undertook deletion and mutational analyses to map functional domains and to identify specific motifs that are essential for EIAV Rev activity. All functional domains are contained within the second exon of EIAV Rev. The overall organization of domains within Rev exon 2 includes a nuclear export signal, a large central region required for RNA binding, a nonessential region, and a C-terminal region required for both nuclear localization and RNA binding. Subcellular localization or green fluorescent protein-Rev mutants indicated that basic residues within the KRRRK motif in the C-terminal region of Rev are necessary for targeting of Rev to the nucleus. Two separate regions of Rev were necessary for RNA binding: a central region encompassing residues 57 to 130 and a C-terminal region spanning residues 144 to 165. Within these regions were two distinct, short arginine-rich motifs essential for RNA binding, including an RRDRW motif in the central region and the KRRRK motif near the C terminus. These findings suggest that EIAV Rev utilizes a bipartite RNA-binding domain.