EBV noncoding RNA EBER2 interacts with host RNA-binding proteins to regulate viral gene expression
EBV noncoding RNA EBER2 interacts with host RNA-binding proteins to regulate viral gene expression
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DOI：
10.1073/pnas.1601773113
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发表时间：
2016-03-22
影响因子：
11.1
通讯作者：
Steitz, Joan A.
中科院分区：
文献类型：
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作者：
Lee, Nara;Yario, Therese A.;Steitz, Joan A.
Epstein-Barr 病毒 (EBV) 产生一种高度丰富的非编码 RNA，称为 EBV 编码 RNA 2 (EBER2)，它与宿主转录因子配对盒蛋白 5 (PAX5) 间接相互作用，调节病毒潜伏膜蛋白 1/2 (LMP1/2) 基因表达以及 EBV 裂解复制。为了鉴定中间蛋白，我们分离了包含 EBER2-PAX5 的复合物，并通过质谱分析了蛋白质成分。最重要的候选者包括三种宿主蛋白富含脯氨酸和谷氨酰胺剪接因子 (SFPQ)、不含 POU 结构域的八聚体结合蛋白 (NONO) 和 RNA 结合基序蛋白 14 (RBM14)，据报道，所有这些都是称为副斑点的核体的组成部分。体内RNA-蛋白质交联表明SFPQ和RBM14直接接触EBER2。使用重组蛋白的结合研究表明，SFPQ 和 NONO 与 PAX5 结合，可能桥接其与 EBER2 的相互作用。与 EBER2 或 PAX5 耗竭类似，三种宿主 RNA 结合蛋白中任何一种的敲低都会导致病毒 LMP2A mRNA 水平上调，支持这些新发现的因子与 EBER2 和 PAX5 之间的生理相关相互作用。这些 EBER2 相互作用蛋白的鉴定使得能够寻找以类似于 EBER2 的方式调节宿主基因表达的细胞非编码 RNA。
Epstein-Barr virus (EBV) produces a highly abundant noncoding RNA called EBV-encoded RNA 2 (EBER2) that interacts indirectly with the host transcription factor paired box protein 5 (PAX5) to regulate viral latent membrane protein 1/2 (LMP1/2) gene expression as well as EBV lytic replication. To identify intermediary proteins, we isolated EBER2-PAX5-containing complexes and analyzed the protein components by mass spectrometry. The top candidates include three host proteins splicing factor proline and glutamine rich (SFPQ), non-POU domain-containing octamer-binding protein (NONO), and RNA binding motif protein 14 (RBM14), all reported to be components of nuclear bodies called paraspeckles. In vivo RNA-protein crosslinking indicates that SFPQ and RBM14 contact EBER2 directly. Binding studies using recombinant proteins demonstrate that SFPQ and NONO associate with PAX5, potentially bridging its interaction with EBER2. Similar to EBER2 or PAX5 depletion, knockdown of any of the three host RNA-binding proteins results in the up-regulation of viral LMP2A mRNA levels, supporting a physiologically relevant interaction of these newly identified factors with EBER2 and PAX5. Identification of these EBER2-interacting proteins enables the search for cellular noncoding RNAs that regulate host gene expression in a manner similar to EBER2.