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首页> 外文期刊>Proceedings of the National Academy of Sciences of the United States of America >Regulation of eukaryotic protein synthesis: Selective influenza viral mRNA translation is mediated by the cellular RNA-binding protein GRSF-1
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Regulation of eukaryotic protein synthesis: Selective influenza viral mRNA translation is mediated by the cellular RNA-binding protein GRSF-1

机译：真核蛋白合成的调节：选择性流感病毒mRNA翻译是由细胞RNA结合蛋白GRSF-1介导的

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摘要

To better understand regulation of eukary- otic protein synthesis, we studied cellular and viral mRNA translation in influenza virus-infected cells. Influenza virus infection results in a dramatic shut-off of cellular protein synthesis that is concomitant with selective viral mRNA translation. Earlier work showed that these events are medi- ated by viral and/or cellular factors binding to the 5' un- translated region (5' UTR) of viral mRNAs. To identify trans-acting cellular proteins responsible for selective viral protein synthesis, we employed the yeast three-hybrid system. Using the 5' UTR of the influenza virus nucleocapsid protein (NP) mRNA as bait, we identified the cellular RNA- recognition motif containing RNA-binding protein G-rich sequence factor 1 (GRSF-1) as a positive-acting translational regulatory factor. The in vivo yeast assay revealed GRSF-1 specifically bound to the NP 5' UTR but not select NP 5' UTR mutants or cellular RNA 5' UTRs. These data were confirmed by gel shift assays using recombinant GRSF-1. Importantly, recombinant GRSF-1 specifically stimulated translation of a NP 5' UTR-driven template in cell-free translation systems. Furthermore, translation efficiency of NP 5' UTR-driven templates was reduced markedly in GRSF-1-depleted HeLa cell extracts, but restored in GRSF-1-reconstituted extracts. GRSF-1 also stimulated translation of an NP 5' UTR-driven templale in HeLa cell extracts that were depleted of essential factors by addition of RNA oligonucleotides representing the viral 5' UTR RNA. Taken together, these data document the functional demon
机译：为了更好地理解真核蛋白合成的调控，我们研究了流感病毒感染细胞中细胞和病毒mRNA的翻译。流感病毒感染导致细胞蛋白质合成急剧关闭，并伴有选择性病毒mRNA翻译。较早的工作表明，这些事件是由与病毒mRNA的5'非翻译区（5'UTR）结合的病毒和/或细胞因子介导的。为了鉴定负责选择性病毒蛋白合成的反式作用细胞蛋白，我们采用了酵母三杂交系统。使用流感病毒核衣壳蛋白（NP）mRNA的5'UTR作为诱饵，我们确定了包含RNA结合蛋白富G序列因子1（GRSF-1）的细胞RNA识别基序为正向翻译调节因子。体内酵母分析显示GRSF-1特异性结合NP 5'UTR，但未选择NP 5'UTR突变体或细胞RNA 5'UTR。这些数据通过使用重组GRSF-1的凝胶位移测定法得到证实。重要的是，重组GRSF-1在无细胞翻译系统中特异性刺激NP 5'UTR驱动的模板的翻译。此外，在耗尽GRSF-1的HeLa细胞提取物中，NP 5'UTR驱动的模板的翻译效率显着降低，但在重组GRSF-1的提取物中得以恢复。 GRSF-1还刺激了HeLa细胞提取物中NP 5'UTR驱动的模板的翻译，该提取物中通过添加代表病毒5'UTR RNA的RNA寡核苷酸而耗尽了必需因子。总而言之，这些数据记录了功能演示

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