Recruitment of the complete hTREX complex is required for Kaposi's sarcoma-associated herpesvirus intronless mRNA nuclear export and virus replication.
Recruitment of the complete hTREX complex is required for Kaposi's sarcoma-associated herpesvirus intronless mRNA nuclear export and virus replication.
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DOI：
10.1371/journal.ppat.1000194
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发表时间：
2008-10
期刊：
影响因子：
6.7
通讯作者：
Whitehouse, Adrian
中科院分区：
文献类型：
--
作者：
Boyne, James R.;Colgan, Kevin J.;Whitehouse, Adrian
细胞前体mRNA在核输出之前经历各种转录后加工事件，包括加帽、剪接和聚腺苷酸化。剪接对于mRNA核输出特别重要，因为两种不同的多蛋白复合物，称为人TREX（hTREX）和外显子连接复合物（EJC），以剪接依赖性方式被募集到mRNA中。相反，一些卡波西肉瘤相关疱疹病毒（KSHV）裂解mRNA缺乏内含子，并通过病毒编码的ORF 57蛋白输出。在本文中，我们表明ORF 57结合到无内含子病毒mRNA，并用于募集完整的hTREX复合物，但不是EJC，以组装输出组分病毒核糖核蛋白颗粒（vRNP）。该vRNP的形成由ORF 57和hTREX输出衔接蛋白Aly之间的直接相互作用介导。Aly又直接与DEAD-盒蛋白UAP 56相互作用，后者作为桥梁将剩余的hTREX蛋白募集到复合物中。此外，我们表明，ORF 57中的点突变破坏了ORF 57-Aly相互作用，导致ORF 57介导的整个hTREX复合物向无内含子病毒mRNA的募集失败，并抑制mRNA随后的核输出和病毒复制。此外，我们利用反式显性Aly突变体来防止完整ORF 57-hTREX复合物的组装;这导致由结合至ORF 57的病毒mRNA、Aly和核输出因子TAP组成的vRNP。引人注目的是，尽管输出衔接子Aly和输出因子TAP都存在于病毒mRNP上，但在不存在剩余hTREX组分（UAP 56和hTH-复合物）的情况下观察到无内含子病毒mRNA输出和病毒复制的显著减少。总之，这些数据提供了完整的hTREX复合物对于KSHV无内含子mRNA的输出和感染性病毒产生至关重要的第一个直接证据。在细胞核中基因表达后，新转录的信使RNA（mRNA）被输出到细胞质，在那里它被翻译成蛋白质。在哺乳动物中，绝大多数mRNA含有内含子，其必须在核输出之前被剪接体去除。除了切除内含子之外，剪接对于将几种蛋白质复合物募集到mRNA中也是必不可少的，一个例子是mRNA输出所需的人类转录/输出复合物。疱疹病毒，如卡波西肉瘤相关疱疹病毒，通过劫持宿主细胞生物机器的组分，包括mRNA输出所需的那些蛋白质来复制。疱疹病毒学中一个有趣的警告是，疱疹病毒，如卡波西肉瘤相关疱疹病毒，产生一些缺乏内含子的mRNA，不进行剪接。那么这些无内含子的mRNA是如何输出到细胞质的呢？答案在于一种名为ORF 57的病毒蛋白，它能够与无内含子的mRNA结合，然后将它们输出到细胞质中。ORF 57通过模拟剪接和募集人转录/输出复合物至无内含子病毒mRNA来实现该功能，从而促进其输出至细胞质中。
A cellular pre-mRNA undergoes various post-transcriptional processing events, including capping, splicing and polyadenylation prior to nuclear export. Splicing is particularly important for mRNA nuclear export as two distinct multi-protein complexes, known as human TREX (hTREX) and the exon-junction complex (EJC), are recruited to the mRNA in a splicing-dependent manner. In contrast, a number of Kaposi's sarcoma–associated herpesvirus (KSHV) lytic mRNAs lack introns and are exported by the virus-encoded ORF57 protein. Herein we show that ORF57 binds to intronless viral mRNAs and functions to recruit the complete hTREX complex, but not the EJC, in order assemble an export component viral ribonucleoprotein particle (vRNP). The formation of this vRNP is mediated by a direct interaction between ORF57 and the hTREX export adapter protein, Aly. Aly in turn interacts directly with the DEAD-box protein UAP56, which functions as a bridge to recruit the remaining hTREX proteins to the complex. Moreover, we show that a point mutation in ORF57 which disrupts the ORF57-Aly interaction leads to a failure in the ORF57-mediated recruitment of the entire hTREX complex to the intronless viral mRNA and inhibits the mRNAs subsequent nuclear export and virus replication. Furthermore, we have utilised a trans-dominant Aly mutant to prevent the assembly of the complete ORF57-hTREX complex; this results in a vRNP consisting of viral mRNA bound to ORF57, Aly and the nuclear export factor, TAP. Strikingly, although both the export adapter Aly and the export factor TAP were present on the viral mRNP, a dramatic decrease in intronless viral mRNA export and virus replication was observed in the absence of the remaining hTREX components (UAP56 and hTHO-complex). Together, these data provide the first direct evidence that the complete hTREX complex is essential for the export of KSHV intronless mRNAs and infectious virus production. Following gene expression in the nucleus, newly transcribed messenger RNA (mRNA) is exported to the cytoplasm, where it is translated into protein. In mammals the vast majority of mRNAs contain introns that must be removed by the spliceosome prior to nuclear export. In addition to excising introns, splicing is also essential for the recruitment of a several protein complexes to mRNA, one example being the human transcription/export complex, which is required for mRNA export. Herpesviruses, such as Kaposi's sarcoma–associated herpesvirus, replicate by hijacking components of the host cells biological machinery, including those proteins necessary for mRNA export. An intriguing caveat in herpesvirology is that herpesviruses, such as Kaposi's sarcoma–associated herpesvirus, produce some mRNAs that lack introns and do not undergo splicing. How then are these intronless mRNAs exported to the cytoplasm? The answer lies in a virus protein called ORF57 that is able to bind to the intronless mRNA and then export them to the cytoplasm. ORF57 achieves this function by mimicking splicing and recruiting the human transcription/export complex to the intronless viral mRNA, thus facilitating its export into the cytoplasm.
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