Hepatitis C virus core protein acts as a trans-modulating factor on internal translation initiation of the viral RNA
Hepatitis C virus core protein acts as a trans-modulating factor on internal translation initiation of the viral RNA
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DOI：
10.1074/jbc.m501826200
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发表时间：
2005-05-06
影响因子：
4.8
通讯作者：
Cahour, A
中科院分区：
文献类型：
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作者：
Boni, S;Lavergne, JP;Cahour, A
丙型肝炎病毒（HCV） RNA的翻译起始是通过位于其5'端的内部核糖体进入位点（IRES）发生的。作为一种正链病毒，HCV使用基因组RNA模板进行翻译和复制，但这两个过程之间的转换仍然知之甚少。HCV核心蛋白（HCV- c）被认为是调节这种调节的一个很好的候选者。然而，目前的数据在将HCV翻译中的任何潜在作用归因于HCV核心蛋白方面仍然存在争议。本研究通过在蔗糖梯度上离心，证明了HCV- c对HCV IRES和40s核糖体亚基具有结合活性。为了进一步了解这些相互作用，我们使用体外报告基因法研究了外源性添加纯化HCV- c对HCV IRES活性的影响。我们发现HCV IRES介导的翻译被HCV- c以剂量依赖的方式特异性调节，在添加低剂量HCV- c时，IRES效率可达到5倍的刺激，然后在高剂量时降低。有趣的是，IRES某些结构域的突变以及上游报告基因的存在都会导致预期效果的变化，这与HCV IRES功能对其整体结构的高度依赖是一致的。总之，这些结果表明HCV核心蛋白参与了HCV翻译起始的紧密调节，这取决于它的浓度，它们表明该蛋白在病毒基因表达中具有重要的生物学作用。
Translation initiation of hepatitis C virus (HCV) RNA occurs through an internal ribosome entry site (IRES) located at its 5' end. As a positive-stranded virus, HCV uses the genomic RNA template for translation and replication, but the transition between these two processes remains poorly understood. HCV core protein (HCV-C) has been proposed as a good candidate to modulate such a regulation. However, current data are still the subject of controversy in attributing any potential role in HCV translation to the HCV core protein. Here we demonstrate that HCV-C displays binding activities toward both HCV IRES and the 40 S ribosomal subunit by using centrifugation on sucrose gradients. To gain further insight into these interactions, we investigated the effect of exogenous addition of purified HCV-C on HCV IRES activity by using an in vitro reporter assay. We found that HCV IRES-mediated translation was specifically modulated by HCV-C provided in trans, in a dose-dependent manner, with up to a 5-fold stimulation of the IRES efficiency upon addition of low amounts of HCV-C, followed by a decrease at high doses. Interestingly, mutations within some domains of the IRES as well as the presence of an upstream reporter gene both lead to changes in the expected effects, consistent with the high dependence of HCV IRES function on its overall structure. Collectively, these results indicate that the HCV core protein is involved in a tight modulation of HCV translation initiation, depending on its concentration, and they suggest an important biological role of this protein in viral gene expression.