Establishment and Application of a High Throughput Screening System Targeting the Interaction between HCV Internal Ribosome Entry Site and Human Eukaryotic Translation Initiation Factor 3.
Establishment and Application of a High Throughput Screening System Targeting the Interaction between HCV Internal Ribosome Entry Site and Human Eukaryotic Translation Initiation Factor 3.
复制标题
HCV内部核糖体进入位点与人类真核翻译起始因子3相互作用的高通量筛选系统的建立及应用
DOI：
10.3389/fmicb.2017.00977
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发表时间：
2017
影响因子：
5.2
通讯作者：
Sun C
中科院分区：
文献类型：
--
作者：
Zhu Y;Huang P;Yang N;Liu R;Liu X;Dai H;Zhang L;Song F;Sun C
病毒是细胞内专性寄生虫，通常会招募宿主细胞机制进行复制。人真核翻译起始因子3（eIF 3）可被丙型肝炎病毒（HCV）内部核糖体进入位点（IRES）直接募集，促进病毒蛋白的翻译。在这项研究中，我们建立了一个基于荧光偏振（FP）的高通量筛选（HTS）系统，针对HCV IRES和eIF 3之间的相互作用。通过用该HTS系统筛选总共894种化合物，发现两种化合物（Mucl 39526和NP 39）干扰HCV IRES和eIF 3之间的相互作用。并进一步证明了这两种化合物在体外对HCV IRES依赖的翻译有抑制作用。因此，该HTS系统可用于筛选针对人eIF 3的潜在HCV复制抑制剂，这有助于克服病毒耐药性问题。令人惊讶的是，一种催产素拮抗剂化合物HP-3被发现通过该HTS系统显著增强HCV IRES与eIF 3之间的相互作用。HP-3在体内外均能与HCV IRES直接相互作用，促进HCV IRES依赖的翻译，提示HP-3具有促进HCV复制的潜力。因此，该系统也可用于筛选潜在的HCV复制增强子，这对了解病毒复制和筛选新型抗病毒药物具有重要意义。据我们所知，这是第一个靶向eIF 3和HCV IRES之间相互作用的HTS系统，其可用于筛选潜在的HCV复制抑制剂和增强剂。
Viruses are intracellular obligate parasites and the host cellular machinery is usually recruited for their replication. Human eukaryotic translation initiation factor 3 (eIF3) could be directly recruited by the hepatitis C virus (HCV) internal ribosome entry site (IRES) to promote the translation of viral proteins. In this study, we establish a fluorescence polarization (FP) based high throughput screening (HTS) system targeting the interaction between HCV IRES and eIF3. By screening a total of 894 compounds with this HTS system, two compounds (Mucl39526 and NP39) are found to disturb the interaction between HCV IRES and eIF3. And these two compounds are further demonstrated to inhibit the HCV IRES-dependent translation in vitro. Thus, this HTS system is functional to screen the potential HCV replication inhibitors targeting human eIF3, which is helpful to overcome the problem of viral resistance. Surprisingly, one compound HP-3, a kind of oxytocin antagonist, is discovered to significantly enhance the interaction between HCV IRES and eIF3 by this HTS system. HP-3 is demonstrated to directly interact with HCV IRES and promote the HCV IRES-dependent translation both in vitro and in vivo, which strongly suggests that HP-3 has potentials to promote HCV replication. Therefore, this HTS system is also useful to screen the potential HCV replication enhancers, which is meaningful for understanding the viral replication and screening novel antiviral drugs. To our knowledge, this is the first HTS system targeting the interaction between eIF3 and HCV IRES, which could be applied to screen both potential HCV replication inhibitors and enhancers.