Kong Li Kuo, Sarnow Peter
Department of Microbiology and Immunology, Stanford University School of Medicine, Sherman Fairchild Science Building, 299 Campus Drive, Stanford, CA 94305, USA.
J Virol. 2002 Dec;76(24):12457-62. doi: 10.1128/jvi.76.24.12457-12462.2002.
Translation initiation in many eukaryotic mRNAs is modulated by an interaction between the cap binding protein complex, bound to the 5' end of the mRNA, and the polyadenosine binding protein, bound to the 3'-terminal polyadenosine sequences. A few cellular and viral mRNAs, such as the hepatitis C virus (HCV) mRNA genome, lack 3'-terminal polyadenosine sequences. For such mRNAs, the question of whether their 3'-end sequences also regulate the initiation phase of protein synthesis via an interaction with their 5' ends has received intense scrutiny. For HCV mRNA, various experimental designs have led to conflicting interpretations, that the 3' end of the RNA can modulate translation initiation either in a positive or in a negative fashion. To examine the possibility of end-to-end communication in HCV in detail, mRNAs containing the HCV internal ribosome entry site linked to a luciferase coding region, followed by different 3' noncoding regions, were expressed in the cytoplasm of cultured cells by T7 RNA polymerase. The intracellular translation efficiencies, steady-state levels, stabilities, and 3'-end sequences of these chimeric RNAs were examined. It was found that the HCV 3' noncoding region modulates neither the translation nor the stability of the mRNAs. Thus, there is no detectable end-to-end communication in cytoplasmically expressed chimeric mRNAs containing the HCV noncoding regions. However, it remains an open question whether end-to-end communication occurs in full-length HCV mRNAs in the infected liver.
许多真核生物mRNA的翻译起始是由结合在mRNA 5'端的帽结合蛋白复合体与结合在3'端多聚腺苷酸序列上的多聚腺苷酸结合蛋白之间的相互作用所调节的。一些细胞和病毒mRNA,如丙型肝炎病毒(HCV)mRNA基因组,缺乏3'端多聚腺苷酸序列。对于这类mRNA,其3'端序列是否也通过与5'端的相互作用来调节蛋白质合成的起始阶段这一问题受到了广泛关注。对于HCV mRNA,各种实验设计得出了相互矛盾的解释,即RNA的3'端可以以正向或负向方式调节翻译起始。为了详细研究HCV中首尾通信的可能性,通过T7 RNA聚合酶在培养细胞的细胞质中表达了含有与荧光素酶编码区相连的HCV内部核糖体进入位点,随后是不同3'非编码区的mRNA。检测了这些嵌合RNA的细胞内翻译效率、稳态水平、稳定性和3'端序列。结果发现,HCV 3'非编码区既不调节mRNA的翻译也不调节其稳定性。因此,在含有HCV非编码区的细胞质表达嵌合mRNA中没有可检测到的首尾通信。然而,在受感染肝脏中的全长HCV mRNA中是否发生首尾通信仍是一个悬而未决的问题。