The 5'-poly(A) leader of poxvirus mRNA confers a translational advantage that can be achieved in cells with impaired cap-dependent translation

The poly(A) leader at the 5’-untranslated region (5’-UTR) is an unusually striking feature of all poxvirus mRNAs transcribed after viral DNA replication (post-replicative mRNAs). These poly(A) leaders are non-templated and of heterogeneous lengths; and their function during poxvirus infection remains a long-standing question. Here, we discovered that a 5’-poly(A) leader conferred a selective translational advantage to mRNA in poxvirus-infected cells. A constitutive and uninterrupted 5’-poly(A) leader with 12 residues was optimal. Because the most frequent lengths of the 5’-poly(A) leaders are 8–12 residues, the result suggests that the poly(A) leader has been evolutionarily optimized to boost poxvirus protein production. A 5’-poly(A) leader also could increase protein production in the bacteriophage T7 promoter-based expression system of vaccinia virus, the prototypic member of poxviruses. Interestingly, although vaccinia virus post-replicative mRNAs do have 5’- methylated guanosine caps and can use cap-dependent translation, in vaccinia virus-infected cells, mRNA with a 5’-poly(A) leader could also be efficiently translated in cells with impaired cap-dependent translation. However, the translation was not mediated through an internal ribosome entry site (IRES). These results point to a fundamental mechanism poxvirus uses to efficiently translate its post-replicative mRNAs.

位于5'非翻译区（5’-untranslated region, 5’-UTR）的poly(A)前导序列（poly(A) leader）是所有痘病毒（poxvirus）在病毒DNA复制后转录的mRNA（即复制后mRNA，post-replicative mRNAs）极为显著的特征。这类poly(A)前导序列属于非模板化序列，且长度不均一；其在痘病毒感染过程中的功能长期以来一直是一个悬而未决的问题。本研究发现，5'端poly(A)前导序列可赋予痘病毒感染细胞内的mRNA选择性翻译优势。由12个残基组成的连续不间断的5'poly(A)前导序列为最优形式。鉴于痘病毒5'poly(A)前导序列的最常见长度为8~12个残基，该结果提示poly(A)前导序列已在进化中得到优化，以提升痘病毒的蛋白质合成效率。在作为痘病毒原型成员的牛痘病毒（vaccinia virus）的噬菌体T7启动子（bacteriophage T7 promoter）表达系统中，5'poly(A)前导序列同样可提高蛋白质产量。有趣的是，尽管牛痘病毒的复制后mRNA带有5'甲基化鸟苷帽（5’-methylated guanosine caps），且可通过帽依赖型翻译（cap-dependent translation）途径进行翻译，但在牛痘病毒感染的细胞中，携带5'poly(A)前导序列的mRNA也可在帽依赖型翻译功能受损的细胞中被高效翻译。不过，该翻译过程并非通过内部核糖体进入位点（internal ribosome entry site, IRES）介导。上述研究结果揭示了痘病毒用以高效翻译其复制后mRNA的一种核心机制。