The mechanism of an exceptional case of reinitiation after translation of a long ORF reveals why such events do not generally occur in mammalian mRNA translation
The mechanism of an exceptional case of reinitiation after translation of a long ORF reveals why such events do not generally occur in mammalian mRNA translation
复制标题
DOI：
10.1101/gad.439507
复制
发表时间：
2007-12-01
影响因子：
10.5
通讯作者：
Jackson, Richard J.
中科院分区：
文献类型：
--
作者：
Poyry, Tuija A. A.;Kaminski, Ann;Jackson, Richard J.
猫杯状病毒的亚基因组mRNA为双顺反子，两顺反子由四个核苷酸重叠，上游顺反子编码一个75 kDa的主要衣壳蛋白前体(Pre-VP1)，下游顺反子编码一个10 kDa的小衣壳蛋白。网织红细胞裂解物中的翻译动力学表明，下游的顺反子是通过终止-重新启动过程翻译的，这是不寻常的，因为不需要eIF4G或eIF4F复合体。重新启动需要VP1前ORF的3‘端87个核苷酸(NT)，而不需要其他病毒序列。再起始位点的选择是因为它与这个87-nT元件的接近，而不是它与前VP1 ORF终止密码子的接近，尽管这必须位于重新启动密码子下游不超过-30个核苷酸。结果表明，该87-NT元件可与40S核糖体亚基和启动因子eIF3结合，添加辅助eIF3可提高再启动效率。重新启动缺陷的突变体对eIF3或缺陷的40S亚单位结合(或两者)的亲和力降低。这些结果表明，在获得eIF2/GTP/Met-tRNA(I)三元复合体后，在终止后核糖体分解过程中形成的一些eIF3/40S复合体与这个87-NT元件结合在适合重新引发的位置。
The subgenomic mRNA of feline caliciviruses is bicistronic with the two cistrons overlapping by four nucleotides, ...AUGA. The upstream cistron encodes a 75-kDa major capsid protein precursor (pre-VP1), and the downstream cistron a 10-kDa minor capsid protein. The kinetics of translation in reticulocyte lysates show that the downstream cistron is translated by a termination-reinitiation process, which is unusual in not requiring eIF4G or the eIF4F complex. Reinitiation requires the 3 '-terminal 87 nucleotides (nt) of the pre-VP1 ORF, but no other viral sequences. The reinitiation site is selected by virtue of its proximity to this 87-nt element, and not its proximity to the pre-VP1 ORF stop codon, although this must be located not more than -30 nt downstream from the restart codon. This 87-nt element was shown to bind 40S ribosomal subunits and initiation factor eIF3, and addition of supplementary eIF3 enhanced reinitiation efficiency. Mutants defective in reinitiation showed reduced affinity for eIF3 or defective 40S subunit binding (or both). These results suggest a mechanism in which some of the eIF3/40S complexes formed during disassembly of post-termination ribosomes bind to this 87-nt element in a position appropriate for reinitiation following acquisition of an eIF2/GTP/Met-tRNA(i) ternary complex.