DHX29 reduces leaky scanning through an upstream AUG codon regardless of its nucleotide context.
DHX29 reduces leaky scanning through an upstream AUG codon regardless of its nucleotide context.
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DOI：
10.1093/nar/gkw240
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发表时间：
2016-05-19
影响因子：
14.9
通讯作者：
Pisarev AV
中科院分区：
文献类型：
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作者：
Pisareva VP;Pisarev AV
在真核生物翻译起始过程中，由40S核糖体亚基、真核起始因子（eIFs）和起始tRNA组成的43S预起始复合物（43S PIC）对mRNA进行扫描以找到合适的起始密码子。在起始密码子选择准确性方面，eIF1和eIF1A起着关键作用，而哺乳动物特有的DHX29解旋酶对结构化mRNA的核糖体扫描有重要贡献。在此，我们表明在核糖体扫描过程中，无论其核苷酸环境如何，DHX29都能刺激对AUG密码子的识别，但不刺激对近同源的CUG密码子的识别。这种刺激作用取决于DHX29和eIF1A之间的接触。独特的DHX29 N末端结构域与mRNA入口附近的核糖体位点结合，并在此处与eIF1A的OB结构域接触。紫外线交联实验表明，DHX29可能会在核糖体复合物的关键核苷酸环境位置重新排列eIF1A和eIF2α。有趣的是，在没有eIF1A的情况下，DHX29会阻碍48S起始复合物的形成，这可能是因为它形成了一个物理屏障，阻止43S PIC加载到mRNA上。突变分析使我们能够区分DHX29的mRNA解旋和密码子选择活性。因此，DHX29是另一个有助于起始密码子选择的起始因子的例子。
During eukaryotic translation initiation, the 43S preinitiation complex (43S PIC), consisting of the 40S ribosomal subunit, eukaryotic initiation factors (eIFs) and initiator tRNA scans mRNA to find an appropriate start codon. Key roles in the accuracy of initiation codon selection belong to eIF1 and eIF1A, whereas the mammalian-specific DHX29 helicase substantially contributes to ribosomal scanning of structured mRNAs. Here, we show that DHX29 stimulates the recognition of the AUG codon but not the near-cognate CUG codon regardless of its nucleotide context during ribosomal scanning. The stimulatory effect depends on the contact between DHX29 and eIF1A. The unique DHX29 N-terminal domain binds to the ribosomal site near the mRNA entrance, where it contacts the eIF1A OB domain. UV crosslinking assays revealed that DHX29 may rearrange eIF1A and eIF2α in key nucleotide context positions of ribosomal complexes. Interestingly, DHX29 impedes the 48S initiation complex formation in the absence of eIF1A perhaps due to forming a physical barrier that prevents the 43S PIC from loading onto mRNA. Mutational analysis allowed us to split the mRNA unwinding and codon selection activities of DHX29. Thus, DHX29 is another example of an initiation factor contributing to start codon selection.