Non-conventional translation initiation in bacteria

原核生物中传统的翻译起始途径依赖于Shine Dalgarno（SD）基序与16 S rRNA 3 '末端的相互作用。最近，我们可以表明，翻译起始盐古菌是占主导地位的两个非传统机制（leaderless转录本和SD-少的转录本）。我们现在已经将翻译起始的表征扩展到大肠杆菌。我们可以证明，在E.大肠杆菌，表明非传统的翻译起始机制也被细菌使用。我们将使用translatome分析的转录本与非平均翻译效率的压力条件下或春雷霉素治疗的全球特征。双报告基因系统将被用来量化在各种条件下的常规和非常规机制的相对翻译效率，顺式作用元件的作用将通过诱变进行分析。起始复合物将被分离，并通过使用超软质谱和蛋白质组学，它将揭示专门的复合物是否存在于不同的条件下或非传统的转录（类似于最近确定的专门61 S核糖体）。

The conventional pathway of translation initiation in prokaryotes relies on the interaction of a Shine Dalgarno (SD) motif with the 3’-end of the 16S rRNA. Recently we could show that translation initiation in haloarchaea is predominated by two non-conventional mechanisms (leaderless transcripts and SD-less transcripts). We have now extended the characterization of translation initiation to Escherichia coli. We could show that SD-less transcripts exist also in E. coli, indicating that non-conventional mechanisms for translation initiation are also used by bacteria. We will use translatome analyses for a global characterization of transcripts with non-average translational efficiencies under stress conditions or kasugamycin treatment. A dual reporter gene system will be used to quantify relative translational efficiencies of the conventional and non-conventional mechanisms under various conditions, and the role of cis-acting elements will be analyzed by mutagenesis. Initiation complexes will be isolated and by using ultrasoft mass spectrometry and proteomics it will be revealed whether specialized complexes exist under different conditions or on non-conventional transcripts (similar to the recently identified specialized 61S ribosomes).