Hussmann Jeffrey A, Patchett Stephanie, Johnson Arlen, Sawyer Sara, Press William H
Institute for Computational Engineering and Sciences, University of Texas at Austin, Austin, Texas, United States of America.
Institute for Cellular and Molecular Biology, University of Texas at Austin, Austin, Texas, United States of America.
PLoS Genet. 2015 Dec 11;11(12):e1005732. doi: 10.1371/journal.pgen.1005732. eCollection 2015 Dec.
Ribosome profiling produces snapshots of the locations of actively translating ribosomes on messenger RNAs. These snapshots can be used to make inferences about translation dynamics. Recent ribosome profiling studies in yeast, however, have reached contradictory conclusions regarding the average translation rate of each codon. Some experiments have used cycloheximide (CHX) to stabilize ribosomes before measuring their positions, and these studies all counterintuitively report a weak negative correlation between the translation rate of a codon and the abundance of its cognate tRNA. In contrast, some experiments performed without CHX report strong positive correlations. To explain this contradiction, we identify unexpected patterns in ribosome density downstream of each type of codon in experiments that use CHX. These patterns are evidence that elongation continues to occur in the presence of CHX but with dramatically altered codon-specific elongation rates. The measured positions of ribosomes in these experiments therefore do not reflect the amounts of time ribosomes spend at each position in vivo. These results suggest that conclusions from experiments in yeast using CHX may need reexamination. In particular, we show that in all such experiments, codons decoded by less abundant tRNAs were in fact being translated more slowly before the addition of CHX disrupted these dynamics.
核糖体谱分析可生成信使核糖核酸（mRNA）上正在进行翻译的核糖体位置的快照。这些快照可用于推断翻译动力学。然而，最近在酵母中进行的核糖体谱分析研究对于每个密码子的平均翻译速率得出了相互矛盾的结论。一些实验在测量核糖体位置之前使用环己酰亚胺（CHX）来稳定核糖体，而这些研究都得出了与直觉相反的结果，即密码子的翻译速率与其同源tRNA的丰度之间存在微弱的负相关。相比之下，一些未使用CHX进行的实验则报告了强正相关。为了解释这一矛盾，我们在使用CHX的实验中发现了每种密码子下游核糖体密度的意外模式。这些模式表明，在存在CHX的情况下延伸仍在继续，但密码子特异性延伸速率发生了显著变化。因此，这些实验中测量的核糖体位置并不能反映核糖体在体内每个位置所花费的时间量。这些结果表明，在酵母中使用CHX进行的实验得出的结论可能需要重新审视。特别是，我们表明，在所有此类实验中，由丰度较低的tRNA解码的密码子在添加CHX破坏这些动力学之前实际上翻译得更慢。
