基于Ribo-Seq的生物进化中翻译缓冲机制的研究

By comparing the ribosome profiling in two related species, it has been found that the divergence at the mRNA level is usually buffered by translation, so that the difference at the protein level is smaller. This buffering effect guarantees that the key proteins have similar expression levels among species, regardless of the potential deleterious effects at the mRNA level. We seek to identify the molecular mechanisms of such interesting and important phenomenon. There are two potential mechanisms: the translational negative feedback within the regulatory circuit could reduce the difference at the protein level; alternatively, the compensatory mutations at the translational level may also attenuate the difference at the protein level. We plan to generate a series of mutants to distinguish those two potential mechanisms, because mutants do not have enough time for compensatory mutations to occur and to be fixed. We will perform Ribo-Seq to gauge the negative feedback at the translational level, and compare it to the buffering effect between species. We further seek to understand the genetic basis of compensatory mutations. Our study will provide significant insights into the molecular mechanism of the pervasive and important phenomenon, translational buffering, and will also shed light on the synthetic biology, which designs and generates useful artificial organisms.

通过Ribo-Seq比较不同物种的翻译组，生物学家们观察到了一个非常有趣的现象：翻译缓冲。即，在物种间转录层面的差异会在翻译层面获得缓冲，从而在蛋白质组层面展现出较小的差异。翻译缓冲可能的生物学机制是什么呢？首先，细胞内部可能存在负反馈机制，保证mRNA水平的波动不会影响蛋白水平，从而保证细胞内的稳态不会受到破坏。第二种可能的机制是在进化过程中在不利突变的DNA背景下发生补偿突变，可以抵消第一个突变带来的不利影响。突变体没有时间积累补偿突变，因此我们将通过系统分析突变体的翻译组，在全基因组层面解析两种机制，从而阐明翻译缓冲的生物学基础。进而，我们将通过一系列遗传学和功能基因组学实验系统检测补偿突变的遗传基础，并尝试阐明翻译层面的负反馈和补偿突变的影响因素。本课题的开展将解析翻译调控这一广泛存在的重要的生物学现象，为理解细胞蛋白质组的稳定性提供理论基础，并为合成生物学提供设计参考。

本项目对翻译调控过程与机制开展了系统的研究。我们在酵母突变体中通过ribo-seq的方法检测翻译组，发现突变体中不但没有出现翻译缓冲现象，反而展现出相反的趋势：mRNA水平的提高与翻译效率的提高同时发生，或者mRNA水平的降低与翻译效率的降低同时发生。具体而言，我们发现5′-非翻译区的长度可以影响翻译效率，这主要是通过mRNA二级结构和起始密码子上游编码的uORF实现的。我们发现同义密码子的使用可以通过影响翻译效率的方式影响mRNA稳定性。我们还发现真核生物的poly(A)尾也可能通过掺入非A碱基的方式影响其与poly(A)结合蛋白的相互作用，从而影响mRNA的翻译效率。这些工作系统地解析了翻译调控的机制，为改良关键基因的翻译调控奠定了坚实的理论基础。