The Impact of Leadered and Leaderless Gene Structures on Translation Efficiency, Transcript Stability, and Predicted Transcription Rates in Mycobacterium smegmatis
The Impact of Leadered and Leaderless Gene Structures on Translation Efficiency, Transcript Stability, and Predicted Transcription Rates in Mycobacterium smegmatis
复制标题
DOI：
10.1128/jb.00746-19
复制
发表时间：
2020-05-01
影响因子：
3.2
通讯作者：
Shell, Scarlet S.
中科院分区：
文献类型：
--
作者：
Nguyen, Tien G.;Vargas-Blanco, Diego A.;Shell, Scarlet S.
基因表达的调节对于结核分枝杆菌耐受感染过程中遇到的应激源以及非致病性分枝杆菌（例如耻垢分枝杆菌）在环境应激源中生存至关重要。与更好研究的模型不同，分枝杆菌将大约 14% 的基因表达为无领导转录本。然而，无前导序列转录结构对分枝杆菌中 mRNA 半衰期和翻译效率的影响尚未得到直接测试。对于引导转录本，5' 非翻译区 (UTR) 对 mRNA 半衰期和翻译效率的贡献同样未知。在结核分枝杆菌和耻垢分枝杆菌中，必需的西格玛因子 SigA 由半衰期相对较短的转录物编码。我们假设 sigA 的长 5' UTR 导致了这种不稳定。为了测试这一点，我们构建了荧光报告基因并测量了蛋白质丰度、mRNA 丰度和 mRNA 半衰期，并计算了相对转录物生产率。与分枝杆菌表达质粒中常用的合成 5' UTR 相比，sigA 5' UTR 提高了转录物生产率、缩短了 mRNA 半衰期并降低了表观翻译率。无领导者转录本的翻译效率似乎与具有 sigA 5' UTR 的转录本相似，但预测的转录本生产率较低。对结核分枝杆菌 mRNA 和蛋白质丰度的全球比较未能揭示有前导转录本和无前导转录本的蛋白质/mRNA 比率的系统差异，这表明翻译效率的变异性很大程度上是由前导状态以外的因素驱动的。我们的数据还根据另一种模型进行了讨论，该模型得出了不同的结论，并表明无领导者转录本的翻译效率确实可能较低。 重要性 结核病由结核分枝杆菌引起，是一个主要的公共卫生问题，每年在全球造成 150 万人死亡。在感染期间，结核分枝杆菌必须改变其基因表达模式以适应其遇到的应激条件。了解结核分枝杆菌如何调节基因表达可能为干扰细菌生存的方法提供线索。基因表达包括转录、mRNA 降解和翻译。在这里，我们使用耻垢分枝杆菌作为模式生物来研究 5' 非翻译区如何以多种方式影响基因表达的这三个方面。我们还深入了解了无前导 mRNA 的表达，这种 mRNA 缺乏 5' 非翻译区，并且在分枝杆菌中异常普遍。
Regulation of gene expression is critical for Mycobacterium tuberculosis to tolerate stressors encountered during infection and for nonpathogenic mycobacteria such as Mycobacterium smegmatis to survive environmental stressors. Unlike better-studied models, mycobacteria express similar to 14% of their genes as leaderless transcripts. However, the impacts of leaderless transcript structures on mRNA half-life and translation efficiency in mycobacteria have not been directly tested. For leadered transcripts, the contributions of 5' untranslated regions (UTRs) to mRNA half-life and translation efficiency are similarly unknown. In M. tuberculosis and M. smegmatis, the essential sigma factor, SigA, is encoded by a transcript with a relatively short half-life. We hypothesized that the long 5' UTR of sigA causes this instability. To test this, we constructed fluorescence reporters and measured protein abundance, mRNA abundance, and mRNA half-life and calculated relative transcript production rates. The sigA 5' UTR conferred an increased transcript production rate, shorter mRNA half-life, and decreased apparent translation rate compared to a synthetic 5' UTR commonly used in mycobacterial expression plasmids. Leaderless transcripts appeared to be translated with similar efficiency as those with the sigA 5' UTR but had lower predicted transcript production rates. A global comparison of M. tuberculosis mRNA and protein abundances failed to reveal systematic differences in protein/mRNA ratios for leadered and leaderless transcripts, suggesting that variability in translation efficiency is largely driven by factors other than leader status. Our data are also discussed in light of an alternative model that leads to different conclusions and suggests leaderless transcripts may indeed be translated less efficiently.IMPORTANCE Tuberculosis, caused by Mycobacterium tuberculosis, is a major public health problem killing 1.5 million people globally each year. During infection, M. tuberculosis must alter its gene expression patterns to adapt to the stress conditions it encounters. Understanding how M. tuberculosis regulates gene expression may provide clues for ways to interfere with the bacterium's survival. Gene expression encompasses transcription, mRNA degradation, and translation. Here, we used Mycobacterium smegmatis as a model organism to study how 5' untranslated regions affect these three facets of gene expression in multiple ways. We furthermore provide insight into the expression of leaderless mRNAs, which lack 5' untranslated regions and are unusually prevalent in mycobacteria.