mRNA-programmed translation pauses in the targeting of E. coli membrane proteins.
mRNA-programmed translation pauses in the targeting of E. coli membrane proteins.
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DOI：
10.7554/elife.03440
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发表时间：
2014-08-18
期刊：
影响因子：
7.7
通讯作者：
Pilpel Y
中科院分区：
文献类型：
--
作者：
Fluman N;Navon S;Bibi E;Pilpel Y
在所有活的生物体中，翻译膜蛋白的核糖体在翻译早期通过普遍存在的信号识别颗粒（SRP）系统靶向膜translocons。在真核生物中，SRP Alu结构域阻止膜蛋白的翻译延伸，直到靶向完成。然而，奇怪的是，大多数真细菌都缺乏Alu结构域。在这项研究中，通过分析全基因组的翻译速率数据，我们确定了一个潜在的补偿机制，在E。在膜蛋白靶向过程中减缓翻译。潜在的机制可能是编程到编码序列中，在翻译的早期阶段，Shine-Dalgarno样元件在战略位置触发延长暂停。我们提供的实验证据表明，在靶向过程中减缓翻译并提高膜蛋白生产保真度，因为它与过表达膜蛋白的更好折叠相关。因此，缓慢延伸对于E.大肠杆菌，它利用不同于真核生物的机制来控制翻译速度。DOI：http://dx.doi.org/10.7554/eLife.03440.001蛋白质是由长链状分子构成的。首先，一段DNA被复制成信使RNA（或mRNA）分子，然后与称为核糖体的大分子复合物结合。核糖体读取并翻译mRNA序列中的代码以构建蛋白质链，然后折叠成特定的三维形状以允许蛋白质执行其功能。许多蛋白质也需要靶向细胞内的正确位置，以发挥其作用。一些蛋白质必须插入细胞膜，这些蛋白质在构建过程中被另一种称为信号识别颗粒（或简称SRP）的分子复合物引导到细胞膜上。当新蛋白从核糖体中出现时，SRP与新蛋白结合，并帮助将其引导到膜上。为了确保膜蛋白正确折叠，当蛋白质被靶向膜时，它们的翻译被暂停。植物、动物和其他真核生物通过SRP复合物的一个独特部分来完成这一过程，该复合物只有在翻译核糖体靠近膜时才允许翻译继续进行。然而，大多数细菌缺乏SRP复合物的这一部分，但它们仍然能够准确地将新的、正确折叠的蛋白质插入它们的膜中。这表明细菌细胞中一定存在另一种机制。Fluman等人研究了一个现有的数据集，该数据集测量了在任何给定时间内，在细菌E中沿着mRNA分子长度的不同点上发现了多少核糖体。杆菌如果核糖体始终存在于给定mRNA分子的特定位点，则表明这些位点是翻译暂停发生的位点。特定的短mRNA序列--与核糖体结合并将其固定在适当的位置--经常在这些暂停位点发现。这些序列与另一种称为Shine-Dalgarno序列的序列相似，该序列通常也存在于mRNA分子的起始处，其功能是招募核糖体并开始开始翻译过程。Fluman等人揭示，膜蛋白的mRNA在其编码区的早期就包含这些相似的序列。有些蛋白质看起来可能会在新形成的蛋白质链从核糖体中出现之前暂停翻译，这可以给核糖体时间靶向细胞膜。其他类似Shine-Dalgarno的序列稍后在mRNA分子中被发现，这些蛋白质链来回穿过膜几次。Fluman等人表明，这种减慢翻译的方式--不同于真核生物所用的方式--有助于确保膜蛋白在大肠杆菌中正确折叠。杆菌虽然这些暂停经常发生，主要是在膜蛋白翻译的早期阶段，但已知在其他mRNA中存在许多其他翻译暂停位点。下一个挑战是了解这些其他暂停位点的功能，以及它们如何与其他机制一起调节细胞内的核糖体翻译。DOI：http://dx.doi.org/10.7554/eLife.03440.002网站
In all living organisms, ribosomes translating membrane proteins are targeted to membrane translocons early in translation, by the ubiquitous signal recognition particle (SRP) system. In eukaryotes, the SRP Alu domain arrests translation elongation of membrane proteins until targeting is complete. Curiously, however, the Alu domain is lacking in most eubacteria. In this study, by analyzing genome-wide data on translation rates, we identified a potential compensatory mechanism in E. coli that serves to slow down the translation during membrane protein targeting. The underlying mechanism is likely programmed into the coding sequence, where Shine–Dalgarno-like elements trigger elongation pauses at strategic positions during the early stages of translation. We provide experimental evidence that slow translation during targeting and improves membrane protein production fidelity, as it correlates with better folding of overexpressed membrane proteins. Thus, slow elongation is important for membrane protein targeting in E. coli, which utilizes mechanisms different from the eukaryotic one to control the translation speed. DOI: http://dx.doi.org/10.7554/eLife.03440.001 Proteins are built as long chain-like molecules. First, a length of DNA is copied to make a messenger RNA (or mRNA) molecule, which then binds to a large molecular complex called a ribosome. The ribosome reads and translates the code in the mRNA sequence to build a protein chain, which then folds into a specific three-dimensional shape to allow the protein to perform its function. Many proteins also need to be targeted to the right location within the cell in order to carry out their role. Some proteins have to be inserted into the membranes of cells and these proteins are directed, as they are being built, to the membrane by another molecular complex called the signal recognition particle (or SRP for short). The SRP binds to the new protein as it emerges from the ribosome and helps to direct it to the membrane. To make sure that membrane proteins fold correctly, their translation is paused whilst the protein is being targeted to the membrane. Plants, animals, and other eukaryotes do this via a unique part of the SRP complex that only allows the translation to continue once the translating ribosome has been brought close to the membrane. However, most bacteria lack this part of the SRP complex, and yet they are still able to accurately insert new, correctly folded, proteins into their membranes. This suggests that an alternative mechanism must exist in bacterial cells. Fluman et al. looked at an existing data set that had measured how many ribosomes are found at different points along the length of mRNA molecules at any given time in the bacterium E. coli. If ribosomes are consistently found at specific sites in given mRNA molecules, it suggests that these are the sites where a pause in translation occurs. Specific short mRNA sequences—that bind to a ribosome and hold it in place—are often found in these pause sites. These sequences are similar to another sequence, called the Shine–Dalgarno sequence that is often also found at the very start of an mRNA molecule, where it functions to recruit a ribosome and begin the translation process. Fluman et al. reveal that mRNAs of membrane proteins contained these similar sequences early on in their coding region. Some looked likely to pause the translation before the newly formed protein chain emerged from the ribosome, which could give the ribosome time to be targeted to the membrane. Other Shine–Dalgarno-like sequences were found slightly later on in the mRNA molecules for protein chains that span back-and-forth through the membrane several times. Fluman et al. show that slowing translation in this manner—which is different to that used by eukaryotes—helps to ensure that membrane proteins are folded correctly in E. coli. Although these pauses occur frequently, mainly in the early stages of the translation of membrane proteins, there are many other translation pause sites that are known to exist in other mRNAs. The next challenge is to understand the function of these other pause sites, and how they work together with other mechanisms to regulate translating ribosomes inside cells. DOI: http://dx.doi.org/10.7554/eLife.03440.002