Functional analysis of stress-dependent RNA-enzyme interactions

应激依赖性RNA-酶相互作用的功能分析

• 批准号: BB/S017747/1
• 负责人: Andre Gerber
• 金额: $ 63.53万
• 依托单位: University of Surrey
• 依托单位国家: 英国
• 项目类别: Research Grant
• 财政年份: 2019
• 资助国家: 英国
• 起止时间: 2019 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=BB%2FS017747%2F1
• 关键词: Functional; analysis; stress; dependent; RNA

Cells have to immediately respond to changing environmental conditions. In particular, unicellular organisms such as baker's yeast Saccharomyces cerevisae must immediately react to cope with an altered environment. Since yeast is also important for production of food, wine and other goods, there is great interest to understand how the cells can deal with stress to optimise industrial applications. Specifically, the response to oxidative stress, which is imposed by the imbalance in the levels of so-called reactive oxygen species (ROS) generated during normal aerobic metabolism but particularly through exposure to certain toxic chemicals or irradiation, is of wider interest because it is connected to the development of diverse pathological processes in humans, such as neurodegenerative disorders, diabetes, arthritis and cancer. Cells have developed a variety of mechanisms to adapt to stress, including immediate changes in the levels of key metabolites and by altering gene expression. Regarding the latter, it has become particularly recognised that stress response is considerably affecting the synthesis of new proteins, and the stability of RNAs. RNA represents an essential intermediate step in gene expression, where DNA is copied into RNA to serve as template for protein synthesis. Importantly, RNA is never naked in cells but covered by a host of proteins, so-called RNA-binding proteins (RBPs). These proteins can remove or rearrange parts of RNA, store, or deliver it to particular locations within the cell, and ultimately degrade it. They also control when and how messenger RNAs (mRNAs; refers to the class of RNAs that encode proteins) are translated into proteins. Due to the tremendous importance of RPPs in gene expression control, we and other researchers have developed an experimental approach to catalogue all of the RBPs that interact with mRNAs in cells. Besides detecting many of the previously known RPBs, it was found that proteins with other well-established functions, such as enzymes, could interact with mRNAs (enzymes are proteins that perform essential chemical reactions in the cell). Furthermore, we have investigated which RBPs change mRNA associations upon oxidative stress in yeast. Interestingly, we found that many of the ones that changed mRNA associations were metabolic enzymes acting in carbon metabolisms, which is central for energy production and storage in cells; and provides building blocks for the synthesis of complex biological molecules. Therefore, we wish to understand the function of these "enigmatic" enzyme-RNA interactions and whether they could play a role in coping with oxidative stress. We will first test whether the observed changes in RNA-enzyme interactions are specific to oxidative stress or apply to other stress conditions as well. We will then comprehensively identify the RNA targets for selected enzymes acting in central carbon metabolisms, and investigate where and how they bind to RNAs. Finally, we wish to explore whether the stress-dependent interactions with RNA affects their fate, or conversely, whether it modulates enzymatic activity and plays a role in the cell's adaptation to stress. With our research, we expect to discover previously unrecognised links between RNA regulation, metabolism and cellular stress response. If so, this knowledge will likely have impact on diverse important aspects of our society, from food-production towards a better understanding of components that contribute to today's most prominent diseases including cancer.

细胞必须立即对变化的环境条件做出反应。特别是，单细胞生物，如面包酵母酿酒酵母，必须立即作出反应，以科普改变的环境。由于酵母对食品，葡萄酒和其他商品的生产也很重要，因此了解细胞如何应对压力以优化工业应用具有很大的兴趣。具体而言，对氧化应激的反应是由正常有氧代谢过程中产生的所谓活性氧（ROS）水平的不平衡引起的，但特别是通过暴露于某些有毒化学物质或辐射，这一反应受到了更广泛的关注，因为它与人类多种病理过程的发展有关，如神经退行性疾病、糖尿病、关节炎和癌症。细胞已经发展出多种机制来适应压力，包括关键代谢物水平的立即变化和改变基因表达。关于后者，已经特别认识到应激反应显著影响新蛋白质的合成和RNA的稳定性。RNA代表基因表达中的重要中间步骤，其中DNA被复制到RNA中作为蛋白质合成的模板。重要的是，RNA在细胞中从来不是裸露的，而是被许多蛋白质，即所谓的RNA结合蛋白（RBP）所覆盖。这些蛋白质可以移除或重新排列RNA的一部分，将其储存或运送到细胞内的特定位置，并最终降解它。它们还控制信使RNA（mRNA;指编码蛋白质的RNA类）何时以及如何翻译成蛋白质。由于RPPs在基因表达控制中的巨大重要性，我们和其他研究人员已经开发出一种实验方法来对细胞中与mRNA相互作用的所有RPPs进行分类。除了检测许多先前已知的RPB外，还发现具有其他既定功能的蛋白质，如酶，可以与mRNA相互作用（酶是在细胞中进行基本化学反应的蛋白质）。此外，我们已经研究了哪些RBP改变酵母中氧化应激后的mRNA相关性。有趣的是，我们发现许多改变mRNA关联的是在碳代谢中起作用的代谢酶，碳代谢是细胞能量产生和储存的核心;并为合成复杂的生物分子提供了基础。因此，我们希望了解这些“神秘”的酶-RNA相互作用的功能，以及它们是否可以在应对氧化应激中发挥作用。我们将首先测试所观察到的RNA-酶相互作用的变化是否特异于氧化应激或也适用于其他应激条件。然后，我们将全面确定在中心碳代谢中起作用的选定酶的RNA靶点，并研究它们在哪里以及如何与RNA结合。最后，我们希望探索与RNA的应激依赖性相互作用是否影响它们的命运，或者相反，它是否调节酶活性并在细胞适应应激中发挥作用。通过我们的研究，我们希望发现RNA调节，代谢和细胞应激反应之间以前未被认识到的联系。如果是这样的话，这些知识可能会对我们社会的各个重要方面产生影响，从粮食生产到更好地了解导致当今最突出的疾病（包括癌症）的成分。

项目成果

Biochemical approach for isolation of polyadenylated RNAs with bound proteins from yeast.

• DOI: 10.1016/j.xpro.2021.100929
• 发表时间: 2021-12-17
• 期刊: STAR protocols
• 作者: Matia-González AM;Jabre I;Gerber AP
• 通讯作者: Gerber AP

Oxidative stress induces coordinated remodeling of RNA-enzyme interactions.

• DOI: 10.1016/j.isci.2021.102753
• 发表时间: 2021-07-23
• 期刊: iScience
• 影响因子: 5.8
• 作者: Matia-González AM;Jabre I;Laing EE;Gerber AP
• 通讯作者: Gerber AP

RNA-Centric Approaches to Profile the RNA-Protein Interaction Landscape on Selected RNAs.

• DOI: 10.3390/ncrna7010011
• 发表时间: 2021-02-15
• 期刊: Non-coding RNA
• 影响因子: 4.3
• 作者: Gerber AP