RNA-dependent proteome solubility maintenance in <i>Escherichia coli</i> lysates analysed by quantitative mass spectrometry: Proteomic characterization in terms of isoelectric point, structural disorder, functional hub, and chaperone network

Protein aggregation, a consequence of misfolding and impaired proteostasis, can lead to cellular malfunctions such as various proteinopathies. The mechanisms protecting proteins from aggregation in complex cellular environments have long been investigated, often from a protein-centric viewpoint. However, our study provides insights into a crucial, yet overlooked actor: RNA. We found that depleting RNAs from Escherichia coli lysates induces global protein aggregation. Our quantitative mass spectrometry analysis identified over 900 statistically significant proteins from the Escherichia coli proteome whose solubility depends on RNAs. Proteome-wide characterization showed that the RNA dependency is particularly enriched among acidic proteins, intrinsically disordered proteins, and structural hub proteins. Moreover, we observed distinct differences in RNA-binding mode and Gene Ontology categories between RNA-dependent acidic and basic proteins. Notably, the solubility of key molecular chaperones [Trigger factor, DnaJ, and GroES] is largely dependent on RNAs, suggesting a yet-to-be-explored hierarchical relationship between RNA-based chaperone (termed as chaperna) and protein-based chaperones, both of which constitute the whole chaperone network. These findings provide new insights into the RNA-centric role in maintaining healthy proteome solubility in vivo, where proteins associate with a variety of RNAs, either stably or transiently.蛋白质聚集是蛋白质错误折叠与蛋白稳态受损的必然结果，可引发多种细胞功能紊乱，例如各类蛋白质病。长期以来，针对复杂细胞环境中保护蛋白质免受聚集的机制的研究多以蛋白质为中心展开。然而本研究揭示了一类长期被忽视的关键参与者：核糖核酸（RNA）。我们发现，从大肠杆菌（Escherichia coli）裂解液中剔除核糖核酸会诱发全局性蛋白质聚集。通过定量质谱分析，我们从大肠杆菌蛋白质组中鉴定出900余种具有统计学显著性的、可溶性依赖于核糖核酸的蛋白质。全蛋白质组表征分析显示，核糖核酸依赖性在酸性蛋白质、内在无序蛋白质以及结构枢纽蛋白中显著富集。此外，我们还观察到，依赖核糖核酸的酸性蛋白质与碱性蛋白质在核糖核酸结合模式以及基因本体（Gene Ontology）分类上存在显著差异。值得注意的是，关键分子伴侣[触发因子（Trigger factor）、DnaJ及GroES]的可溶性在很大程度上依赖于核糖核酸，这提示基于核糖核酸的分子伴侣（命名为chaperna）与蛋白质类分子伴侣之间存在尚未被阐明的层级调控关系，二者共同构成完整的分子伴侣网络。上述研究结果为阐明以核糖核酸为中心的、维持活体中健康蛋白质组可溶性的机制提供了全新视角——在活体环境内，蛋白质会以稳定或瞬时的方式与多种核糖核酸结合。