Table_1_Multiple Small RNAs Interact to Co-regulate Ethanol Tolerance in Zymomonas mobilis.XLSX

sRNAs represent a powerful class of regulators that influences multiple mRNA targets in response to environmental changes. However, very few direct sRNA–sRNA interactions have been deeply studied in any organism. Zymomonas mobilis is a bacterium with unique ethanol-producing metabolic pathways in which multiple small RNAs (sRNAs) have recently been identified, some of which show differential expression in ethanol stress. In this study, we show that two sRNAs (Zms4 and Zms6) are upregulated under ethanol stress and have significant impacts on ethanol tolerance and production in Z. mobilis. We conducted multi-omics analysis (combining transcriptomics and sRNA-immunoprecipitation) to map gene networks under the influence of their regulation. We confirmed that Zms4 and Zms6 bind multiple RNA targets and regulate their expressions, influencing many downstream pathways important to ethanol tolerance and production. In particular, Zms4 and Zms6 interact with each other as well as many other sRNAs, forming a novel sRNA–sRNA direct interaction network. This study thus uncovers a sRNA network that co-orchestrates multiple ethanol related pathways through a diverse set of mRNA targets and a large number of sRNAs. To our knowledge, this study represents one of the largest sRNA–sRNA direct interactions uncovered so far.

sRNA（小核RNA）代表一类强大的调控因子，能够响应环境变化影响多种mRNA靶标。然而，在任一生物体中，sRNA与sRNA之间的直接相互作用研究寥寥无几。Zymomonas mobilis 是一种具有独特乙醇产生代谢途径的细菌，近期在其体内鉴定出多种小RNA（sRNA），其中一些在乙醇胁迫下表现出差异表达。在本研究中，我们揭示了在乙醇胁迫下，两种sRNA（Zms4和Zms6）的表达上调，并对Z. mobilis的乙醇耐受性和生产具有显著影响。我们通过多组学分析（结合转录组学和sRNA免疫沉淀技术）绘制了其调控作用下的基因网络。我们证实Zms4和Zms6能够结合多个RNA靶标并调节其表达，影响众多与乙醇耐受性和生产相关的下游途径。特别是，Zms4和Zms6不仅相互交互，还与其他多种sRNA相互作用，形成了一种新型的sRNA-sRNA直接相互作用网络。因此，本研究揭示了通过多样化的mRNA靶标和大量sRNA协同调控多个乙醇相关途径的sRNA网络。据我们所知，本研究是目前公开报道的最大的sRNA-sRNA直接相互作用研究之一。