Institute of Molecular Biology, Academia Sinica, Taipei, 11529, Taiwan.
Department of Immunology, Microbiology and Parasitology, University of the Basque Country UPV/EHU, Leioa, Spain.
Mol Genet Genomics. 2019 Oct;294(5):1359-1371. doi: 10.1007/s00438-019-01597-1. Epub 2019 Jul 30.
Previous studies revealed important roles of small RNAs (sRNAs) in regulation of bacterial metabolism, stress responses and virulence. However, only a minor fraction of sRNAs is well characterized with respect to the spectra of their targets, conditional expression profiles and actual mechanisms they use to regulate gene expression to control particular biological pathways. To learn more about the specific contribution of sRNAs to the global regulatory network controlling the Escherichia coli central carbon metabolism (CCM), we employed microarray analysis and compared transcriptome profiles of E. coli cells grown on two alternative minimal media supplemented with either pyruvate or glucose, respectively. Microarray analysis revealed that utilization of these alternative carbon sources led to profound differences in gene expression affecting all major gene clusters associated with CCM as well as expression of several known (CyaR, RyhB, GcvB and RyeA) and putative (C0652) sRNAs. To assess the impact of transcriptional reprogramming of gene expression on E. coli protein abundance, we also employed two-dimensional protein gel electrophoresis. Our experimental data made it possible to determine the major pathways for pyruvate assimilation when it is used as a sole carbon source and reveal the impact of other key processes (i.e., energy production, molecular transport and cell resistance to stress) associated with the CCM in E. coli. Moreover, some of these processes were apparently controlled by GcvB, RyhB and CyaR at the post-transcriptional level, thus indicating the complexity and interconnection of the regulatory networks that control CCM in bacteria.
先前的研究表明,小 RNA(sRNA)在调节细菌代谢、应激反应和毒力方面发挥着重要作用。然而,只有一小部分 sRNA 就其靶标的谱、条件表达谱以及它们用于调节基因表达以控制特定生物途径的实际机制得到了很好的描述。为了更多地了解 sRNA 对控制大肠杆菌中心碳代谢(CCM)的全局调控网络的特定贡献,我们采用了微阵列分析,并比较了在两种不同的基础培养基上生长的大肠杆菌细胞的转录组图谱,培养基分别补充了丙酮酸或葡萄糖。微阵列分析显示,这些替代碳源的利用导致了基因表达的深刻差异,影响了与 CCM 相关的所有主要基因簇,以及几个已知(CyaR、RyhB、GcvB 和 RyeA)和推定(C0652)sRNA 的表达。为了评估基因表达转录重编程对大肠杆菌蛋白质丰度的影响,我们还采用了二维蛋白质凝胶电泳。我们的实验数据使我们能够确定当丙酮酸被用作唯一碳源时丙酮酸同化的主要途径,并揭示了与大肠杆菌 CCM 相关的其他关键过程(即能量产生、分子运输和细胞应激抗性)的影响。此外,其中一些过程显然在转录后水平受到 GcvB、RyhB 和 CyaR 的控制,这表明了控制细菌 CCM 的调控网络的复杂性和相互关联性。