Helmholtz Centre for Infection Research, Inhoffenstraße 7, 38124, Brunswick, Germany.
Technical Chemistry - Life Science, Leibniz University of Hannover, Callinstr. 5, 30167, Hannover, Germany.
Microb Cell Fact. 2022 Apr 21;21(1):67. doi: 10.1186/s12934-022-01787-4.
Escherichia coli adapted to carbon-limiting conditions is generally geared for energy-efficient carbon utilization. This includes also the efficient utilization of glucose, which serves as a source for cellular building blocks as well as energy. Thus, catabolic and anabolic functions are balanced under these conditions to minimize wasteful carbon utilization. Exposure to glucose excess interferes with the fine-tuned coupling of anabolism and catabolism leading to the so-called carbon overflow metabolism noticeable through acetate formation and eventually growth inhibition.
Cellular adaptations towards sudden but timely limited carbon excess conditions were analyzed by exposing slow-growing cells in steady state glucose-limited continuous culture to a single glucose pulse. Concentrations of metabolites as well as time-dependent transcriptome alterations were analyzed and a transcriptional network analysis performed to determine the most relevant transcription and sigma factor combinations which govern these adaptations. Down-regulation of genes related to carbon catabolism is observed mainly at the level of substrate uptake and downstream of pyruvate and not in between in the glycolytic pathway. It is mainly accomplished through the reduced activity of CRP-cAMP and through an increased influence of phosphorylated ArcA. The initiated transcriptomic change is directed towards down-regulation of genes, which contribute to active movement, carbon uptake and catabolic carbon processing, in particular to down-regulation of genes which contribute to efficient energy generation. Long-term changes persisting after glucose depletion and consumption of acetete encompassed reduced expression of genes related to active cell movement and enhanced expression of genes related to acid resistance, in particular acid resistance system 2 (GABA shunt) which can be also considered as an inefficient bypass of the TCA cycle.
Our analysis revealed that the major part of the trancriptomic response towards the glucose pulse is not directed towards enhanced cell proliferation but towards protection against excessive intracellular accumulation of potentially harmful concentration of metabolites including among others energy rich compounds such as ATP. Thus, resources are mainly utilized to cope with "overfeeding" and not for growth including long-lasting changes which may compromise the cells future ability to perform optimally under carbon-limiting conditions (reduced motility and ineffective substrate utilization).
适应碳限制条件的大肠杆菌通常倾向于高效利用碳。这包括有效地利用葡萄糖，葡萄糖既是细胞构建块的来源，也是能量的来源。因此，在这些条件下，分解代谢和合成代谢功能平衡，以最小化浪费的碳利用。暴露于葡萄糖过剩会干扰新陈代谢的合成与分解之间的精细耦合，导致所谓的碳溢出代谢，这种代谢可通过形成乙酸盐和最终的生长抑制来观察到。
通过将缓慢生长的细胞暴露于稳定的葡萄糖限制连续培养中的单个葡萄糖脉冲中，分析了细胞对突然但及时的碳过量条件的适应。分析了代谢物的浓度以及时变转录组变化，并进行了转录网络分析，以确定控制这些适应的最相关的转录和σ因子组合。观察到与碳分解代谢相关的基因下调主要发生在底物摄取水平上，并且在丙酮酸下游，而不是在糖酵解途径中间。这主要是通过 CRP-cAMP 活性降低和磷酸化 ArcA 的影响增加来完成的。启动的转录组变化方向是下调有助于主动运动、碳摄取和碳分解代谢的基因，特别是下调有助于有效能量产生的基因。在葡萄糖耗尽和乙酸消耗后持续存在的长期变化包括与主动细胞运动相关的基因表达减少，以及与酸抗性相关的基因表达增强，特别是与酸抗性系统 2（GABA 分流）相关的基因表达增强，这也可以被认为是 TCA 循环的低效旁路。
我们的分析表明，转录组对葡萄糖脉冲的主要反应不是指向增强细胞增殖，而是指向保护细胞免受潜在有害浓度的代谢物的过度积累，包括能量丰富的化合物如 ATP 等。因此，资源主要用于应对“过度喂养”，而不是用于生长，包括可能损害细胞在碳限制条件下最佳发挥作用的未来能力的持久变化（运动能力降低和底物利用效率降低）。
