Trimethoprim, a preferred treatment for urinary tract infections, is becoming obsolete owing to the rapid dissemination of resistant E. coli. Although direct resistance mechanisms such as overexpression of a mutant FolA and dfr enzymes are well characterized, associated alterations that drive or sustain resistance are unknown. We identify the repertoire of resistance-associated perturbations by constructing and interrogating a transcriptome-integrated functional interactome. From the cross talk between perturbations in stress-response and metabolic pathways, we identify the critical dependence on serine hydroxymethyltransferase (GlyA) as an emergent vulnerability. Through its deletion, we demonstrate that GlyA is necessary to sustain high levels of resistance in both laboratory-evolved resistant E. coli and a multidrug-resistant clinical isolate. Through comparative evolution, we show that the absence of GlyA activity decelerates the acquisition of resistance in E. coli. Put together, our results identify GlyA as a promising target, providing a basis for the rational design of drug combinations.

甲氧苄氨嘧啶，一种用于尿路感染的首选治疗方法，由于耐药性大肠杆菌的快速传播而变得过时。虽然直接抗性机制例如突变FolA和过表达DFR酶具有很好的特征，尚不清楚驱动或维持抗性的相关变化。我们通过构建和询问转录组整合的功能性相互作用组来鉴定与抗药性相关的摄动库。从应激反应和代谢途径中的干扰之间的相互影响，我们确定了对丝氨酸羟甲基转移酶（GlyA）的关键依赖性是一种紧急情况。通过删除它，我们证明了GlyA在维持实验室进化的抗性大肠杆菌和多药耐药性临床分离株中维持高水平抗药性方面是必需的。通过比较进化，我们表明，缺乏GlyA活性会降低大肠杆菌中抗药性的获得。综上所述，我们的研究结果确定GlyA是有前途的靶标，为合理设计药物组合提供了基础。