Polymyxin B lethality requires energy-dependent outer membrane disruption

Research data underpinning publication <b>Polymyxin B lethality requires energy-dependent outer membrane disruption</b>, accepted for publication at Nature Microbiology (2025), and manuscript deposited at https://www.biorxiv.org/content/10.1101/2025.04.16.649083v1.Polymyxin antibiotics target lipopolysaccharide (LPS) in both membranes of the bacterial cell envelope, leading to bacterial killing through a mechanism that remains poorly understood. Here, we demonstrate that metabolic activity is essential for polymyxin lethality and leverage this insight to determine its mode of action. Polymyxin B (PmB) efficiently killed exponential phase <i>E. coli</i> but was unable to eliminate stationary phase cells unless a carbon source was available. Antibiotic lethality correlated with surface protrusions, LPS loss from the outer membrane (OM), and a corresponding reduction in barrier function, processes that required LPS synthesis and transport, but were blocked by the MCR-1 polymyxin resistance determinant.<i> </i>While the energy-dependent OM disruption was not directly lethal, it facilitated PmB access to the inner membrane (IM), which the antibiotic permeabilised in an energy-independent manner, leading to cell death. This work reveals how metabolic inactivity confers tolerance of a clinically important, membrane-targeting antibiotic, leading to new insight into mechanism of action.Data include microscopy data (in generic image format or if not, accessible by open-source software, https://gwyddion.net/) and spreadsheets with tabulated data for plots shown in figures (quantification of cell death, fluorescence, etc.).

本研究数据支撑已被《自然-微生物学》（Nature Microbiology，2025年已接收）录用的论文**《多粘菌素B的致死作用依赖能量依赖性外膜破坏》**，预印本已上传至https://www.biorxiv.org/content/10.1101/2025.04.16.649083v1。多粘菌素类抗生素可靶向细菌细胞被膜两层膜中的脂多糖（lipopolysaccharide, LPS），但其介导细菌杀伤的具体机制仍不甚明晰。本研究证实代谢活性是多粘菌素发挥致死效应的必要条件，并依托这一发现解析其作用模式。实验显示，多粘菌素B（Polymyxin B, PmB）可高效杀灭对数生长期大肠杆菌（E. coli），但无法清除静止期细胞，除非补充碳源。抗生素的致死效应与细胞表面突起、外膜（outer membrane, OM）脂多糖丢失以及相应的屏障功能降低相关，上述过程依赖脂多糖的合成与转运，但可被MCR-1多粘菌素耐药决定因子阻断。尽管能量依赖性的外膜破坏本身并不直接致死，但它可促进PmB进入内膜（inner membrane, IM），而抗生素可通过不依赖能量的方式使内膜通透，最终引发细胞死亡。本研究揭示了代谢失活如何使细菌对临床重要的膜靶向抗生素产生耐受性，为解析其作用机制提供了新视角。本数据集包含显微镜成像数据（通用图像格式，或可通过开源软件Gwyddion（https://gwyddion.net/）读取），以及包含论文各图表制表数据的电子表格（涵盖细胞死亡量化、荧光信号量化等内容）。