Hart Elizabeth M, Lyerly Evan, Bernhardt Thomas G
Department of Microbiology, Harvard Medical School, Boston, Massachusetts, United States of America.
Howard Hughes Medical Institute, Harvard Medical School, Boston, Massachusetts, United States of America.
PLoS Genet. 2024 Jun 3;20(6):e1011127. doi: 10.1371/journal.pgen.1011127. eCollection 2024 Jun.
The cell envelope fortifies bacterial cells against antibiotics and other insults. Species in the Mycobacteriales order have a complex envelope that includes an outer layer of mycolic acids called the mycomembrane (MM) and a cell wall composed of peptidoglycan and arabinogalactan. This envelope architecture is unique among bacteria and contributes significantly to the virulence of pathogenic Mycobacteriales like Mycobacterium tuberculosis. Characterization of pathways that govern envelope biogenesis in these organisms is therefore critical in understanding their biology and for identifying new antibiotic targets. To better understand MM biogenesis, we developed a cell sorting-based screen for mutants defective in the surface exposure of a porin normally embedded in the MM of the model organism Corynebacterium glutamicum. The results revealed a requirement for the conserved σD envelope stress response in porin export and identified MarP as the site-1 protease, respectively, that activate the response by cleaving the membrane-embedded anti-sigma factor. A reporter system revealed that the σD pathway responds to defects in mycolic acid and arabinogalactan biosynthesis, suggesting that the stress response has the unusual property of being induced by activating signals that arise from defects in the assembly of two distinct envelope layers. Our results thus provide new insights into how C. glutamicum and related bacteria monitor envelope integrity and suggest a potential role for members of the σD regulon in protein export to the MM.
细胞包膜增强了细菌细胞对抗生素和其他损害的能力。分枝杆菌目物种具有复杂的包膜，包括一层称为霉菌膜（MM）的分枝菌酸外层以及由肽聚糖和阿拉伯半乳聚糖组成的细胞壁。这种包膜结构在细菌中是独一无二的，并且对诸如结核分枝杆菌等致病性分枝杆菌目的毒力有重大贡献。因此，表征这些生物体中控制包膜生物合成的途径对于理解它们的生物学特性以及确定新的抗生素靶点至关重要。为了更好地理解MM生物合成，我们开发了一种基于细胞分选的筛选方法，用于筛选在模型生物谷氨酸棒杆菌的MM中正常嵌入的孔蛋白表面暴露有缺陷的突变体。结果分别揭示了孔蛋白输出中对保守的σD包膜应激反应的需求，并确定MarP为1型位点蛋白酶，其通过切割膜嵌入的抗σ因子来激活该反应。一个报告系统显示，σD途径对分枝菌酸和阿拉伯半乳聚糖生物合成中的缺陷有反应，这表明应激反应具有由两个不同包膜层组装缺陷产生的激活信号诱导的不寻常特性。因此，我们的结果为谷氨酸棒杆菌和相关细菌如何监测包膜完整性提供了新的见解，并暗示了σD调节子成员在蛋白质输出到MM中的潜在作用。
