Essential Role of the ESX-5 Secretion System in Outer Membrane Permeability of Pathogenic Mycobacteria

作者：Louis S. Ates, Roy Ummels, Susanna Commandeur, Robert van de Weerd, Marion Sparrius, Eveline M. Weerdenburg, Marina Alber, Rainer Kalscheuer, Sander R. Piersma, Abdallah M. Abdallah, Moataz Abd El Ghany, Alyaa M. Abdel‐Haleem, Arnab Pain, Connie R. Jiménez, Wilbert Bitter, Edith N. G. Houben · 发表于：PLoS Genetics · 年份：2015 · DOI：10.1371/journal.pgen.1005190 · 被引用次数：196 · 研究领域：Mycobacterium research and diagnosis、Tuberculosis Research and Epidemiology、Antibiotic Resistance in Bacteria

Mycobacteria possess different type VII secretion (T7S) systems to secrete proteins across their unusual cell envelope. One of these systems, ESX-5, is only present in slow-growing mycobacteria and responsible for the secretion of multiple substrates. However, the role of ESX-5 substrates in growth and/or virulence is largely unknown. In this study, we show that esx-5 is essential for growth of both Mycobacterium marinum and Mycobacterium bovis. Remarkably, this essentiality can be rescued by increasing the permeability of the outer membrane, either by altering its lipid composition or by the introduction of the heterologous porin MspA. Mutagenesis of the first nucleotide-binding domain of the membrane ATPase EccC5 prevented both ESX-5-dependent secretion and bacterial growth, but did not affect ESX-5 complex assembly. This suggests that the rescuing effect is not due to pores formed by the ESX-5 membrane complex, but caused by ESX-5 activity. Subsequent proteomic analysis to identify crucial ESX-5 substrates confirmed that all detectable PE and PPE proteins in the cell surface and cell envelope fractions were routed through ESX-5. Additionally, saturated transposon-directed insertion-site sequencing (TraDIS) was applied to both wild-type M. marinum cells and cells expressing mspA to identify genes that are not essential anymore in the presence of MspA. This analysis confirmed the importance of esx-5, but we could not identify essential ESX-5 substrates, indicating that multi...