The conserved σD envelope stress response monitors multiple aspects of envelope integrity in corynebacteria

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 {sigma}D envelope stress response in porin export and identified MarPas the site-1 protease, respectively, that activate the response by cleaving the membrane-embedded anti-sigma factor. A reporter system revealed that the {sigma}D 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 {sigma}D regulon in protein export to the MM. AUTHOR SUMMARYBacteria within the Mycobacteriales order, which includes the pathogen Mycobacterium tuberculosis, have a unique multilayered cell surface architecture. How they sense and respond to defects in the construction of this distinct envelope to maintain homeostasis remains poorly understood. Here, we used the model organism Corynebacterium glutamicum to reveal that the conserved {sigma}D envelope stress response pathway responds to two distinct signals resulting from defects in different envelope layers. The results provide new insight into how members of the Mycobacteriales monitor the integrity of their surface and pave the way for the identification of novel vulnerabilities in envelope biogenesis that may be useful for targeting with antibiotics.