Identification of new components of the RipC-FtsEX cell separation pathway of Corynebacterineae

• Published in: PLoS genetics 2019-08, Vol.15 (8), p.e1008284
• Main Authors: Lim, Hoong Chuin, Sher, Joel W, Rodriguez-Rivera, Frances P, Fumeaux, Coralie, Bertozzi, Carolyn R, Bernhardt, Thomas G
• Format: Article
• Language: English
• Subjects: Acids; Active biological transport; Analysis; Antitubercular Agents - pharmacology; Arabinogalactan; Bacteria; Bacterial Outer Membrane - drug effects; Bacterial Outer Membrane - metabolism; Bacterial Proteins - genetics; Bacterial Proteins - metabolism; Biology and Life Sciences; Biosynthesis; Biosynthetic Pathways - drug effects; Cell division; Cell Division - genetics; Cell separation; Cell walls; Corynebacteria; Corynebacterium glutamicum - drug effects; Corynebacterium glutamicum - physiology; Cytoplasmic membranes; DNA Transposable Elements - genetics; Drug Resistance, Bacterial - genetics; E coli; Ethambutol; Ethambutol - pharmacology; Evolution; Galactans - biosynthesis; Genes; Genetic aspects; Genetic code; Genetic Loci; Genomes; Hypersensitivity; Identification and classification; Integrated software; Medical schools; Microscopy; Mutation; Mycolic acids; Mycolic Acids - metabolism; Organisms; Peptidoglycan - metabolism; Peptidoglycans; Polysaccharides; Proteins; Research and Analysis Methods; Streptococcus infections; Transposons; Tuberculosis
• ISSN: 1553-7404; 1553-7390; 1553-7404
• DOI: 10.1371/journal.pgen.1008284

Several important human pathogens are represented in the Corynebacterineae suborder, including Mycobacterium tuberculosis and Corynebacterium diphtheriae. These bacteria are surrounded by a multilayered cell envelope composed of a cytoplasmic membrane, a peptidoglycan (PG) cell wall, a second polysaccharide layer called the arabinogalactan (AG), and finally an outer membrane-like layer made of mycolic acids. Several anti-tuberculosis drugs target the biogenesis of this complex envelope, but their efficacy is declining due to resistance. New therapies are therefore needed to treat diseases caused by these organisms, and a better understanding of the mechanisms of envelope assembly should aid in their discovery. To this end, we generated the first high-density library of transposon insertion mutants in the model organism C. glutamicum. Transposon-sequencing was then used to define its essential gene set and identify loci that, when inactivated, confer hypersensitivity to ethambutol (EMB), a drug that targets AG biogenesis. Among the EMBs loci were genes encoding RipC and the FtsEX complex, a PG cleaving enzyme required for proper cell division and its predicted regulator, respectively. Inactivation of the conserved steAB genes (cgp_1603-1604) was also found to confer EMB hypersensitivity and cell division defects. A combination of quantitative microscopy, mutational analysis, and interaction studies indicate that SteA and SteB form a complex that localizes to the cytokinetic ring to promote cell separation by RipC-FtsEX and may coordinate its PG remodeling activity with the biogenesis of other envelope layers during cell division.