title: A genetic strategy to identify targets for the development of drugs that prevent bacterial persistence.
abstract: Antibacterial drug development suffers from a paucity of targets whose inhibition kills replicating and nonreplicating bacteria. The latter include phenotypically dormant cells, known as persisters, which are tolerant to many antibiotics and often contribute to failure in the treatment of chronic infections. This is nowhere more apparent than in tuberculosis caused by Mycobacterium tuberculosis, a pathogen that tolerates many antibiotics once it ceases to replicate. We developed a strategy to identify proteins that Mycobacterium tuberculosis requires to both grow and persist and whose inhibition has the potential to prevent drug tolerance and persister formation. This strategy is based on a tunable dual-control genetic switch that provides a regulatory range spanning three orders of magnitude, quickly depletes proteins in both replicating and nonreplicating mycobacteria, and exhibits increased robustness to phenotypic reversion. Using this switch, we demonstrated that depletion of the nicotinamide adenine dinucleotide synthetase (NadE) rapidly killed Mycobacterium tuberculosis under conditions of standard growth and nonreplicative persistence induced by oxygen and nutrient limitation as well as during the acute and chronic phases of infection in mice. These findings establish the dual-control switch as a robust tool with which to probe the essentiality of Mycobacterium tuberculosis proteins under different conditions, including those that induce antibiotic tolerance, and NadE as a target with the potential to shorten current tuberculosis chemotherapies.
authors.affiliation: Department of Microbiology and Immunology, Weill Cornell Medical College, New York, NY 10065.
authors.forename: Jee-Hyun
authors.initials: JH
authors.lastname: Kim
authors.forename: Kathryn M
authors.initials: KM
authors.lastname: O'Brien
authors.forename: Ritu
authors.initials: R
authors.lastname: Sharma
authors.forename: Helena I M
authors.initials: HI
authors.lastname: Boshoff
authors.forename: German
authors.initials: G
authors.lastname: Rehren
authors.forename: Sumit
authors.initials: S
authors.lastname: Chakraborty
authors.forename: Joshua B
authors.initials: JB
authors.lastname: Wallach
authors.forename: Mercedes
authors.initials: M
authors.lastname: Monteleone
authors.forename: Daniel J
authors.initials: DJ
authors.lastname: Wilson
authors.forename: Courtney C
authors.initials: CC
authors.lastname: Aldrich
authors.forename: Clifton E
authors.initials: CE
authors.lastname: Barry
authors.forename: Kyu Y
authors.initials: KY
authors.lastname: Rhee
authors.forename: Sabine
authors.initials: S
authors.lastname: Ehrt
authors.forename: Dirk
authors.initials: D
authors.lastname: Schnappinger
mesh_terms: D019732:Amide Synthases / Q000037:antagonists & inhibitors*; D000818:Animals; D002352:Carrier Proteins; D055808:Drug Discovery / Q000379:methods*; D004361:Drug Tolerance / Q000502:physiology*; D029968:Escherichia coli Proteins; D015971:Gene Expression Regulation, Enzymologic / Q000502:physiology*; D005818:Genetic Engineering / Q000379:methods; D008156:Luciferases; D051379:Mice; D009169:Mycobacterium tuberculosis / Q000187:drug effects* / Q000254:growth & development; D014376:Tuberculosis / Q000517:prevention & control*
chemical_list: D002352:Carrier Proteins; D029968:Escherichia coli Proteins; C477700:sspB protein, E coli; D008156:Luciferases; D019732:Amide Synthases; C038479:NAD+ synthase