title: Mycobacterium tuberculosis has diminished capacity to counteract redox stress induced by elevated levels of endogenous superoxide.
abstract: Mycobacterium tuberculosis (Mtb) has evolved protective and detoxification mechanisms to maintain cytoplasmic redox balance in response to exogenous oxidative stress encountered inside host phagocytes. In contrast, little is known about the dynamic response of this pathogen to endogenous oxidative stress generated within Mtb. Using a noninvasive and specific biosensor of cytoplasmic redox state of Mtb, we for first time discovered a surprisingly high sensitivity of this pathogen to perturbation in redox homeostasis induced by elevated endogenous reactive oxygen species (ROS). We synthesized a series of hydroquinone-based small molecule ROS generators and found that ATD-3169 permeated mycobacteria to reliably enhance endogenous ROS including superoxide radicals. When Mtb strains including multidrug-resistant (MDR) and extensively drug-resistant (XDR) patient isolates were exposed to this compound, a dose-dependent, long-lasting, and irreversible oxidative shift in intramycobacterial redox potential was detected. Dynamic redox potential measurements revealed that Mtb had diminished capacity to restore cytoplasmic redox balance in comparison with Mycobacterium smegmatis (Msm), a fast growing nonpathogenic mycobacterial species. Accordingly, Mtb strains were extremely susceptible to inhibition by ATD-3169 but not Msm, suggesting a functional linkage between dynamic redox changes and survival. Microarray analysis showed major realignment of pathways involved in redox homeostasis, central metabolism, DNA repair, and cell wall lipid biosynthesis in response to ATD-3169, all consistent with enhanced endogenous ROS contributing to lethality induced by this compound. This work provides empirical evidence that the cytoplasmic redox poise of Mtb is uniquely sensitive to manipulation in steady-state endogenous ROS levels, thus revealing the importance of targeting intramycobacterial redox metabolism for controlling TB infection.
authors.affiliation: Department of Microbiology and Cell Biology, Centre for Infectious Disease Research (CIDR), Indian Institute of Science, Bangalore-12, India; International Centre for Genetic Engineering and Biotechnology, New Delhi 67, India.
authors.forename: Priyanka
authors.initials: P
authors.lastname: Tyagi
authors.affiliation: Department of Chemistry, Indian Institute of Science Education and Research, Pune 08, India.
authors.forename: Allimuthu T
authors.initials: AT
authors.lastname: Dharmaraja
authors.affiliation: Department of Microbiology and Cell Biology, Centre for Infectious Disease Research (CIDR), Indian Institute of Science, Bangalore-12, India.
authors.forename: Ashima
authors.initials: A
authors.lastname: Bhaskar
authors.affiliation: Department of Chemistry, Indian Institute of Science Education and Research, Pune 08, India. Electronic address: harinath@iiserpune.ac.in.
authors.forename: Harinath
authors.initials: H
authors.lastname: Chakrapani
authors.affiliation: Department of Microbiology and Cell Biology, Centre for Infectious Disease Research (CIDR), Indian Institute of Science, Bangalore-12, India. Electronic address: asingh@mcbl.iisc.ernet.in.
authors.forename: Amit
authors.initials: A
authors.lastname: Singh
mesh_terms: D000995:Antitubercular Agents / Q000494:pharmacology; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D002991:Clofazimine / Q000494:pharmacology; D045643:DNA Repair Enzymes / Q000235:genetics / Q000378:metabolism; D024881:Drug Resistance, Bacterial; D024901:Drug Resistance, Multiple, Bacterial; D015870:Gene Expression; D015964:Gene Expression Regulation, Bacterial / Q000187:drug effects; D005798:Genes, Bacterial; D008826:Microbial Sensitivity Tests; D020102:Mycobacterium smegmatis / Q000187:drug effects / Q000502:physiology; D009169:Mycobacterium tuberculosis / Q000187:drug effects / Q000502:physiology*; D010084:Oxidation-Reduction; D018384:Oxidative Stress; D013481:Superoxides / Q000378:metabolism*
chemical_list: D000995:Antitubercular Agents; D001426:Bacterial Proteins; D013481:Superoxides; D002991:Clofazimine; D045643:DNA Repair Enzymes
keywords: Drug resistance; Mycobacterium smegmatis; Mycobacterium tuberculosis; Reactive oxygen species (ROS); Redox regulation; Superoxide
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