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Mycobacterium tuberculosis growth following aerobic expression of the DosR regulon.

ABSTRACT: The Mycobacterium tuberculosis regulator DosR is induced by multiple stimuli including hypoxia, nitric oxide and redox stress. Overlap of these stimuli with conditions thought to promote latency in infected patients fuels a model in which DosR regulon expression is correlated with bacteriostasis in vitro and a proxy for latency in vivo. Here, we find that inducing the DosR regulon to wildtype levels in aerobic, replicating M. tuberculosis does not alter bacterial growth kinetics. We conclude that DosR regulon expression alone is insufficient for bacterial latency, but rather is expressed during a range of growth states in a dynamic environment.

SUBMITTER: Minch K

PROVIDER: S-EPMC3338750 | biostudies-literature | 2012

REPOSITORIES: biostudies-literature

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Mycobacterium tuberculosis growth following aerobic expression of the DosR regulon.

Minch Kyle K Rustad Tige T Sherman David R DR

PloS one 20120427 4

The Mycobacterium tuberculosis regulator DosR is induced by multiple stimuli including hypoxia, nitric oxide and redox stress. Overlap of these stimuli with conditions thought to promote latency in infected patients fuels a model in which DosR regulon expression is correlated with bacteriostasis in vitro and a proxy for latency in vivo. Here, we find that inducing the DosR regulon to wildtype levels in aerobic, replicating M. tuberculosis does not alter bacterial growth kinetics. We conclude tha ...[more]

PMID: 22558276

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Project description:The DosR regulon, a set of 48 genes normally expressed in Mycobacterium tuberculosis under conditions that inhibit aerobic respiration, is controlled via the DosR-DosS/DosT two-component system. While the regulon requires induction in most M. tuberculosis isolates, for members of the Beijing lineage, its expression is uncoupled from the need for signaling. In our attempts to understand the mechanistic basis for this uncoupling in the Beijing background, we previously reported the identification of two synonymous single-nucleotide polymorphisms (SNPs) within the adjacent Rv3134c gene. In the present study, we have interrogated the impact of these SNPs on dosR expression in wild-type strains, as well as a range of dosR-dosS-dosT mutants, for both Beijing and non-Beijing M. tuberculosis backgrounds. In this manner, we have unequivocally determined that the C601T dosR promoter SNP is the sole requirement for the dramatic shift in the pattern of DosR regulon expression seen in this globally important lineage. Interestingly, we also show that DosT is completely nonfunctional within these strains. Thus, a complex series of evolutionary steps has led to the present-day Beijing DosR phenotype that, in turn, potentially confers a fitness advantage in the face of some form of host-associated selective pressure. Mycobacterium tuberculosis strains of the Beijing lineage have been described as being of enhanced virulence compared to other lineages, and in certain regions, they are associated with the dramatic spread of multidrug-resistant tuberculosis (TB). In terms of trying to understand the functional basis for these broad epidemiological phenomena, it is interesting that, in contrast to the other major lineages, the Beijing strains all constitutively overexpress members of the DosR regulon. Here, we identify the mutational events that led to the evolution of this unique phenotype. In addition, our work highlights the fact that important phenotypic differences exist between distinct M. tuberculosis lineages, with the potential to impact the efficacy of diagnosis, vaccination, and treatment programs.

Project description:Mycobacterium tuberculosis is a strict aerobe capable of prolonged survival in the absence of oxygen. We investigated the ability of anaerobic M. tuberculosis to counter challenges to internal pH homeostasis in the absence of aerobic respiration, the primary mechanism of proton efflux for aerobic bacilli. Anaerobic M. tuberculosis populations were markedly impaired for survival under a mildly acidic pH relative to standard culture conditions. An acidic environmental pH greatly increased the susceptibilities of anaerobic bacilli to the collapse of the proton motive force by protonophores, to antimicrobial compounds that target entry into the electron transport system, and to small organic acids with uncoupling activity. However, anaerobic bacilli exhibited high tolerance against these challenges at a near-neutral pH. At a slightly alkaline pH, which was near the optimum intracellular pH, the addition of protonophores even improved the long-term survival of bacilli. Although anaerobic M. tuberculosis bacilli under acidic conditions maintained 40% lower ATP levels than those of bacilli under standard culture conditions, ATP loss alone could not explain the drop in viability. Protonophores decreased ATP levels by more than 90% regardless of the extracellular pH but were bactericidal only under acidic conditions, indicating that anaerobic bacilli could survive an extreme ATP loss provided that the external pH was within viable intracellular parameters. Acidic conditions drastically decreased the anaerobic survival of a DosR mutant, while an alkaline environment improved the survival of the DosR mutant. Together, these findings indicate that intracellular acidification is a primary challenge for the survival of anaerobic M. tuberculosis and that the DosR regulon plays a critical role in sustaining internal pH homeostasis.IMPORTANCE During infection, M. tuberculosis bacilli are prevalent in environments largely devoid of oxygen, yet the factors that influence the survival of these severely growth-limited and metabolically limited bacilli remain poorly understood. We determined how anaerobic bacilli respond to fluctuations in environmental pH and observed that these bacilli were highly susceptible to stresses that promoted internal acidic stress, whereas conditions that promoted an alkaline internal pH promoted long-term survival even during severe ATP depletion. The DosR regulon, a major regulator of general hypoxic stress, played an important role in maintaining internal pH homeostasis under anaerobic conditions. Together, these findings indicate that in the absence of aerobic respiration, protection from internal acidification is crucial for long-term M. tuberculosis survival.

Dataset Information

Mycobacterium tuberculosis growth following aerobic expression of the DosR regulon.

Publications

Mycobacterium tuberculosis growth following aerobic expression of the DosR regulon.

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