Silencing of Oxidative Stress Response in Mycobacterium tuberculosis : Expression Patterns of ahpC in Virulent and Avirulent Strains and Effect of ahpC Inactivation

Abstract

Intracellular pathogens such as Mycobacterium tuberculosis are able to survive in the face of antimicrobial products generated by the host cell in response to infection. The product of the alkyl hydroperoxide reductase gene (ahpC) of M. tuberculosis is thought to be involved in protecting the organism against both oxidative and nitrosative stress encountered within the infected macrophage. Here we report that, contrary to expectations, ahpC expression in virulent strains of M. tuberculosis and Mycobacterium bovis grown in vitro is repressed, often below the level of detection, whereas expression in the avirulent vaccine strain M. bovis BCG is constitutively high. The repression of the ahpC gene of the virulent strains is independent of the naturally occurring lesions of central regulator oxyR. Using a green fluorescence protein vector (gfp)-ahpC reporter construct we present data showing that repression of ahpC of virulent M. tuberculosis also occurred during growth inside macrophages, whereas derepression in BCG was again seen under identical conditions. Inactivation of ahpC on the chromosome of M. tuberculosis by homologous recombination had no effect on its growth during acute infection in mice and did not affect in vitro sensitivity to H 2 O 2 . However, consistent with AhpC function in detoxifying organic peroxides, sensitivity to cumene hydroperoxide exposure was increased in the ahpC::Km r mutant strain. The preservation of a functional ahpC gene in M. tuberculosis in spite of its repression under normal growth conditions suggests that, while AhpC does not play a significant role in establishing infection, it is likely to be important under certain, as yet undefined conditions. This is supported by the observation that repression of ahpC expression in vitro was lifted under conditions of static growth.

The timing of TNF and IFN-γ signaling affects macrophage activation strategies during Mycobacterium tuberculosis infection

“…This model phenomenologically captures several effects of NO/RNIs; for example, g bNO captures a possible dormancy program in Mtb induced by NO (Ohno et al, 2003;Voskuil et al, 2004). The relative insensitivity of Mtb to superoxide and other non-RNI killing mechanisms (Bryk et al, 2000;Ehrt et al, 1997;Nathan & Shiloh, 2000;Ruan et al, 1999;Springer et al, 2001) allows us to omit these effectors, which are more important against other pathogens (Miller & Britigan, 1997). The effect of elevated intracellular iron (represented by g bLIP ) is stimulatory, capturing the effects of irongathering siderophores produced by Mtb (DeVoss et al, 2000).…”

“…NO and some reactive nitrogen intermediates (RNIs) derived from it are effective at killing Mtb in vitro (Yu et al, 1999), but other antimicrobial molecules are not (Bryk et al, 2000;Ehrt et al, 1997;Nathan & Shiloh, 2000;Ruan et al, 1999;Springer et al, 2001). NO or RNIs may also induce a latent phase of the Mtb growth cycle (Couture et al, 1999;Ohno et al, 2003).…”

Oxidation of glutathione peroxidase–deficient red cells by organic peroxides

“…[5][6][7][8][9][10] Although these enzymes are peroxiredoxins and have no structural relationship to eucaryotic GSHPx, they exhibit similar catalytic capacities and are able to reduce cumene peroxide and t-butyl peroxide. These enzymes protect the bacterium against damage by organic peroxides, strengthening their functional similarity to GSHPx.…”

Thiol-Dependent Peroxidases in Mycobacterium tuberculosis Antioxidant Defense

“…However, data obtained using M. tuberculosis strains lacking MtAhpC are not straightforward. AhpC expression in virulent strains of M. tuberculosis grown in vitro was repressed and increased under conditions of static growth, probably reflecting adaptation of the bacterium during its infection cycle (Springer et al, 2001). AhpC expression was also induced by hypoxia (Sherman et al, 2001).…”

“…The enzyme also protected human cells from toxicity caused by reactive nitrogen species (Chen et al, 1998). Whereas inactivation of MtAhpC caused no effect on bacterial growth during acute infection in mice and had no effect on in vitro sensitivity to H 2 O 2 , it caused an increase susceptibility to organic hydroperoxide and peroxynitrite-mediated toxicity (Springer et al, 2001;Master et al, 2002). Inactivation of MtAhpC caused a decrease in the survival of M. tuberculosis in non-stimulated macrophages but not in macrophages stimulated with interferon- (Master et al, 2002).…”