Redox homeostasis in mycobacteria: the key to tuberculosis control?

Abstract

Mycobacterium tuberculosis (Mtb) is a metabolically flexible pathogen that has the extraordinary ability to sense and adapt to the continuously changing host environment experienced during decades of persistent infection. Mtb is continually exposed to endogenous reactive oxygen species (ROS) as part of normal aerobic respiration, as well as exogenous ROS and reactive nitrogen species (RNS) generated by the host immune system in response to infection. The magnitude of tuberculosis (TB) disease is further amplified by exposure to xenobiotics from the environment such as cigarette smoke and air pollution, causing disruption of the intracellular prooxidant-antioxidant balance. Both oxidative and reductive stresses induce redox cascades that alter Mtb signal transduction, DNA and RNA synthesis, protein synthesis and antimycobacterial drug resistance. As reviewed in this article, Mtb has evolved specific mechanisms to protect itself against endogenously produced oxidants, as well as defend against host and environmental oxidants and reductants found specifically within the microenvironments of the lung. Maintaining an appropriate redox balance is critical to the clinical outcome because several antimycobacterial prodrugs are only effective upon bioreductive activation. Proper homeostasis of oxido-reductive systems is essential for Mtb survival, persistence and subsequent reactivation. The progress and remaining deficiencies in understanding Mtb redox homeostasis are also discussed.

Figure 1

Virulence life cycle of Mycobacterium tuberculosis and progression of TB. Mtb is transmitted by aerosol, and in 95% of cases, wherein the tubercle bacilli are inhaled, a primary infection is established. This is either cleared by the surge of the cell-mediated immunity or contained inside the granuloma in the form of latent TB, defined by no visible symptom of disease, but persistent, yet dormant, live bacilli within the host. The progress of TB can be stalled at this stage in some cases by isoniazid preventive therapy. This state might last for the lifespan of the infected individual, or progress to active TB by reactivation of the existing infection, with a lifetime risk of 5–10%. This risk of progression is exacerbated by immune-compromising factors such as HIV-AIDS, diabetes, indoor air pollution and tobacco smoke. Reactivation of TB is shown to occur at the upper and more oxygenated lobe of the lung, which can be cured by compliance with drug therapy. However, untreated or poorly treated TB might lead to the formation of tuberculous lesions in the lung. The development of cavities close to airway spaces allows shedding (e.g. coughing) of the bacilli through the airway, a stage of transmission. Subsequently, in a cyclic manner, the TB bacilli are transmitted to other individuals to establish primary infection.

Figure 2

Effect of exogenous environmental and endogenous host redox factors on the pathogenesis of TB. Infectious Mycobacterium tuberculosis (Mtb) bacilli are inhaled as aerosols from the atmosphere and phagocytosed by alveolar macrophages in the lung. A localised proinflammatory immune response causes the recruitment of mononuclear cells, leading to the establishment of a granuloma. However, Mtb cells are also present in lesion-free tissue. During the course of infection, caseous (typically hypoxic), fibrotic and non-necrotic granulomas can develop. The containment of Mtb by these granulomas never operates in isolation, and can fail as a consequence of malnutrition, diabetes, indoor air pollution, tobacco smoke and HIV infection, which are major risk factors for TB. Thus, any condition that weakens the immune status (in particular, a decrease in the function of CD4⁺ T cells) of the host can lead to TB. Exogenous environmental pollutants, which consist largely of redox-active molecules, not only affect the host immune response, but also target the infecting bacilli. Exposure to these environmental agents, production of host redox molecules such as O₂^•−, NO, ONOO⁻, etc. that are generated during the oxidative burst, and the pathological and physiological host responses induced on infection (e.g. hypoxic granuloma, dysregulated host lipid production) can collectively cause an imbalance in Mtb redox homeostasis, leading to oxidative stress or damage. Conversely, exogenous factors and the dysregulation of endogenous host redox factors might lead to the establishment of Mtb infection, maintaining a persistent state or allowing the bacillus to emerge from persistence. Dormant Mtb cells residing inside hypoxic granulomas are resistant to current antimycobacterial drugs and therefore have substantial implications on therapeutic intervention strategies. Moreover, the dynamic physiology and structure of the lung further complicate the situation because no two regions inside the lungs are similar in terms of their architecture and oxygen tension. This also makes it extremely difficult to study the progression of TB using animal models. Inside the lung, Mtb cells are exposed during transmission to a range of oxygen levels that varies from 150 to 180 mmHg in the upper respiratory tract to 1.9 mmHg within the granuloma, compared with pO₂ levels of healthy lungs (~59 mmHg). In addition, host pH and the type of in vivo carbon source, along with its concentration, will also have an impact on Mtb redox homeostasis. Nonetheless, it is still not clear how exposure of Mtb to these exogenous and endogenous redox molecules affects Mtb physiology and redox homeostasis in vivo to favour disease.

Figure 3

Mycobacterial mechanisms of sensing and countering endogenous or exogenous stress. The host generates free radicals, non-free-radical molecules and numerous gases as a mechanism to counter Mtb infection. These molecules target mycobacterial DNA, proteins and lipids, may alter Mtb gene expression, and change the overall metabolic profile or peptide pool. Free radicals can react with prosthetic groups such as Fe–S clusters or haem groups of the respiratory complexes. Mtb responds to these free radical stresses by adjusting its energy metabolism, physiological response and signal transduction cascade. Most of the radical-mediated damage is countered by detoxification processes comprising (a) enzymes such as catalase, superoxide dismutase and alkyl hydroperoxide, (b) redox buffering systems (thioredoxins, mycothiol, ergothioneine and protein thiols) and (c) truncated haemoglobins and cofactors (NAD⁺, FAD⁺ and coenzyme A). Mtb also possesses sensing mechanisms to detect environmental gases such as gradients of O₂, NO and alterations in its intracellular redox state to allow its survival. Well-studied examples are the Dos dormancy regulon and the WhiB3 redox sensor. The Dos regulon senses O₂, NO and CO through the DosS and DosT haem proteins. The signal is relayed to DosR, which leads to the induction of the 48-member Dos dormancy regulon that includes genes involved in energy production, dissipating reducing equivalents and assimilation of storage lipids, which is thought to facilitate mycobacterial persistence. WhiB3 functions as a regulator of cellular metabolism, which responds to O₂ and NO through its Fe–S cluster and integrates it with intermediary metabolic pathways. WhiB3 is an intracellular redox regulator that dissipates reductive stress generated by utilisation of host fatty acids through β-oxidation. Through the transcriptional activation of genes involved in lipid anabolism, WhiB3 is thought to direct reducing equivalents into the production of cell wall components and virulence lipids such as sulfolipids, phthiocerol dimycocerosates, polyacyltrehaloses and DAT. Under certain conditions, WhiB3 regulates the production and accumulation of triacylglycerol, indicating a link with the Dos dormancy signalling pathway.