Mycobacterial responses to in vivo stressors and conditions.
2. Adaptation to growth in the phagosomal compartment of macrophages
Macrophages are the preferred intracellular location for
3. Adaptation to granulomas and caseation
Once infection has progressed, tubercle bacilli replicate within incompletely activated macrophages. Additional macrophages arrive to the site of infection, and engulf newly liberated mycobacteria. The immune cells, T-cells, arrive to this location and an immune structure, the granuloma, composed of macrophages and a mantel of T-cells develops. If the host is resistant, and can robustly activate the body’s macrophages, then
4. Liquefied lesions and sputum
Later in infection caseating granulomas continue to breakdown. At a certain point these granulomas begin to liquefy, and host lipases and proteases are present which damage host tissues. Dead macrophages release lytic enzymes, and bacterial products may also result in host tissue damage and liquefaction ensues. As tissue is damaged, a cavity erodes into the lung airspace. In rabbit studies,
Mycobacterium tuberculosis and dormancy
One third of the world’s population is infected with
The gene encoding a transcriptional regulator,
The ability of
Mycobacterium tuberculosis responses to acidic stress
Mycobacteria seem to bear an intrinsic ability to resist acidic stress. They have a thick waxy cell wall as well as an outer membrane that can resist acidic stress. This physical barrier may serve to inhibit entry of toxic protons, and anything that interferes with this barrier could increase acid susceptibility. Many mutants that are acid susceptible lie in genes that affect cell wall and lipid metabolism (Table 1.). Environmental mycobacteria are found in conditions that may be acidic and can grow at pHs as low as 4.0 (Santos et al, 2007). Pathogenic mycobacteria have evolved to resist acidic stress, and potentially share similar mechanisms with their environmental cousins (Kirschner et al, 1992; Kirschner et al, 1999).
A number of genes that are upregulated by acidic stress have been identified in previous studies. Looking at rapid response to acidity at 15 or 30 minutes it was found that genes involved in cell wall ultrastructure were induced (Fisher et al, 2002). The
The type VII secretion system, Esx-1, may also may be involved in response to acid stress (Abdallah et al, 2007). The 6 kDa early secreted antigenic target (Esat-6) and the 10kDa culture filtrate protein (CFP-10) are secreted by Esx-1. These two proteins form a heterodimer that can dissociate at acidic pH. Esat-6 is capable of lysing membranes, and
7. Response to oxidative damage
Inside phagosomes of activated macrophages tubercle bacilli are exposed to reactive oxygen intermediates.
Mycobacteria contain a unique substance, mycothiol, which combats oxidative stress. Other bacterial species utilize glutathione which can also neutralize oxidative stress. Mycothiol contains cysteine residues which are oxidized when that condition predominates thus forming disulfide bonds, creating mycothione, and preventing other molecules in the mycobacterial cell from becoming oxidized (Table 1.). Human cells produce glutathione to combat oxidative damage, and glutathione is toxic to mycobacterial cells perhaps due to a redox imbalance generated by this substance in the mycobacteria (Venketaraman et al, 2008; Connell et al, 2008)). Mycobacteria also contain other molecules to detoxify oxidative damage including superoxide dismutase (SOD) and catalase (KatG) which can inactivate superoxide (Table 1.) (Shi et al, 2008). SOD and KatG are upregulated early in infection indicating an increase in oxidative damage due to superoxide. Oxidative damage is capable of harming DNA, and histone like proteins (LSR2) can protect against damage by compacting DNA and acting as a physical barrier. UvrB which repairs mycobacterial DNA damage also protects against oxidative damage (Darwin and Nathan, 2005; Colangeli et al, 2009).
8. Heat shock
One of the hallmarks of tuberculosis is fever and night sweats in which body temperature increases and is suboptimal for
Many proteins that are upregulated in
9. Low iron
Normally iron taken up by intestinal epithelial cells and bound to transferrin circulates within the body. This complex binds to cell surface receptors, and is internalized where it releases its iron to be bound by the host cellular factor ferritin. Infection and inflammation are natural signals to the host to limit availability of iron. Proinflammatory cytokines stimulate hepcidin production, decrease iron uptake from the gut, and inhibits the iron efflux protein ferroportin (Johnson and Wessingling-Resnick, 2012). Inflammation thus inhibits iron uptake by the intestinal epithelium thus preventing iron from being loaded onto transferrin. Interfering with uptake limits iron availability in the host, and
Mycobacteria have a variety of systems which aid in the uptake of iron and the regulation of iron responsive genes. As mycobacteria have been shown to be somewhat novel among gram positive bacteria, they possess an outer mycolic acid based membrane, as well as an inner membrane and periplasmic space. Porins in the outer membrane appear to transport iron in the presence of high iron conditions (Jones and Niederweis, 2010).
Iron responsive genes in
10. Hypoxic growth
11. Toxin-antitoxin systems
Interestingly there are many toxin-antitoxin systems within the
12. Two component systems
Two components systems are common in many bacteria. These systems are comprised of a sensor kinase which phosphorylates the response regulator as a result of an environmental signal, which is often a stress. The sensor kinases are trans membrane proteins which are embedded into membranes. They sense external stresses and transmit these signals internally into the bacterial cell by phosphorylating a response regulator that binds to its cognate promoter DNA, and regulates transcription. The mycobacterial genome contains 11 two component systems (Hett and Rubin, 2008). The large number of these systems in the mycobacterial coding regions is likely the result of evolution to accommodate bacterial responses to diverse stresses.
DosS/DosT-DosR was previously described, and responds to initial hypoxic stress (Table 1.) (Park et al, 2003). Some of the genes controlled by the transcriptional regulator DosR are upregulated by hypoxic stress, and are also part of the transcriptional regulator PhoP regulon, a member of the PhoP/R two component system. While it is unknown what environmental signal PhoP or the sensor kinase PhoR are responding to, genes controlled by PhoP either directly or indirectly are upregulated by such stresses as acidity and low oxygen (Table 1.) (Gonzalo-Asensio et al, 2008).
13. Sigma factors
Mycobacterial RNA polymerase catalyzes RNA synthesis from specific promoter sequences. This RNA polymerase is composed of subunits that comprise the core holoenzyme, and include two α subunits, a β, a β' and a ω subunit. The core enzyme, however, cannot target specific promoter sequences. A sigma factor is required for this function, and can bind and recognize specific -10 and -35 promoter sequences. As the mycobacterial genome possesses many different sigma factors, these RNA polymerase components can recognize diverse mycobacterial promoter sequences to activate a whole class of genes. This activity is in addition to specific transcription factors which bind to promoters, regulate transcription, and are not part of the RNA polymerase enzyme.
The mycobacterial genome possesses many different sigma factors that belong to different categories. The
As mycobacteria invade their human hosts they must respond to a plethora of stresses many of which are generated by the host's immune system. Under this selective pressure,
Understanding the specific steps in infection, the stresses associated with each step, and the mycobacterial response may be of clinical relevance. The knowledge that oxidative stress and acidic stress may predominate as adaptive immunity makes the host’s macrophages more activated, may lead to the development of chemotherapeutic agents that target mycobacterial components produced by these stressors during this infective stage. In addition, the knowledge that mycobacteria may utilize toxin-antitoxin systems to slow their growth and to enhance their innate antibiotic resistance may spur the development of therapies that target these systems which could be used in conjunction with traditional antibiotic treatments. Chemotherapeutic agents given to decrease activity of triacylglycerol synthase may decrease infectivity of sputum positive individuals by inhibiting lipid body production in the bacilli while antibiotic treatment lags in its sterilizing activity. Ultimately treatments may be developed which target inducible systems upregulated by stresses, and may interfere with mycobacterial responses to these stressors. By thwarting these adaptive responses potentially with chemotherapeutic agents, mycobacteria may be rendered more fragile and susceptible to the host's immune system. In addition a greater understanding of how
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