Metabolic Switching of Mycobacterium tuberculosis during Hypoxia Is Controlled by the Virulence Regulator PhoP

Abstract

Mycobacterium tuberculosis retains the ability to establish an asymptomatic latent infection. A fundamental question in mycobacterial physiology is to understand the mechanisms involved in hypoxic stress, a critical player in persistence. Here, we show that the virulence regulator PhoP responds to hypoxia, the dormancy signal, and effectively integrates hypoxia with nitrogen metabolism. We also provide evidence to demonstrate that both under nitrogen limiting conditions and during hypoxia, phoP locus controls key genes involved in nitrogen metabolism. Consistently, under hypoxia a ΔphoP strain shows growth attenuation even with surplus nitrogen, the alternate electron acceptor, and complementation of the mutant restores bacterial growth. Together, our observations provide new biological insights into the role of PhoP in integrating nitrogen metabolism with hypoxia by the assistance of the hypoxia regulator DosR. The results have significant implications on the mechanism of intracellular survival and growth of the tubercle bacilli under a hypoxic environment within the phagosome. IMPORTANCE M. tuberculosis retains the unique ability to establish an asymptomatic latent infection. To understand the mechanisms involved in hypoxic stress which play a critical role in persistence, we show that the virulence regulator PhoP is linked to hypoxia, the dormancy signal. In keeping with this, phoP was shown to play a major role in M. tuberculosis growth under hypoxia even in the presence of surplus nitrogen, the alternate electron acceptor. Our results showing regulation of hypoxia-responsive genes provide new biological insights into role of the virulence regulator in metabolic switching by sensing hypoxia and integrating nitrogen metabolism with hypoxia by the assistance of the hypoxia regulator DosR.

Convergence of two global regulators to coordinate expression of essential virulence determinants of Mycobacterium tuberculosis

“…Because PhoP was shown to interact with other regulators via its N-terminal domain ( Sevalkar et al, 2019 ; Singh et al, 2020 ), we next assessed the role of different stretches of N-terminal domain of PhoP (referred to as PhoP N ) in CRP-PhoP interactions. In vitro pull-down assays using mutant GST-PhoP proteins (each with three potential CRP- contacting residues of PhoP replaced with Ala) with His-tagged CRP displayed effective protein-protein interaction as that of the WT PhoP (compare lane 1 with lanes 2–6) ( Figure 5—figure supplement 1A ).…”

Exposure of Mycobacterium avium subsp. homonissuis to Metal Concentrations of the Phagosome Environment Enhances the Selection of Persistent Subpopulation to Antibiotic Treatment

“…For instance, iron is seen in greater concentration in the mycobacterial vacuoles of cultured as well as mice macrophages at 24 h of infection than 1 h post-infection [ 17 ]. For establishing the intracellular niche and survival within the host, mycobacteria require iron [ 11 , 13 , 14 ]. The iron acquisition molecules such as siderophore and mycobactins assist bacteria to recruit transferrin, a host iron-transport protein, to the phagosome and, thus, increase the vacuolar iron concentration [ 41 ].…”

“…M. tuberculosis has 88 toxin-antitoxin genes, many of them associated with persistence [ 12 ]. The hspR deletion, an important regulatory gene for heat-shock proteins, results in a persistence defect and the virulence regulator PhoP responds to hypoxic stresses by metabolic switch of nitrogen metabolism for long-term survival [ 13 ]. In addition, the oxidative stress and reactive oxygen species (ROS) can activate bacterial efflux system for recycling of damaged proteins and, thus, enhancing mycobacterial tolerance to antibiotics by increased drug efflux from bacterial cells [ 14 , 15 ].…”

Phenotypic adaptation of Mycobacterium tuberculosis to host-associated stressors that induce persister formation

“…Upon persister formation, an increase in nitrate reductase occurs, indicating that M. tuberculosis utilizes nitrate as an alternative electron acceptor during anaerobic respiration since reduction in electron flow associated with decreasing oxygen levels results in an increased concentration of reduced cofactors, such as NADH ( Sohaskey, 2008 ; Williams et al., 2015 ). Furthermore, PhoP and DosR regulates expression of nitrite and nitrate reductases for persistence during hypoxia ( Singh et al., 2020 ).…”