scholarly journals Unique Roles of DosT and DosS in DosR Regulon Induction and Mycobacterium tuberculosis Dormancy

2009 ◽  
Vol 77 (8) ◽  
pp. 3258-3263 ◽  
Author(s):  
Ryan W. Honaker ◽  
Rachel L. Leistikow ◽  
Iona L. Bartek ◽  
Martin I. Voskuil
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Latent Infection ◽  
Transcriptional Regulator ◽  
Wild Type ◽  
Hypoxic Conditions ◽  
Sensor Kinases ◽  
The Individual ◽  
Anaerobic Dormancy ◽  
Dosr Regulon

ABSTRACT In Mycobacterium tuberculosis, the sensor kinases DosT and DosS activate the transcriptional regulator DosR, resulting in the induction of the DosR regulon, which is important for anaerobic survival and perhaps latent infection. The individual and collective roles of these sensors have been postulated biochemically, but their roles in vivo have remained unclear. This work demonstrates distinct and additive roles for each sensor during anaerobic dormancy. Both sensors are necessary for wild-type levels of DosR regulon induction, and concomitantly, full induction of the regulon is required for wild-type anaerobic survival. In the anaerobic model, DosT plays an early role, responding to hypoxia. DosT then induces the regulon and with it DosS, which sustains and further induces the regulon. DosT then loses its functionality as oxygen becomes limited, and DosS alone maintains induction of the genes from that point forward. Thus, M. tuberculosis has evolved a system whereby it responds to hypoxic conditions in a stepwise fashion as it enters an anaerobic state.

scholarly journals Domain II plays a crucial role in the function of ribosome recycling factor

2006 ◽  
Vol 393 (3) ◽  
pp. 767-777 ◽  
Author(s):  
Peng Guo ◽  
Liqiang Zhang ◽  
Hongjie Zhang ◽  
Yanming Feng ◽  
Guozhong Jing
Keyword(s):  
Crucial Role ◽  
Specific Interaction ◽  
Elongation Factor ◽  
Wild Type ◽  
Ribosome Recycling Factor ◽  
E Coli ◽  
Thermoanaerobacter Tengcongensis ◽  
The Individual

RRF (ribosome recycling factor) consists of two domains, and in concert with EF-G (elongation factor-G), triggers dissociation of the post-termination ribosomal complex. However, the function of the individual domains of RRF remains unclear. To clarify this, two RRF chimaeras, EcoDI/TteDII and TteDI/EcoDII, were created by domain swaps between the proteins from Escherichia coli and Thermoanaerobacter tengcongensis. The ribosome recycling activity of the RRF chimaeras was compared with their wild-type RRFs by using in vivo and in vitro activity assays. Like wild-type TteRRF (T. tengcongensis RRF), the EcoDI/TteDII chimaera is non-functional in E. coli, but both wild-type TteRRF, and EcoDI/TteDII can be activated by coexpression of T. tengcongensis EF-G in E. coli. By contrast, like wild-type E. coli RRF (EcoRRF), TteDI/EcoDII is fully functional in E. coli. These findings suggest that domain II of RRF plays a crucial role in the concerted action of RRF and EF-G for the post-termination complex disassembly, and the specific interaction between RRF and EF-G on ribosomes mainly depends on the interaction between domain II of RRF and EF-G. This study provides direct genetic and biochemical evidence for the function of the individual domains of RRF.

scholarly journals Deletion of the Mycobacterium tuberculosis α-Crystallin-Like hspX Gene Causes Increased Bacterial Growth In Vivo

2006 ◽  
Vol 74 (2) ◽  
pp. 861-868 ◽  
Author(s):  
Yanmin Hu ◽  
Farahnaz Movahedzadeh ◽  
Neil G. Stoker ◽  
Anthony R. M. Coates
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Constitutive Expression ◽  
Active Role ◽  
Structural Genes ◽  
Wild Type ◽  
Major Antigen ◽  
Oxide Stress ◽  
Unusual Phenotype

ABSTRACT Hypervirulent mutants of Mycobacterium tuberculosis, whose growth rates are higher in vivo, have now been reported to have mutations in both regulatory and structural genes, but the basis for this unusual phenotype is not understood. One hypervirulence gene, dosR (devR, Rv2031c), activates transcription of approximately 50 genes in this pathogen in response to hypoxia and nitric oxide stress. The most dramatic activation (∼80-fold) is activation of the hspX (acr, Rv2031c) gene, which encodes a 16-kDa α-crystallin-like protein that is a major antigen. In this study we found that a Δacr mutant exhibited increased growth following infection of BALB/c mice in vivo and in both resting and activated macrophages in vitro (as measured by the number of CFU). The increased growth in macrophages was equal to that of a ΔdosR mutant, while introduction of a constitutively expressed hspX gene reduced the ΔdosR virulence to wild-type levels. These results suggest that the increased number of CFU of the ΔdosR mutant was largely due to loss of hspX expression. We also confirmed that constitutive expression of hspX slows growth in vitro, and we propose that hspX plays an active role in slowing the growth of M. tuberculosis in vivo immediately following infection.

scholarly journals Construction and Phenotypic Characterization of an Auxotrophic Mutant of Mycobacterium tuberculosis Defective in l-Arginine Biosynthesis

2002 ◽  
Vol 70 (6) ◽  
pp. 3080-3084 ◽  
Author(s):  
Bhavna G. Gordhan ◽  
Debbie A. Smith ◽  
Heidi Alderton ◽  
Ruth A. McAdam ◽  
Gregory J. Bancroft ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Mutant Strain ◽  
Auxotrophic Mutant ◽  
Phenotypic Characterization ◽  
Wild Type ◽  
Arginine Biosynthesis ◽  
High Concentrations

ABSTRACT A mutant of Mycobacterium tuberculosis defective in the metabolism of l-arginine was constructed by allelic exchange mutagenesis. The argF mutant strain required exogenous l-arginine for growth in vitro, and in the presence of 0.96 mM l-arginine, it achieved a growth rate and cell density in stationary phase comparable to those of the wild type. The mutant strain was also able to grow in the presence of high concentrations of argininosuccinate, but its auxotrophic phenotype could not be rescued by l-citrulline, suggesting that the ΔargF::hyg mutation exerted a polar effect on the downstream argG gene but not on argH. The mutant strain displayed reduced virulence in immunodeficient SCID mice and was highly attenuated in immunocompetent DBA/2 mice, suggesting that l-arginine availability is restricted in vivo.

scholarly journals Exaggerated in vivo IL-17 responses discriminate recall responses in active TB

2019 ◽  
Author(s):  
Gabriele Pollara ◽  
Carolin T Turner ◽  
Gillian S Tomlinson ◽  
Lucy CK Bell ◽  
Ayesha Khan ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Immune Responses ◽  
Th17 Cells ◽  
Latent Infection ◽  
Inflammatory Diseases ◽  
Chronic Inflammatory Diseases ◽  
Host Immune Responses ◽  
Matrix Metalloproteinase 1 ◽  
First Time

AbstractHost immune responses at the site of Mycobacterium tuberculosis (Mtb) infection serve to contain the pathogen, but also mediate the pathogenesis of tuberculosis (TB) and onward transmission of infection. Interferon gamma (IFNγ) responses do not discriminate between protection and pathogenicity, but IL-17A/F responses, known to drive pathology in diverse chronic inflammatory diseases, have also been associated with TB pathogenesis in animal models. At the site of in vivo immune recall responses to Mtb modelled by the tuberculin skin test, we show for the first time that active TB in humans is also associated with exaggerated IL-17A/F expression, accumulation of Th17 cells and IL-17A/F bioactivity, including increased neutrophil recruitment and matrix metalloproteinase-1 expression directly implicated in TB pathogenesis. These features discriminate recall responses in patients with active TB from those with cured or latent infection and are also evident at the site of TB disease. Our data support targeting of this pathway in host-directed therapy for TB.

scholarly journals Role of the dosR-dosS Two-Component Regulatory System in Mycobacterium tuberculosis Virulence in Three Animal Models

2008 ◽  
Vol 77 (3) ◽  
pp. 1230-1237 ◽  
Author(s):  
Paul J. Converse ◽  
Petros C. Karakousis ◽  
Lee G. Klinkenberg ◽  
Anup K. Kesavan ◽  
Lan H. Ly ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Animal Models ◽  
Hollow Fiber ◽  
Regulatory System ◽  
Growth Defect ◽  
Bacterial Survival ◽  
Wild Type ◽  
Fiber Model

ABSTRACT The Mycobacterium tuberculosis dosR gene (Rv3133c) is part of an operon, Rv3134c-Rv3132c, and encodes a response regulator that has been shown to be upregulated by hypoxia and other in vitro stress conditions and may be important for bacterial survival within granulomatous lesions found in tuberculosis. DosR is activated in response to hypoxia and nitric oxide by DosS (Rv3132c) or DosT (Rv2027c). We compared the virulence levels of an M. tuberculosis dosR-dosS deletion mutant (ΔdosR-dosS [ΔdosR-S]), a dosR-complemented strain, and wild-type H37Rv in rabbits, guinea pigs, and mice infected by the aerosol route and in a mouse hollow-fiber model that may mimic in vivo granulomatous conditions. In the mouse and the guinea pig models, the ΔdosR-S mutant exhibited a growth defect. In the rabbit, the ΔdosR-S mutant did not replicate more than the wild type. In the hollow-fiber model, the mutant phenotype was not different from that of the wild-type strain. Our analyses reveal that the dosR and dosS genes are required for full virulence and that there may be differences in the patterns of attenuation of this mutant between the animal models studied.

scholarly journals Regulation of Mycobacterium tuberculosis whiB3 in the Mouse Lung and Macrophages

2006 ◽  
Vol 74 (11) ◽  
pp. 6449-6457 ◽  
Author(s):  
N. Banaiee ◽  
W. R. Jacobs ◽  
J. D. Ernst
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Mouse Lung ◽  
Wild Type ◽  
Necrosis Factor Alpha ◽  
Environmental Signals ◽  
Immunodeficient Mice ◽  
Reverse Transcription Pcr ◽  
Factor Alpha

ABSTRACT Mycobacterium tuberculosis is a highly successful human pathogen, with ∼2 × 109 individuals infected globally. To understand the responses of M. tuberculosis to the in vivo environment, we studied the in vivo regulation of M. tuberculosis genes whose M. marinum homologs are induced in chronically infected frog tissues. The expression of 16S rRNA was shown to remain constant in M. tuberculosis under in vivo and in vitro conditions and therefore could be used for internal normalization in quantitative reverse transcription-PCR assays. We found whiB3, a putative transcriptional regulator implicated in mediating tissue damage, to be maximally induced at 2 weeks postinfection in the lungs of wild-type and immunodeficient (gamma interferon receptor−/−, Rag1−/−, and tumor necrosis factor alpha−/−) mice. At later time points in wild-type mice, whiB3 induction was decreased and gradually declined over the course of infection. In immunodeficient mice, whiB3 induction declined rapidly and was completely abolished in moribund animals. whiB3 was also found to be induced in naïve bone marrow-derived macrophages after 6 h of infection. whiB3 expression in vivo and in vitro was found to be inversely correlated with bacterial density. These results indicate that M. tuberculosis regulates the expression of whiB3 in response to environmental signals present in vivo and are consistent with a model of regulation by quorum sensing.

scholarly journals Fibrinogen Regulates the Cytotoxicity of Mycobacterial Trehalose Dimycolate but Is Not Required for Cell Recruitment, Cytokine Response, or Control of Mycobacterial Infection

2009 ◽  
Vol 78 (3) ◽  
pp. 1004-1011 ◽  
Author(s):  
Kaori Sakamoto ◽  
Rachel E. Geisel ◽  
Mi-Jeong Kim ◽  
Bryce T. Wyatt ◽  
Llewelyn B. Sellers ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Granulation Tissue ◽  
Inflammatory Responses ◽  
Mycobacterial Infection ◽  
Cytokine Response ◽  
Wild Type ◽  
Cell Recruitment ◽  
Trehalose Dimycolate ◽  
Deficient Mice

ABSTRACT During inflammatory responses and wound healing, the conversion of soluble fibrinogen to fibrin, an insoluble extracellular matrix, long has been assumed to create a scaffold for the migration of leukocytes and fibroblasts. Previous studies concluded that fibrinogen is a necessary cofactor for mycobacterial trehalose 6,6′-dimycolate-induced responses, because trehalose dimycolate-coated beads, to which fibrinogen was adsorbed, were more inflammatory than those to which other plasma proteins were adsorbed. Herein, we investigate roles for fibrin(ogen) in an in vivo model of mycobacterial granuloma formation and in infection with Mycobacterium tuberculosis, the causative agent of tuberculosis. In wild-type mice, the subcutaneous injection of trehalose dimycolate-coated polystyrene microspheres, suspended within Matrigel, elicited a pyogranulomatous response during the course of 12 days. In fibrinogen-deficient mice, neutrophils were recruited but a more suppurative lesion developed, with the marked degradation and disintegration of the matrix. Compared to that in wild-type mice, the early formation of granulation tissue in fibrinogen-deficient mice was edematous, hypocellular, and disorganized. These deficiencies were complemented by the addition of exogenous fibrinogen. The absence of fibrinogen had no effect on cell recruitment or cytokine production in response to trehalose dimycolate, nor was there a difference in lung histopathology or overall bacterial burden in mice infected with Mycobacterium tuberculosis. In this model, fibrin(ogen) was not required for cell recruitment, cytokine response, or response to infection, but it promoted granulation tissue formation and suppressed leukocyte necrosis.

scholarly journals The cGAS/STING Pathway Is Important for Dendritic Cell Activation but Is Not Essential to Induce Protective Immunity against Mycobacterium tuberculosis Infection

2018 ◽  
Vol 10 (3) ◽  
pp. 239-252 ◽  
Author(s):  
Fabio V. Marinho ◽  
Sulayman Benmerzoug ◽  
Stephanie  Rose ◽  
Priscila C. Campos ◽  
João T. Marques ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Cell Activation ◽  
Body Weight Gain ◽  
Host Cells ◽  
Health Concern ◽  
Wild Type ◽  
Activation Markers ◽  
Sting Pathway

Mycobacterium tuberculosis (Mtb) infection remains a major public health concern. The STING (stimulator of interferon genes) pathway contributes to the cytosolic surveillance of host cells. Most studies on the role of STING activation in Mtb infection have focused on macrophages. Moreover, a detailed investigation of the role of STING during Mtb infection in vivo is required. Here, we deciphered the involvement of STING in the activation of dendritic cells (DCs) and the host response to Mtb infection in vivo. In DCs, this adaptor molecule was important for Ifn-β expression and IL-12 production as well as for the surface expression of the activation markers CD40 and CD86. We also documented that Mtb DNA induces STING activation in murine fibroblasts. In vivo Mtb aerogenic infection induced the upregulation of the STING and cGAS (cyclic GMP-AMP synthase) genes, and Ifn-β pulmonary expression was dependent on both sensors. However, mice deficient for STING or cGAS presented a similar outcome to wild-type controls, with no major alterations in body weight gain, bacterial burden, or survival. Lung inflammation, proinflammatory cytokine production, and inflammatory cell recruitment were similar in STING- and cGAS-deficient mice compared to wild-type controls. In summary, although the STING pathway seems to be crucial for DC activation during Mtb infection, it is dispensable for host protection in vivo.

scholarly journals Altered Oligomerization Properties of N316 Mutants of Escherichia coli TyrR

2008 ◽  
Vol 190 (24) ◽  
pp. 8238-8243 ◽  
Author(s):  
Takashi Koyanagi ◽  
Takane Katayama ◽  
Hideyuki Suzuki ◽  
Hidehiko Kumagai
Keyword(s):  
Gene Expression ◽  
Escherichia Coli ◽  
Amino Acids ◽  
Gel Filtration ◽  
Aromatic Amino Acids ◽  
Transcriptional Regulator ◽  
Wild Type ◽  
Oligomer Formation ◽  
Regulatory Properties

ABSTRACT The transcriptional regulator TyrR is known to undergo a dimer-to-hexamer conformational change in response to aromatic amino acids, through which it controls gene expression. In this study, we identified N316D as the second-site suppressor of Escherichia coli TyrRE274Q, a mutant protein deficient in hexamer formation. N316 variants exhibited altered in vivo regulatory properties, and the most drastic changes were observed for TyrRN316D and TyrRN316R mutants. Gel filtration analyses revealed that the ligand-mediated oligomer formation was enhanced and diminished for TyrRN316D and TyrRN316R, respectively, compared with the wild-type TyrR. ADP was substituted for ATP in the oligomer formation of TyrRN316D.

scholarly journals The Mycobacterium tuberculosis DosR Regulon Assists in Metabolic Homeostasis and Enables Rapid Recovery from Nonrespiring Dormancy

2009 ◽  
Vol 192 (6) ◽  
pp. 1662-1670 ◽  
Author(s):  
Rachel L. Leistikow ◽  
Russell A. Morton ◽  
Iona L. Bartek ◽  
Isaac Frimpong ◽  
Karleen Wagner ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Mycobacterium Tuberculosis ◽  
Energy Levels ◽  
Redox Balance ◽  
Bacterial Metabolism ◽  
Aerobic Respiration ◽  
Latently Infected ◽  
The Face ◽  
Anaerobic Dormancy ◽  
Dosr Regulon

ABSTRACT Mycobacterium tuberculosis survives in latently infected individuals, likely in a nonreplicating or dormancy-like state. The M. tuberculosis DosR regulon is a genetic program induced by conditions that inhibit aerobic respiration and prevent bacillus replication. In this study, we used a mutant incapable of DosR regulon induction to investigate the contribution of this regulon to bacterial metabolism during anaerobic dormancy. Our results confirm that the DosR regulon is essential for M. tuberculosis survival during anaerobic dormancy and demonstrate that it is required for metabolic processes that occur upon entry into and throughout the dormant state. Specifically, we showed that regulon mechanisms shift metabolism away from aerobic respiration in the face of dwindling oxygen availability and are required for maintaining energy levels and redox balance as the culture becomes anaerobic. We also demonstrated that the DosR regulon is crucial for rapid resumption of growth once M. tuberculosis exits an anaerobic or nitric oxide-induced nonrespiring state. In summary, the DosR regulon encodes novel metabolic mechanisms essential for M. tuberculosis to survive in the absence of respiration and to successfully transition rapidly between respiring and nonrespiring conditions without loss of viability.

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