scholarly journals The Alternative Sigma Factor RpoE2 Is Involved in the Stress Response to Hypochlorite and in vivo Survival of Haemophilus influenzae

2021 ◽  
Vol 12 ◽  
Author(s):  
Marufa Nasreen ◽  
Aidan Fletcher ◽  
Jennifer Hosmer ◽  
Qifeng Zhong ◽  
Ama-Tawiah Essilfie ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Sigma Factor ◽  
Target Genes ◽  
Cell Envelope ◽  
Sigma Factors ◽  
Methionine Sulfoxide Reductase ◽  
Respiratory Pathogen ◽  
Human Pathogens

Extracytoplasmic function (ECF) sigma factors underpin the ability of bacteria to adapt to changing environmental conditions, a process that is particularly relevant in human pathogens that inhabit niches where human immune cells contribute to high levels of extracellular stress. Here, we have characterized the previously unstudied RpoE2 ECF sigma factor from the human respiratory pathogen H. influenzae (Hi) and its role in hypochlorite-induced stress. Exposure of H. influenzae to oxidative stress (HOCl, H2O2) increased rpoE2 gene expression, and the activity of RpoE2 was controlled by a cytoplasmic 67-aa anti-sigma factor, HrsE. RpoE2 regulated the expression of the periplasmic MsrAB peptide methionine sulfoxide reductase that, in H. influenzae, is required for HOCl resistance, thus linking RpoE2 to HOCl stress. Interestingly, a HiΔrpoE2 strain had wild-type levels of resistance to oxidative stress in vitro, but HiΔrpoE2 survival was reduced 26-fold in a mouse model of lung infection, demonstrating the relevance of this sigma factor for H. influenzae pathogenesis. The HiRpoE2 system has some similarity to the ECF sigma factors described in Streptomyces and Neisseria sp. that also control the expression of msr genes. However, HiRpoE2 regulation extended to genes encoding other periplasmic damage repair proteins, an operon containing a DoxX-like protein, and also included selected OxyR-controlled genes. Based on our results, we propose that the highly conserved HiRpoE2 sigma factor is a key regulator of H. influenzae responses to oxidative damage in the cell envelope region that controls a variety of target genes required for survival in the host.

scholarly journals Examination of Mycobacterium tuberculosis sigma factor mutants using low-dose aerosol infection of guinea pigs suggests a role for SigC in pathogenesis

Microbiology ◽  
2006 ◽  
Vol 152 (6) ◽  
pp. 1591-1600 ◽  
Author(s):  
Russell K. Karls ◽  
Jeannette Guarner ◽  
David N. McMurray ◽  
Kristin A. Birkness ◽  
Frederick D. Quinn
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Sigma Factor ◽  
Guinea Pigs ◽  
Low Dose ◽  
Sigma Factors ◽  
Respiratory Pathogen ◽  
Growth Deficiency ◽  
Aerosol Infection

Secondary sigma factors in bacteria direct transcription of defence regulons in response to specific stresses. To identify which sigma factors in the human respiratory pathogen Mycobacterium tuberculosis are important for adaptive survival in vivo, defined null mutations were created in individual sigma factor genes. In this study, in vitro growth virulence and guinea pig pathology of M. tuberculosis mutants lacking functional sigma factors (SigC, SigF, or SigM) were compared to the parent strain, H37Rv. None of the mutant strains exhibited a growth deficiency in Middlebrook 7H9 broth, nor were any impaired for intracellular replication in the human monocytic macrophage cell-line THP-1. Following low-dose aerosol infection of guinea pigs, however, differences could be detected. While a SigM mutant resulted in lung and spleen granulomas of comparable composition to those found in H37Rv-infected animals, a SigF mutant was partially attenuated, exhibiting necrotic spleen granulomas and ill-defined lung granulomas. SigC mutants exhibited attenuation in the lung and spleen; notably, necrotic granulomas were absent. These data suggest that while SigF may be important for survival in the lung, SigC is likely a key regulator of pathogenesis and adaptive survival in the lung and spleen. Understanding how SigC mediates survival in the host should prove useful in the development of anti-tuberculosis therapies.

scholarly journals The Mycobacterium tuberculosis Sigma Factor σB Is Required for Full Response to Cell Envelope Stress and Hypoxia In Vitro, but It Is Dispensable for In Vivo Growth

2009 ◽  
Vol 191 (18) ◽  
pp. 5628-5633 ◽  
Author(s):  
P. A. Fontán ◽  
M. I. Voskuil ◽  
M. Gomez ◽  
D. Tan ◽  
M. Pardini ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Mycobacterium Tuberculosis ◽  
Sigma Factor ◽  
Cell Envelope ◽  
Hypoxic Conditions ◽  
Envelope Stress ◽  
Stress Oxidative ◽  
In Vivo Growth

ABSTRACT The numerous sigma (σ) factors present in Mycobacterium tuberculosis are indicative of the adaptability of this pathogen to different environmental conditions. In this report, we describe the M. tuberculosis σB regulon and the phenotypes of an M. tuberculosis sigB mutant strain exposed to cell envelope stress, oxidative stress, and hypoxia. The sigB mutant was especially defective in survival under hypoxic conditions in vitro, but it was not attenuated for growth in THP-1 cells or during mouse and guinea pig infection.

scholarly journals The Mycobacterium tuberculosis Extracytoplasmic-Function Sigma Factor SigL Regulates Polyketide Synthases and Secreted or Membrane Proteins and Is Required for Virulence

2005 ◽  
Vol 187 (20) ◽  
pp. 7062-7071 ◽  
Author(s):  
Mi-Young Hahn ◽  
Sahadevan Raman ◽  
Mauricio Anaya ◽  
Robert N. Husson
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Stress Responses ◽  
Sigma Factor ◽  
Sigma Factors ◽  
Infection Model ◽  
Transcription Start Sites ◽  
Promoter Sequences ◽  
Extracytoplasmic Function Sigma Factor

ABSTRACT Mycobacterium tuberculosis sigL encodes an extracytoplasmic function (ECF) sigma factor and is adjacent to a gene for a membrane protein (Rv0736) that contains a conserved HXXXCXXC sequence. This motif is found in anti-sigma factors that regulate several ECF sigma factors, including those that control oxidative stress responses. In this work, SigL and Rv0736 were found to be cotranscribed, and the intracellular domain of Rv0736 was shown to interact specifically with SigL, suggesting that Rv0736 may encode an anti-sigma factor of SigL. An M. tuberculosis sigL mutant was not more susceptible than the parental strain to several oxidative and nitrosative stresses, and sigL expression was not increased in response to these stresses. In vivo, sigL is expressed from a weak SigL-independent promoter and also from a second SigL-dependent promoter. To identify SigL-regulated genes, sigL was overexpressed and microarray analysis of global transcription was performed. Four small operons, sigL (Rv0735)-Rv0736, mpt53 (Rv2878c)-Rv2877c, pks10 (Rv1660)-pks7 (Rv1661), and Rv1139c-Rv1138c, were among the most highly upregulated genes in the sigL-overexpressing strain. SigL-dependent transcription start sites of these operons were mapped, and the consensus promoter sequences TGAACC in the −35 region and CGTgtc in the −10 region were identified. In vitro, purified SigL specifically initiated transcription from the promoters of sigL, mpt53, and pks10. Additional genes, including four PE_PGRS genes, appear to be regulated indirectly by SigL. In an in vivo murine infection model, the sigL mutant strain showed marked attenuation, indicating that the sigL regulon is important in M. tuberculosis pathogenesis.

scholarly journals Control of Thioredoxin Reductase Gene (trxB) Transcription by SarA in Staphylococcus aureus

2009 ◽  
Vol 192 (1) ◽  
pp. 336-345 ◽  
Author(s):  
Anand Ballal ◽  
Adhar C. Manna
Keyword(s):  
Oxidative Stress ◽  
Staphylococcus Aureus ◽  
Dna Binding ◽  
Target Genes ◽  
Thioredoxin Reductase ◽  
Negative Regulator ◽  
Binding Studies ◽  
Oxidizing Agents

ABSTRACT Thioredoxin reductase (encoded by trxB) protects Staphylococcus aureus against oxygen or disulfide stress and is indispensable for growth. Among the different sarA family mutants analyzed, transcription of trxB was markedly elevated in the sarA mutant under conditions of aerobic as well as microaerophilic growth, indicating that SarA acts as a negative regulator of trxB expression. Gel shift analysis showed that purified SarA protein binds directly to the trxB promoter region DNA in vitro. DNA binding of SarA was essential for repression of trxB transcription in vivo in S. aureus. Northern blot analysis and DNA binding studies of the purified wild-type SarA and the mutant SarAC9G with oxidizing agents indicated that oxidation of Cys-9 reduced the binding of SarA to the trxB promoter DNA. Oxidizing agents, in particular diamide, could further enhance transcription of the trxB gene in the sarA mutant, suggesting the presence of a SarA-independent mode of trxB induction. Analysis of two oxidative stress-responsive sarA regulatory target genes, trxB and sodM, with various mutant sarA constructs showed a differential ability of the SarA to regulate expression of the two above-mentioned genes in vivo. The overall data demonstrate the important role played by SarA in modulating expression of genes involved in oxidative stress resistance in S. aureus.

scholarly journals Sigma Factor F Does Not Prevent Rifampin Inhibition of RNA Polymerase or Cause Rifampin Tolerance in Mycobacterium tuberculosis

2010 ◽  
Vol 192 (20) ◽  
pp. 5472-5479 ◽  
Author(s):  
Ruben C. Hartkoorn ◽  
Claudia Sala ◽  
Sophie J. Magnet ◽  
Jeffrey M. Chen ◽  
Florence Pojer ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Rna Polymerase ◽  
Stationary Phase ◽  
Sigma Factor ◽  
Potent Inhibitor ◽  
Sigma Factors ◽  
Antituberculosis Drugs ◽  
Axenic Cultures

ABSTRACT The tolerance of Mycobacterium tuberculosis to antituberculosis drugs is a major reason for the lengthy therapy needed to treat a tuberculosis infection. Rifampin is a potent inhibitor of RNA polymerase (RNAP) in vivo but has been shown to be less effective against stationary-phase bacteria. Sigma factor F is associated with bacteria entering stationary phase and has been proposed to impact rifampin activity. Here we investigate whether RNAP containing SigF is more resistant to rifampin inhibition in vitro and whether overexpression of sigF renders M. tuberculosis more tolerant to rifampin. Real-time and radiometric in vitro transcription assays revealed that rifampin equally inhibits transcription by RNAP containing sigma factors SigA and SigF, therefore ruling out the hypothesis that SigF may be responsible for increased resistance of the enzyme to rifampin in vitro. In addition, overexpression or deletion of sigF did not alter rifampin susceptibility in axenic cultures of M. tuberculosis, indicating that SigF does not affect rifampin tolerance in vivo.

scholarly journals ςBldN, an Extracytoplasmic Function RNA Polymerase Sigma Factor Required for Aerial Mycelium Formation in Streptomyces coelicolor A3(2)

2000 ◽  
Vol 182 (16) ◽  
pp. 4606-4616 ◽  
Author(s):  
Maureen J. Bibb ◽  
Virginie Molle ◽  
Mark J. Buttner
Keyword(s):  
Rna Polymerase ◽  
Sigma Factor ◽  
Streptomyces Coelicolor ◽  
Aerial Mycelium ◽  
Sigma Factors ◽  
Colony Development ◽  
Null Mutant ◽  
Wild Type

ABSTRACT Sporulation mutants of Streptomyces coelicolor appear white because they are defective in the synthesis of the gray polyketide spore pigment, and such white (whi) mutants have been used to define 13 sporulation loci. whiN, one of five new whi loci identified in a recent screen of NTG (N-methyl-N′-nitro-N-nitrosoguanidine)-inducedwhi strains (N. J. Ryding et al., J. Bacteriol. 181:5419–5425, 1999), was defined by two mutants, R112 and R650. R650 produced frequent spores that were longer than those of the wild type. In contrast, R112 produced long, straight, undifferentiated hyphae, although rare spore chains were observed, sometimes showing highly irregular septum placement. Subcloning and sequencing showed thatwhiN encodes a member of the extracytoplasmic function subfamily of RNA polymerase sigma factors and that the sigma factor has an unusual N-terminal extension of approximately 86 residues that is not present in other sigma factors. A constructed whiN null mutant failed to form aerial mycelium (the “bald” phenotype) and, as a consequence, whiN was renamed bldN. This observation was not totally unexpected because, on some media, the R112 point mutant produced substantially less aerial mycelium than its parent, M145. The bldN null mutant did not fit simply into the extracellular signaling cascade proposed for S. coelicolor bld mutants. Expression of bldN was analyzed during colony development in wild-type and aerial mycelium-deficientbld strains. bldN was transcribed from a single promoter, bldNp. bldN transcription was developmentally regulated, commencing approximately at the time of aerial mycelium formation, and depended on bldG and bldH, but not on bldA, bldB, bldC,bldF, bldK, or bldJ or onbldN itself. Transcription from the p1 promoter of the response-regulator gene bldM depended onbldN in vivo, and the bldMp1 promoter was shown to be a direct biochemical target for ςBldN holoenzyme in vitro.

scholarly journals The Involvement of Mycobacterium Type III-A CRISPR-Cas System in Oxidative Stress

2021 ◽  
Vol 12 ◽  
Author(s):  
Fan Yang ◽  
Lingqing Xu ◽  
Lujie Liang ◽  
Wanfei Liang ◽  
Jiachen Li ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Cell Envelope ◽  
Reduction Process ◽  
Intracellular Survival ◽  
Host Immunity ◽  
Type I ◽  
Type Iii

Type I and type II CRISPR-Cas systems are employed to evade host immunity by targeting interference of bacteria’s own genes. Although Mycobacterium tuberculosis (M. tuberculosis), the causative agent of tuberculosis, possesses integrated type III-A CRISPR-Cas system, its role in mycobacteria remains obscure. Here, we observed that seven cas genes (csm2∼5, cas10, cas6) were upregulated in Mycobacterium bovis BCG under oxidative stress treatment, indicating the role of type III-A CRISPR-Cas system in oxidative stress. To explore the functional role of type III-A CRISPR-Cas system, TCC (Type III-A CRISPR-Cas system, including cas6, cas10, and csm2-6) mutant was generated. Deletion of TCC results in increased sensitivity in response to hydrogen peroxide and reduced cell envelope integrity. Analysis of RNA-seq dataset revealed that TCC impacted on the oxidation-reduction process and the composition of cell wall which is essential for mycobacterial envelop integrity. Moreover, disrupting TCC led to poor intracellular survival in vivo and in vitro. Finally, we showed for the first time that TCC contributed to the regulation of regulatory T cell population, supporting a role of TCC in modulating host immunity. Our finding reveals the important role of TCC in cell envelop homeostasis. Our work also highlights type III-A CRISPR-Cas system as an important factor for intracellular survival and host immunoregulation in mycobacteria, thus may be a potential target for therapy.

scholarly journals Oncogenic hijacking of a developmental transcription factor evokes vulnerability toward oxidative stress in Ewing sarcoma

2020 ◽  
Vol 11 (1) ◽  
Author(s):  
Aruna Marchetto ◽  
Shunya Ohmura ◽  
Martin F. Orth ◽  
Maximilian M. L. Knott ◽  
Maria V. Colombo ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Transcription Factor ◽  
Ewing Sarcoma ◽  
Target Genes ◽  
Transcriptome Profiling ◽  
In Vivo Experiments ◽  
Ets Family ◽  
Pdx Models

AbstractEwing sarcoma (EwS) is an aggressive childhood cancer likely originating from mesenchymal stem cells or osteo-chondrogenic progenitors. It is characterized by fusion oncoproteins involving EWSR1 and variable members of the ETS-family of transcription factors (in 85% FLI1). EWSR1-FLI1 can induce target genes by using GGAA-microsatellites as enhancers.Here, we show that EWSR1-FLI1 hijacks the developmental transcription factor SOX6 – a physiological driver of proliferation of osteo-chondrogenic progenitors – by binding to an intronic GGAA-microsatellite, which promotes EwS growth in vitro and in vivo. Through integration of transcriptome-profiling, published drug-screening data, and functional in vitro and in vivo experiments including 3D and PDX models, we discover that constitutively high SOX6 expression promotes elevated levels of oxidative stress that create a therapeutic vulnerability toward the oxidative stress-inducing drug Elesclomol.Collectively, our results exemplify how aberrant activation of a developmental transcription factor by a dominant oncogene can promote malignancy, but provide opportunities for targeted therapy.

scholarly journals Regulation of σB by an Anti- and an Anti-Anti-Sigma Factor in Streptomyces coelicolor in Response to Osmotic Stress

2004 ◽  
Vol 186 (24) ◽  
pp. 8490-8498 ◽  
Author(s):  
Eun-Jin Lee ◽  
You-Hee Cho ◽  
Hyo-Sub Kim ◽  
Bo-Eun Ahn ◽  
Jung-Hye Roe
Keyword(s):  
Osmotic Stress ◽  
Sigma Factor ◽  
Target Genes ◽  
Sequence Similarity ◽  
Streptomyces Coelicolor ◽  
Dependent Manner ◽  
Its Gene ◽  
Genes Encoding

ABSTRACT σB, a homolog of stress-responsive σB of Bacillus subtilis, controls both osmoprotection and differentiation in Streptomyces coelicolor A3 (2). Its gene is preceded by rsbA and rsbB genes encoding homologs of an anti-sigma factor, RsbW, and its antagonist, RsbV, of B. subtilis, respectively. Purified RsbA bound to σB and prevented σB-directed transcription from the sigBp1 promoter in vitro. An rsbA-null mutant exhibited contrasting behavior to the sigB mutant, with elevated sigBp1 transcription, no actinorhodin production, and precocious aerial mycelial formation, reflecting enhanced activity of σB in vivo. Despite sequence similarity to RsbV, RsbB lacks the conserved phosphorylatable serine residue and its gene disruption produced no distinct phenotype. RsbV (SCO7325) from a putative six-gene operon (rsbV-rsbR-rsbS-rsbT-rsbU1-rsbU) was strongly induced by osmotic stress in a σB-dependent manner. It antagonized the inhibitory action of RsbA on σB-directed transcription and was phosphorylated by RsbA in vitro. These results support the hypothesis that the rapid induction of σB target genes by osmotic stress results from modulation of σB activity by the kinase-anti-sigma factor RsbA and its phosphorylatable antagonist RsbV, which function by a partner-switching mechanism. Amplified induction could result from a rapid increase in the synthesis of both σB and its inhibitor antagonist.

scholarly journals Transposon mobilization in the human fungal pathogen Cryptococcus is mutagenic during infection and promotes drug resistance in vitro

2020 ◽  
Vol 117 (18) ◽  
pp. 9973-9980 ◽  
Author(s):  
Asiya Gusa ◽  
Jonathan D. Williams ◽  
Jang-Eun Cho ◽  
Anna Floyd Averette ◽  
Sheng Sun ◽  
...  
Keyword(s):  
Drug Resistance ◽  
Target Genes ◽  
Opportunistic Infections ◽  
Temperature Shift ◽  
Antifungal Drugs ◽  
Rnai Pathway ◽  
Human Pathogens ◽  
Human Fungal Pathogen

When transitioning from the environment, pathogenic microorganisms must adapt rapidly to survive in hostile host conditions. This is especially true for environmental fungi that cause opportunistic infections in immunocompromised patients since these microbes are not well adapted human pathogens. Cryptococcus species are yeastlike fungi that cause lethal infections, especially in HIV-infected patients. Using Cryptococcus deneoformans in a murine model of infection, we examined contributors to drug resistance and demonstrated that transposon mutagenesis drives the development of 5-fluoroorotic acid (5FOA) resistance. Inactivation of target genes URA3 or URA5 primarily reflected the insertion of two transposable elements (TEs): the T1 DNA transposon and the TCN12 retrotransposon. Consistent with in vivo results, increased rates of mutagenesis and resistance to 5FOA and the antifungal drugs rapamycin/FK506 (rap/FK506) and 5-fluorocytosine (5FC) were found when Cryptococcus was incubated at 37° compared to 30° in vitro, a condition that mimics the temperature shift that occurs during the environment-to-host transition. Inactivation of the RNA interference (RNAi) pathway, which suppresses TE movement in many organisms, was not sufficient to elevate TE movement at 30° to the level observed at 37°. We propose that temperature-dependent TE mobilization in Cryptococcus is an important mechanism that enhances microbial adaptation and promotes pathogenesis and drug resistance in the human host.

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