scholarly journals Tuning theMycobacterium tuberculosisAlternative Sigma Factor SigF through the Multidomain Regulator Rv1364c and Osmosensory Kinase Protein Kinase D

2019 ◽  
Vol 201 (7) ◽  
Author(s):  
Richa Misra ◽  
Dilip Menon ◽  
Gunjan Arora ◽  
Richa Virmani ◽  
Mohita Gaur ◽  
...  
Keyword(s):  
Gene Expression ◽  
Mycobacterium Tuberculosis ◽  
Osmotic Stress ◽  
Cross Talk ◽  
Sigma Factor ◽  
Pathogenic Bacteria ◽  
Sigma Factors ◽  
Content Type ◽  
The Cross ◽  
Stress Dependent

ABSTRACTBacterial alternative sigma factors are mostly regulated by a partner-switching mechanism. Regulation of the virulence-associated alternative sigma factor SigF ofMycobacterium tuberculosishas been an area of intrigue, with SigF having more predicted regulators than other sigma factors in this organism. Rv1364c is one such predicted regulator, the mechanism of which is confounded by the presence of both anti-sigma factor and anti-sigma factor antagonist functions in a single polypeptide. Using protein binding and phosphorylation assays, we demonstrate that the anti-sigma factor domain of Rv1364c mediates autophosphorylation of its antagonist domain and binds efficiently to SigF. Furthermore, we identified a direct role for the osmosensor serine/threonine kinase PknD in regulating the SigF-Rv1364c interaction, adding to the current understanding about the intersection of these discrete signaling networks. Phosphorylation of SigF also showed functional implications in its DNA binding ability, which may help in activation of the regulon. InM. tuberculosis, osmotic stress-dependent induction ofespA, a SigF target involved in maintaining cell wall integrity, is curtailed upon overexpression of Rv1364c, showing its role as an anti-SigF factor. Overexpression of Rv1364c led to induction of another target,pks6, involved in lipid metabolism. This induction was, however, curtailed in the presence of osmotic stress conditions, suggesting modulation of SigF target gene expression via Rv1364c. These data provide evidence that Rv1364c acts an independent SigF regulator that is sensitive to the osmosensory signal, mediating the cross talk of PknD with the SigF regulon.IMPORTANCEMycobacterium tuberculosis, capable of latently infecting the host and causing aggressive tissue damage during active tuberculosis, is endowed with a complex regulatory capacity built of several sigma factors, protein kinases, and phosphatases. These proteins regulate expression of genes that allow the bacteria to adapt to various host-derived stresses, like nutrient starvation, acidic pH, and hypoxia. The cross talk between these systems is not well understood. SigF is one such regulator of gene expression that helpsM. tuberculosisto adapt to stresses and imparts virulence. This work provides evidence for its inhibition by the multidomain regulator Rv1364c and activation by the kinase PknD. The coexistence of negative and positive regulators of SigF in pathogenic bacteria reveals an underlying requirement for tight control of virulence factor expression.

scholarly journals Mycobacterium tuberculosis Transcription Machinery: Ready To Respond to Host Attacks

2016 ◽  
Vol 198 (9) ◽  
pp. 1360-1373 ◽  
Author(s):  
Kelly Flentie ◽  
Ashley L. Garner ◽  
Christina L. Stallings
Keyword(s):  
Gene Expression ◽  
Mycobacterium Tuberculosis ◽  
Stress Responses ◽  
Transcription Initiation ◽  
Pathogenic Bacteria ◽  
Hostile Environment ◽  
Tissue Architecture ◽  
Content Type ◽  
Response To Stress ◽  
Successful Colonization

Regulating responses to stress is critical for all bacteria, whether they are environmental, commensal, or pathogenic species. For pathogenic bacteria, successful colonization and survival in the host are dependent on adaptation to diverse conditions imposed by the host tissue architecture and the immune response. Once the bacterium senses a hostile environment, it must enact a change in physiology that contributes to the organism's survival strategy. Inappropriate responses have consequences; hence, the execution of the appropriate response is essential for survival of the bacterium in its niche. Stress responses are most often regulated at the level of gene expression and, more specifically, transcription. This minireview focuses on mechanisms of regulating transcription initiation that are required byMycobacterium tuberculosisto respond to the arsenal of defenses imposed by the host during infection. In particular, we highlight how certain features ofM. tuberculosisphysiology allow this pathogen to respond swiftly and effectively to host defenses. By enacting highly integrated and coordinated gene expression changes in response to stress,M. tuberculosisis prepared for battle against the host defense and able to persist within the human population.

scholarly journals Transcriptome Profiling of the Endophyte Burkholderia phytofirmans PsJN Indicates Sensing of the Plant Environment and Drought Stress

mBio ◽  
2015 ◽  
Vol 6 (5) ◽  
Author(s):  
Raheleh Sheibani-Tezerji ◽  
Thomas Rattei ◽  
Angela Sessitsch ◽  
Friederike Trognitz ◽  
Birgit Mitter
Keyword(s):  
Gene Expression ◽  
Drought Stress ◽  
Sigma Factor ◽  
Plant Stress ◽  
Sigma Factors ◽  
Content Type ◽  
Potato Plants ◽  
Ecf Sigma Factor ◽  
Plant Environment ◽  
Its Gene

ABSTRACT It is widely accepted that bacterial endophytes actively colonize plants, interact with their host, and frequently show beneficial effects on plant growth and health. However, the mechanisms of plant-endophyte communication and bacterial adaption to the plant environment are still poorly understood. Here, whole-transcriptome sequencing of B. phytofirmans PsJN colonizing potato (Solanum tuberosum L.) plants was used to analyze in planta gene activity and the response of strain PsJN to plant stress. The transcriptome of PsJN colonizing in vitro potato plants showed a broad array of functionalities encoded in the genome of strain PsJN. Transcripts upregulated in response to plant drought stress were mainly involved in transcriptional regulation, cellular homeostasis, and the detoxification of reactive oxygen species, indicating an oxidative stress response in PsJN. Genes with modulated expression included genes for extracytoplasmatic function (ECF) group IV sigma factors. These cell surface signaling elements allow bacteria to sense changing environmental conditions and to adjust their metabolism accordingly. TaqMan quantitative PCR (TaqMan-qPCR) was performed to identify ECF sigma factors in PsJN that were activated in response to plant stress. Six ECF sigma factor genes were expressed in PsJN colonizing potato plants. The expression of one ECF sigma factor was upregulated whereas that of another one was downregulated in a plant genotype-specific manner when the plants were stressed. Collectively, our study results indicate that endophytic B. phytofirmans PsJN cells are active inside plants. Moreover, the activity of strain PsJN is affected by plant drought stress; it senses plant stress signals and adjusts its gene expression accordingly. IMPORTANCE In recent years, plant growth-promoting endophytes have received steadily growing interest as an inexpensive alternative to resource-consuming agrochemicals in sustainable agriculture. Even though promising effects are recurrently observed under controlled conditions, these are rarely reproducible in the field or show undesirably strong variations. Obviously, a better understanding of endophyte activities in plants and the influence of plant physiology on these activities is needed to develop more-successful application strategies. So far, research has focused mainly on analyzing the plant response to bacterial inoculants. This prompted us to study the gene expression of the endophyte Burkholderia phytofirmans PsJN in potato plants. We found that endophytic PsJN cells express a wide array of genes and pathways, pointing to high metabolic activity inside plants. Moreover, the strain senses changes in the plant physiology due to plant stress and adjusts its gene expression pattern to cope with and adapt to the altered conditions.

scholarly journals A Tripartite, Hierarchical Sigma Factor Cascade Promotes Hormogonium Development in the Filamentous Cyanobacterium Nostoc punctiforme

mSphere ◽  
2019 ◽  
Vol 4 (3) ◽  
Author(s):  
Alfonso Gonzalez ◽  
Kelsey W. Riley ◽  
Thomas V. Harwood ◽  
Esthefani G. Zuniga ◽  
Douglas D. Risser
Keyword(s):  
Sigma Factor ◽  
Dominant Role ◽  
Sigma Factors ◽  
Oxygenic Photosynthesis ◽  
Model Organisms ◽  
Filamentous Cyanobacterium ◽  
Nostoc Punctiforme ◽  
Filamentous Cyanobacteria ◽  
Content Type ◽  
Nitrogen Cycles

ABSTRACT Cyanobacteria are prokaryotes capable of oxygenic photosynthesis, and frequently, nitrogen fixation as well. As a result, they contribute substantially to global primary production and nitrogen cycles. Furthermore, the multicellular filamentous cyanobacteria in taxonomic subsections IV and V are developmentally complex, exhibiting an array of differentiated cell types and filaments, including motile hormogonia, making them valuable model organisms for studying development. To investigate the role of sigma factors in the gene regulatory network (GRN) controlling hormogonium development, a combination of genetic, immunological, and time-resolved transcriptomic analyses were conducted in the model filamentous cyanobacterium Nostoc punctiforme, which, unlike other common model cyanobacteria, retains the developmental complexity of field isolates. The results support a model where the hormogonium GRN is driven by a hierarchal sigma factor cascade, with sigJ activating the expression of both sigC and sigF, as well as a substantial portion of additional hormogonium-specific genes, including those driving changes to cellular architecture. In turn, sigC regulates smaller subsets of genes for several processes, plays a dominant role in promoting reductive cell division, and may also both positively and negatively regulate sigJ to reinforce the developmental program and coordinate the timing of gene expression, respectively. In contrast, the sigF regulon is extremely limited. Among genes with characterized roles in hormogonium development, only pilA shows stringent sigF dependence. For sigJ-dependent genes, a putative consensus promoter was also identified, consisting primarily of a highly conserved extended −10 region, here designated a J-Box, which is widely distributed among diverse members of the cyanobacterial lineage. IMPORTANCE Cyanobacteria are integral to global carbon and nitrogen cycles, and their metabolic capacity coupled with their ease of genetic manipulation make them attractive platforms for applications such as biomaterial and biofertilizer production. Achieving these goals will likely require a detailed understanding and precise rewiring of these organisms’ GRNs. The complex phenotypic plasticity of filamentous cyanobacteria has also made them valuable models of prokaryotic development. However, current research has been limited by focusing primarily on a handful of model strains which fail to reflect the phenotypes of field counterparts, potentially limiting biotechnological advances and a more comprehensive understanding of developmental complexity. Here, using Nostoc punctiforme, a model filamentous cyanobacterium that retains the developmental range of wild isolates, we define previously unknown definitive roles for a trio of sigma factors during hormogonium development. These findings substantially advance our understanding of cyanobacterial development and gene regulation and could be leveraged for future applications.

scholarly journals Modulating Pathogenesis with Mobile-CRISPRi

2019 ◽  
Vol 201 (22) ◽  
Author(s):  
Jiuxin Qu ◽  
Neha K. Prasad ◽  
Michelle A. Yu ◽  
Shuyan Chen ◽  
Amy Lyden ◽  
...  
Keyword(s):  
Gene Expression ◽  
Pathogenic Bacteria ◽  
Bacterial Species ◽  
Effector Proteins ◽  
Infection Model ◽  
Secretion Systems ◽  
Bacterial Genomes ◽  
Gene Encoding ◽  
Content Type ◽  
Animal Infection

ABSTRACT Conditionally essential (CE) genes are required by pathogenic bacteria to establish and maintain infections. CE genes encode virulence factors, such as secretion systems and effector proteins, as well as biosynthetic enzymes that produce metabolites not found in the host environment. Due to their outsized importance in pathogenesis, CE gene products are attractive targets for the next generation of antimicrobials. However, the precise manipulation of CE gene expression in the context of infection is technically challenging, limiting our ability to understand the roles of CE genes in pathogenesis and accordingly design effective inhibitors. We previously developed a suite of CRISPR interference-based gene knockdown tools that are transferred by conjugation and stably integrate into bacterial genomes that we call Mobile-CRISPRi. Here, we show the efficacy of Mobile-CRISPRi in controlling CE gene expression in an animal infection model. We optimize Mobile-CRISPRi in Pseudomonas aeruginosa for use in a murine model of pneumonia by tuning the expression of CRISPRi components to avoid nonspecific toxicity. As a proof of principle, we demonstrate that knock down of a CE gene encoding the type III secretion system (T3SS) activator ExsA blocks effector protein secretion in culture and attenuates virulence in mice. We anticipate that Mobile-CRISPRi will be a valuable tool to probe the function of CE genes across many bacterial species and pathogenesis models. IMPORTANCE Antibiotic resistance is a growing threat to global health. To optimize the use of our existing antibiotics and identify new targets for future inhibitors, understanding the fundamental drivers of bacterial growth in the context of the host immune response is paramount. Historically, these genetic drivers have been difficult to manipulate precisely, as they are requisite for pathogen survival. Here, we provide the first application of Mobile-CRISPRi to study conditionally essential virulence genes in mouse models of lung infection through partial gene perturbation. We envision the use of Mobile-CRISPRi in future pathogenesis models and antibiotic target discovery efforts.

scholarly journals The Impact of Leadered and Leaderless Gene Structures on Translation Efficiency, Transcript Stability, and Predicted Transcription Rates in Mycobacterium smegmatis

2020 ◽  
Vol 202 (9) ◽  
Author(s):  
Tien G. Nguyen ◽  
Diego A. Vargas-Blanco ◽  
Louis A. Roberts ◽  
Scarlet S. Shell
Keyword(s):  
Gene Expression ◽  
Mycobacterium Tuberculosis ◽  
Mycobacterium Smegmatis ◽  
Half Life ◽  
Untranslated Regions ◽  
Translation Efficiency ◽  
Production Rates ◽  
Content Type ◽  
The Impact ◽  
Mrna Half Life

ABSTRACT Regulation of gene expression is critical for Mycobacterium tuberculosis to tolerate stressors encountered during infection and for nonpathogenic mycobacteria such as Mycobacterium smegmatis to survive environmental stressors. Unlike better-studied models, mycobacteria express ∼14% of their genes as leaderless transcripts. However, the impacts of leaderless transcript structures on mRNA half-life and translation efficiency in mycobacteria have not been directly tested. For leadered transcripts, the contributions of 5′ untranslated regions (UTRs) to mRNA half-life and translation efficiency are similarly unknown. In M. tuberculosis and M. smegmatis, the essential sigma factor, SigA, is encoded by a transcript with a relatively short half-life. We hypothesized that the long 5′ UTR of sigA causes this instability. To test this, we constructed fluorescence reporters and measured protein abundance, mRNA abundance, and mRNA half-life and calculated relative transcript production rates. The sigA 5′ UTR conferred an increased transcript production rate, shorter mRNA half-life, and decreased apparent translation rate compared to a synthetic 5′ UTR commonly used in mycobacterial expression plasmids. Leaderless transcripts appeared to be translated with similar efficiency as those with the sigA 5′ UTR but had lower predicted transcript production rates. A global comparison of M. tuberculosis mRNA and protein abundances failed to reveal systematic differences in protein/mRNA ratios for leadered and leaderless transcripts, suggesting that variability in translation efficiency is largely driven by factors other than leader status. Our data are also discussed in light of an alternative model that leads to different conclusions and suggests leaderless transcripts may indeed be translated less efficiently. IMPORTANCE Tuberculosis, caused by Mycobacterium tuberculosis, is a major public health problem killing 1.5 million people globally each year. During infection, M. tuberculosis must alter its gene expression patterns to adapt to the stress conditions it encounters. Understanding how M. tuberculosis regulates gene expression may provide clues for ways to interfere with the bacterium’s survival. Gene expression encompasses transcription, mRNA degradation, and translation. Here, we used Mycobacterium smegmatis as a model organism to study how 5′ untranslated regions affect these three facets of gene expression in multiple ways. We furthermore provide insight into the expression of leaderless mRNAs, which lack 5′ untranslated regions and are unusually prevalent in mycobacteria.

scholarly journals Transcriptional Regulation and Mechanism of SigN (ZpdN), a pBS32-Encoded Sigma Factor in Bacillus subtilis

mBio ◽  
2019 ◽  
Vol 10 (5) ◽  
Author(s):  
Aisha T. Burton ◽  
Aaron DeLoughery ◽  
Gene-Wei Li ◽  
Daniel B. Kearns
Keyword(s):  
Dna Damage ◽  
Bacillus Subtilis ◽  
Sigma Factor ◽  
Consensus Sequence ◽  
Sigma Factors ◽  
Sequencing Analysis ◽  
Dependent Manner ◽  
Content Type ◽  
Alternative Sigma Factors ◽  
Bona Fide

ABSTRACT Laboratory strains of Bacillus subtilis encode many alternative sigma factors, each dedicated to expressing a unique regulon such as those involved in stress resistance, sporulation, and motility. The ancestral strain of B. subtilis also encodes an additional sigma factor homolog, ZpdN, not found in lab strains due to being encoded on the large, low-copy-number plasmid pBS32, which was lost during domestication. DNA damage triggers pBS32 hyperreplication and cell death in a manner that depends on ZpdN, but how ZpdN mediates these effects is unknown. Here, we show that ZpdN is a bona fide sigma factor that can direct RNA polymerase to transcribe ZpdN-dependent genes, and we rename ZpdN SigN accordingly. Rend-seq (end-enriched transcriptome sequencing) analysis was used to determine the SigN regulon on pBS32, and the 5′ ends of transcripts were used to predict the SigN consensus sequence. Finally, we characterize the regulation of SigN itself and show that it is transcribed by at least three promoters: PsigN1, a strong SigA-dependent LexA-repressed promoter; PsigN2, a weak SigA-dependent constitutive promoter; and PsigN3, a SigN-dependent promoter. Thus, in response to DNA damage SigN is derepressed and then experiences positive feedback. How cells die in a pBS32-dependent manner remains unknown, but we predict that death is the product of expressing one or more genes in the SigN regulon. IMPORTANCE Sigma factors are utilized by bacteria to control and regulate gene expression. Some sigma factors are activated during times of stress to ensure the survival of the bacterium. Here, we report the presence of a sigma factor that is encoded on a plasmid that leads to cellular death after DNA damage.

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 Resilience in the Face of Uncertainty: Sigma Factor B Fine-Tunes Gene Expression To Support Homeostasis in Gram-Positive Bacteria

2016 ◽  
Vol 82 (15) ◽  
pp. 4456-4469 ◽  
Author(s):  
Claudia Guldimann ◽  
Kathryn J. Boor ◽  
Martin Wiedmann ◽  
Veronica Guariglia-Oropeza
Keyword(s):  
Gene Expression ◽  
Sigma Factor ◽  
Environmental Changes ◽  
Regulation Of Gene Expression ◽  
Bacterial Survival ◽  
Gram Positive ◽  
Content Type ◽  
Gram Positive Bacteria ◽  
Changing Environments ◽  
The Face

ABSTRACTGram-positive bacteria are ubiquitous and diverse microorganisms that can survive and sometimes even thrive in continuously changing environments. The key to such resilience is the ability of members of a population to respond and adjust to dynamic conditions in the environment. In bacteria, such responses and adjustments are mediated, at least in part, through appropriate changes in the bacterial transcriptome in response to the conditions encountered. Resilience is important for bacterial survival in diverse, complex, and rapidly changing environments and requires coordinated networks that integrate individual, mechanistic responses to environmental cues to enable overall metabolic homeostasis. In many Gram-positive bacteria, a key transcriptional regulator of the response to changing environmental conditions is the alternative sigma factor σB. σBhas been characterized in a subset of Gram-positive bacteria, including the generaBacillus,Listeria, andStaphylococcus. Recent insight from next-generation-sequencing results indicates that σB-dependent regulation of gene expression contributes to resilience, i.e., the coordination of complex networks responsive to environmental changes. This review explores contributions of σBto resilience inBacillus,Listeria, andStaphylococcusand illustrates recently described regulatory functions of σB.

scholarly journals Ankyrin-Like Protein AnkB Interacts with CatB, Affects Catalase Activity, and Enhances Resistance ofXanthomonas oryzaepv. oryzae andXanthomonas oryzaepv. oryzicola to Phenazine-1-Carboxylic Acid

2017 ◽  
Vol 84 (4) ◽  
Author(s):  
Xiayan Pan ◽  
Shu Xu ◽  
Jian Wu ◽  
Yabing Duan ◽  
Zhitian Zheng ◽  
...  
Keyword(s):  
Gene Expression ◽  
Carboxylic Acid ◽  
Catalase Activity ◽  
Pathogenic Bacteria ◽  
Adaptor Protein ◽  
Bacterial Biofilm ◽  
Xanthomonas Oryzae ◽  
Content Type ◽  
Plant Pathogenic Bacteria ◽  
Ankyrin Protein

ABSTRACTXanthomonas oryzaepv. oryzae, which causes rice bacterial leaf blight, andXanthomonas oryzaepv. oryzicola, which causes rice bacterial leaf streak, are important plant-pathogenic bacteria. A member of the adaptor protein family, ankyrin protein, has been investigated largely in humans but rarely in plant-pathogenic bacteria. In this study, a novel ankyrin-like protein, AnkB, was identified inX. oryzaepv. oryzae andX. oryzaepv. oryzicola. The expression ofankBwas significantly upregulated when these bacteria were treated with phenazine-1-carboxylic acid (PCA).ankBis located 58 bp downstream of the genecatB(which encodes a catalase) in both bacteria, and the gene expression ofcatBand catalase activity were reduced followingankBdeletion inX. oryzaepv. oryzae andX. oryzaepv. oryzicola. Furthermore, we demonstrated that AnkB directly interacts with CatB by glutathioneS-transferase (GST) pulldown assays. Deletion ofankBincreased the sensitivity ofX. oryzaepv. oryzae andX. oryzaepv. oryzicola to H2O2and PCA, decreased bacterial biofilm formation, swimming ability, and exopolysaccharide (EPS) production, and also reduced virulence on rice. Together our results indicate that the ankyrin-like protein AnkB has important and conserved roles in antioxidant systems and pathogenicity inX. oryzaepv. oryzae andX. oryzaepv. oryzicola.IMPORTANCEThis study demonstrates that the ankyrin protein AnkB directly interacts with catalase CatB inXanthomonas oryzaepv. oryzae andXanthomonas oryzaepv. oryzicola. Ankyrin protein AnkB can affect the gene expression ofcatB, catalase activity, and sensitivity to H2O2. InXanthomonasspp., the locations of genesankBandcatBand the amino acid sequence of AnkB are highly conserved. It is suggested that in prokaryotes, AnkB plays a conserved role in the defense against oxidative stress.

Sign in / Sign up

Export Citation Format

Share Document