Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 6.250
Filtrar
1.
PLoS Genet ; 20(3): e1011059, 2024 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-38466775

RESUMO

RpoS is an alternative sigma factor needed for the induction of the general stress response in many gammaproteobacteria. Tight regulation of RpoS levels and activity is required for bacterial growth and survival under stress. In Escherichia coli, various stresses lead to higher levels of RpoS due to increased translation and decreased degradation. During non-stress conditions, RpoS is unstable, because the adaptor protein RssB delivers RpoS to the ClpXP protease. RpoS degradation is prevented during stress by the sequestration of RssB by anti-adaptors, each of which is induced in response to specific stresses. Here, we examined how the stabilization of RpoS is reversed during recovery of the cell from stress. We found that RpoS degradation quickly resumes after recovery from phosphate starvation, carbon starvation, and when transitioning from stationary phase back to exponential phase. This process is in part mediated by the anti-adaptor IraP, known to promote RpoS stabilization during phosphate starvation via the sequestration of adaptor RssB. The rapid recovery from phosphate starvation is dependent upon a feedback loop in which RpoS transcription of rssB, encoding the adaptor protein, plays a critical role. Crl, an activator of RpoS that specifically binds to and stabilizes the complex between the RNA polymerase and RpoS, is also required for the feedback loop to function efficiently, highlighting a critical role for Crl in restoring RpoS basal levels.


Assuntos
Proteínas de Escherichia coli , Escherichia coli , Escherichia coli/metabolismo , Proteínas de Escherichia coli/genética , Proteínas de Escherichia coli/metabolismo , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Retroalimentação , Fator sigma/genética , Fator sigma/metabolismo , Fosfatos/metabolismo , Regulação Bacteriana da Expressão Gênica
2.
Proc Natl Acad Sci U S A ; 121(13): e2400584121, 2024 Mar 26.
Artigo em Inglês | MEDLINE | ID: mdl-38502707

RESUMO

When faced with starvation, the bacterium Bacillus subtilis transforms itself into a dormant cell type called a "spore". Sporulation initiates with an asymmetric division event, which requires the relocation of the core divisome components FtsA and FtsZ, after which the sigma factor σF is exclusively activated in the smaller daughter cell. Compartment-specific activation of σF requires the SpoIIE phosphatase, which displays a biased localization on one side of the asymmetric division septum and associates with the structural protein DivIVA, but the mechanism by which this preferential localization is achieved is unclear. Here, we isolated a variant of DivIVA that indiscriminately activates σF in both daughter cells due to promiscuous localization of SpoIIE, which was corrected by overproduction of FtsA and FtsZ. We propose that the core components of the redeployed cell division machinery drive the asymmetric localization of DivIVA and SpoIIE to trigger the initiation of the sporulation program.


Assuntos
Bacillus subtilis , Proteínas de Bactérias , Bacillus subtilis/metabolismo , Ativação Transcricional , Proteínas de Bactérias/metabolismo , Esporos Bacterianos/genética , Esporos Bacterianos/metabolismo , Divisão Celular/genética , Fator sigma/genética , Fator sigma/metabolismo
3.
Proc Natl Acad Sci U S A ; 121(10): e2321910121, 2024 Mar 05.
Artigo em Inglês | MEDLINE | ID: mdl-38422018

RESUMO

Bacteroidota are abundant members of the human gut microbiota that shape the enteric landscape by modulating host immunity and degrading dietary- and host-derived glycans. These processes are mediated in part by Outer Membrane Vesicles (OMVs). Here, we developed a high-throughput screen to identify genes required for OMV biogenesis and its regulation in Bacteroides thetaiotaomicron (Bt). We identified a family of Dual membrane-spanning anti-sigma factors (Dma) that control OMV biogenesis. We conducted molecular and multiomic analyses to demonstrate that deletion of Dma1, the founding member of the Dma family, modulates OMV production by controlling the activity of the ECF21 family sigma factor, Das1, and its downstream regulon. Dma1 has a previously uncharacterized domain organization that enables Dma1 to span both the inner and outer membrane of Bt. Phylogenetic analyses reveal that this common feature of the Dma family is restricted to the phylum Bacteroidota. This study provides mechanistic insights into the regulation of OMV biogenesis in human gut bacteria.


Assuntos
Bacteroides thetaiotaomicron , Humanos , Bacteroides thetaiotaomicron/genética , Fator sigma , Filogenia
4.
FEMS Microbiol Rev ; 48(2)2024 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-38383636

RESUMO

Promoter sequences are important genetic control elements. Through their interaction with RNA polymerase they determine transcription strength and specificity, thereby regulating the first step in gene expression. Consequently, they can be targeted as elements to control predictability and tuneability of a genetic circuit, which is essential in applications such as the development of robust microbial cell factories. This review considers the promoter elements implicated in the three stages of transcription initiation, detailing the complex interplay of sequence-specific interactions that are involved, and highlighting that DNA sequence features beyond the core promoter elements work in a combinatorial manner to determine transcriptional strength. In particular, we emphasize that, aside from promoter recognition, transcription initiation is also defined by the kinetics of open complex formation and promoter escape, which are also known to be highly sequence specific. Significantly, we focus on how insights into these interactions can be manipulated to lay the foundation for a more rational approach to promoter engineering.


Assuntos
RNA Polimerases Dirigidas por DNA , Transcrição Gênica , Regiões Promotoras Genéticas/genética , Transcrição Gênica/genética , RNA Polimerases Dirigidas por DNA/genética , RNA Polimerases Dirigidas por DNA/metabolismo , Regulação da Expressão Gênica , DNA , Fator sigma/genética , Fator sigma/metabolismo
5.
Nature ; 627(8003): 424-430, 2024 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-38418874

RESUMO

Mycobacterium tuberculosis (Mtb) is a bacterial pathogen that causes tuberculosis (TB), an infectious disease that is responsible for major health and economic costs worldwide1. Mtb encounters diverse environments during its life cycle and responds to these changes largely by reprogramming its transcriptional output2. However, the mechanisms of Mtb transcription and how they are regulated remain poorly understood. Here we use a sequencing method that simultaneously determines both termini of individual RNA molecules in bacterial cells3 to profile the Mtb transcriptome at high resolution. Unexpectedly, we find that most Mtb transcripts are incomplete, with their 5' ends aligned at transcription start sites and 3' ends located 200-500 nucleotides downstream. We show that these short RNAs are mainly associated with paused RNA polymerases (RNAPs) rather than being products of premature termination. We further show that the high propensity of Mtb RNAP to pause early in transcription relies on the binding of the σ-factor. Finally, we show that a translating ribosome promotes transcription elongation, revealing a potential role for transcription-translation coupling in controlling Mtb gene expression. In sum, our findings depict a mycobacterial transcriptome that prominently features incomplete transcripts resulting from RNAP pausing. We propose that the pausing phase constitutes an important transcriptional checkpoint in Mtb that allows the bacterium to adapt to environmental changes and could be exploited for TB therapeutics.


Assuntos
Regulação Bacteriana da Expressão Gênica , Mycobacterium tuberculosis , RNA Bacteriano , Transcriptoma , RNA Polimerases Dirigidas por DNA/metabolismo , Mycobacterium tuberculosis/genética , Mycobacterium tuberculosis/metabolismo , RNA Bacteriano/análise , RNA Bacteriano/biossíntese , RNA Bacteriano/genética , Transcriptoma/genética , Tuberculose/microbiologia , RNA Mensageiro/análise , RNA Mensageiro/biossíntese , RNA Mensageiro/genética , Sítio de Iniciação de Transcrição , Fator sigma/metabolismo , Ribossomos/metabolismo , Biossíntese de Proteínas
6.
Nat Struct Mol Biol ; 31(1): 141-149, 2024 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-38177674

RESUMO

Gene expression in Escherichia coli is controlled by well-established mechanisms that activate or repress transcription. Here, we identify CedA as an unconventional transcription factor specifically associated with the RNA polymerase (RNAP) σ70 holoenzyme. Structural and biochemical analysis of CedA bound to RNAP reveal that it bridges distant domains of ß and σ70 subunits to stabilize an open-promoter complex. CedA does so without contacting DNA. We further show that cedA is strongly induced in response to amino acid starvation, oxidative stress and aminoglycosides. CedA provides a basal level of tolerance to these clinically relevant antibiotics, as well as to rifampicin and peroxide. Finally, we show that CedA modulates transcription of hundreds of bacterial genes, which explains its pleotropic effect on cell physiology and pathogenesis.


Assuntos
Proteínas de Escherichia coli , Fatores Genéricos de Transcrição , Escherichia coli/metabolismo , Fator sigma/química , Fator sigma/genética , Fator sigma/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Proteínas de Escherichia coli/metabolismo , RNA Polimerases Dirigidas por DNA/metabolismo , Fatores Genéricos de Transcrição/genética , Fatores Genéricos de Transcrição/metabolismo , Transcrição Gênica , Proteínas de Bactérias/metabolismo
7.
Commun Biol ; 7(1): 46, 2024 01 06.
Artigo em Inglês | MEDLINE | ID: mdl-38184746

RESUMO

σ factors are considered as positive regulators of gene expression. Here we reveal the opposite, inhibitory role of these proteins. We used a combination of molecular biology methods and computational modeling to analyze the regulatory activity of the extracytoplasmic σE factor from Streptomyces coelicolor. The direct activator/repressor function of σE was then explored by experimental analysis of selected promoter regions in vivo. Additionally, the σE interactome was defined. Taken together, the results characterize σE, its regulation, regulon, and suggest its direct inhibitory function (as a repressor) in gene expression, a phenomenon that may be common also to other σ factors and organisms.


Assuntos
Streptomyces coelicolor , Streptomyces coelicolor/genética , Simulação por Computador , Fator sigma/genética
8.
Mol Microbiol ; 121(2): 291-303, 2024 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-38169053

RESUMO

Pseudomonas aeruginosa is an important opportunistic pathogen. Several of its virulence-related processes, including the synthesis of pyocyanin (PYO) and biofilm formation, are controlled by quorum sensing (QS). It has been shown that the alternative sigma factor RpoS regulates QS through the reduction of lasR and rhlR transcription (encoding QS regulators). However, paradoxically, the absence of RpoS increases PYO production and biofilm development (that are RhlR dependent) by unknown mechanisms. Here, we show that RpoS represses pqsE transcription, which impacts the stability and activity of RhlR. In the absence of RpoS, rhlR transcript levels are reduced but not the RhlR protein concentration, presumably by its stabilization by PqsE, whose expression is increased. We also report that PYO synthesis and the expression of pqsE and phzA1B1C1D1E1F1G1 operon exhibit the same pattern at different RpoS concentrations, suggesting that the RpoS-dependent PYO production is due to its ability to modify PqsE concentration, which in turn modulates the activation of the phzA1 promoter by RhlR. Finally, we demonstrate that RpoS favors the expression of Vfr, which activates the transcription of lasR and rhlR. Our study contributes to the understanding of how RpoS modulates the QS response in P. aeruginosa, exerting both negative and positive regulation.


Assuntos
Percepção de Quorum , Fator sigma , Percepção de Quorum/genética , Fator sigma/genética , Fator sigma/metabolismo , Pseudomonas aeruginosa/metabolismo , Biofilmes , Piocianina , Óperon , Proteínas de Bactérias/metabolismo , Regulação Bacteriana da Expressão Gênica
9.
J Bacteriol ; 206(2): e0035523, 2024 02 22.
Artigo em Inglês | MEDLINE | ID: mdl-38197669

RESUMO

In Escherichia coli, one of the best understood microorganisms, much can still be learned about the basic interactions between transcription factors and promoters. When a cAMP-deficient cya mutant is supplied with maltose as the main carbon source, mutations develop upstream from the two genes malT and sdaC. Here, we explore the regulation of the two promoters, using fluorescence-based genetic reporters in combination with both spontaneously evolved and systematically engineered cis-acting mutations. We show that in the cya mutant, regulation of malT and sdaC evolves toward cAMP-independence and increased expression in the stationary phase. Furthermore, we show that the location of the cAMP receptor protein (Crp) binding site upstream of malT is important for alternative sigma factor usage. This provides new insights into the architecture of bacterial promoters and the global interplay between Crp and sigma factors in different growth phases.IMPORTANCEThis work provides new general insights into (1) the architecture of bacterial promoters, (2) the importance of the location of Class I Crp-dependent promoters, and (3) the global interplay between Crp and sigma factors in different growth phases.


Assuntos
Proteínas de Escherichia coli , Escherichia coli , Proteínas de Bactérias/metabolismo , Proteína Receptora de AMP Cíclico/genética , Proteína Receptora de AMP Cíclico/metabolismo , Escherichia coli/genética , Escherichia coli/metabolismo , Proteínas de Escherichia coli/genética , Proteínas de Escherichia coli/metabolismo , Regulação Bacteriana da Expressão Gênica , Mutação , Fator sigma/genética , Fator sigma/metabolismo , Transcrição Gênica
10.
Appl Environ Microbiol ; 90(2): e0203923, 2024 Feb 21.
Artigo em Inglês | MEDLINE | ID: mdl-38259108

RESUMO

The build-up of formaldehyde, a highly reactive molecule is cytotoxic and must be eliminated for the organism's survival. Formaldehyde detoxification system is found in nearly all organisms including both pathogenic and non-pathogenic mycobacteria. MscR, a formaldehyde dehydrogenase from Mycobacterium smegmatis (Msm), is an indispensable part of this system and forms a bicistronic operon with its downstream uncharacterized gene, fmh. We here show that Fmh, a putative metallo-beta-lactamase, is essential in tolerating higher amounts of formaldehyde when co-overexpressed with mscR in vivo. Our NMR studies indicate that MscR, along with Fmh, enhances formate production through a mycothiol (MSH)-dependent pathway, emphasizing the importance of Fmh in detoxifying formaldehyde. Although another aldehyde dehydrogenase, MSMEG_1543, induces upon formaldehyde addition, it is not involved in its detoxification. We also show that the expression of the mscR operon is constitutive and remains unchanged upon formaldehyde addition, as displayed by the promoter activity of PmscR and by the transcript and protein levels of MscR. Furthermore, we establish the role of a thiol-responsive sigma factor SigH in formaldehyde detoxification. We show that SigH, and not SigE, is crucial for formaldehyde detoxification, even though it does not directly regulate mscR operon expression. In addition, sensitivity to formaldehyde in sigH-knockout could be alleviated by overexpression of mscR. Taken together, our data demonstrate the importance of MSH-dependent pathways in detoxifying formaldehyde in a mycobacterial system. An absence of such MSH-dependent proteins in eukaryotes and its complete conservation in M. tuberculosis, the causative agent of tuberculosis, further unravel new drug targets for this pathogen.IMPORTANCEExtensive research has been done on formaldehyde detoxification in different bacteria. However, our current understanding of the mechanisms underlying this process in mycobacteria remains exceedingly little. We previously showed that MscR, a formaldehyde dehydrogenase from Mycobacterium smegmatis, plays a pivotal role in this detoxification pathway. Here, we present a potential S-formyl-mycothiol hydrolase named Fmh, thought to be a metallo-beta-lactamase, which functions along with mycothiol (MSH) and MscR to enhance formate production within this detoxification pathway. Co-expression of Fmh with MscR significantly enhances the efficiency of formaldehyde detoxification in M. smegmatis. Our experiments establish that Fmh catalyzes the final step of this detoxification pathway. Although an alternative sigma factor SigH was found to be involved in formaldehyde detoxification, it did not directly regulate the expression of mscR. Since formaldehyde detoxification is essential for bacterial survival, we envisage this process to be a potential drug target for M. tuberculosis eradication.


Assuntos
Cisteína , Glicopeptídeos , Inositol , Mycobacterium tuberculosis , Tuberculose , Humanos , Mycobacterium smegmatis/genética , Mycobacterium smegmatis/metabolismo , Fator sigma/genética , Mycobacterium tuberculosis/genética , Mycobacterium tuberculosis/metabolismo , Formaldeído/metabolismo , beta-Lactamases/metabolismo , Formiatos/metabolismo , Proteínas de Bactérias/metabolismo
11.
ACS Synth Biol ; 13(1): 402-407, 2024 Jan 19.
Artigo em Inglês | MEDLINE | ID: mdl-38176073

RESUMO

Expanding sigma70 promoter libraries can support the engineering of metabolic pathways and enhance recombinant protein expression. Herein, we developed an artificial intelligence (AI) and knowledge-based method for the rational design of sigma70 promoters. Strong sigma70 promoters were identified by using high-throughput screening (HTS) with enhanced green fluorescent protein (eGFP) as a reporter gene. The features of these strong promoters were adopted to guide promoter design based on our previous reported deep learning model. In the following case study, the obtained strong promoters were used to express collagen and microbial transglutaminase (mTG), resulting in increased expression levels by 81.4% and 33.4%, respectively. Moreover, these constitutive promoters achieved soluble expression of mTG-activating protease and contributed to active mTG expression in Escherichia coli. The results suggested that the combined method may be effective for promoter engineering.


Assuntos
Inteligência Artificial , Escherichia coli , Escherichia coli/genética , Escherichia coli/metabolismo , RNA Polimerases Dirigidas por DNA/genética , Fator sigma/genética , Fator sigma/metabolismo , Regiões Promotoras Genéticas/genética
12.
mSphere ; 9(2): e0071923, 2024 Feb 28.
Artigo em Inglês | MEDLINE | ID: mdl-38236030

RESUMO

The Gram-positive model organism Bacillus subtilis responds to environmental stressors by activating the alternative sigma factor σB. The sensing apparatus upstream of σB activation is thought to consist of cytoplasmic stressosomes-megadalton-sized protein complexes that include five paralogous proteins known as RsbRs. The RsbRs are presumed to be involved in stress sensing and the subsequent response. Perturbations to the RsbR complement in stressosomes by engineering cells that produce only one of the RsbR paralogs ("single-RsbR strains") lead to altered σB response dynamics with respect to timing and magnitude. Here, we asked whether such changes to σB response dynamics impact the relative fitness of a strain. We competed strain pairs with different RsbR complements under ethanol and sodium chloride stress and found not only differences in relative fitness among wild-type and single-RsbR strains but also different relative fitness values in the two different stressors. We found that the presence of RsbRA, which dominates the wild-type σB response, enhances fitness in ethanol but is detrimental to fitness in NaCl. Meanwhile, RsbRD-only cells were among the most fit in NaCl. Strains producing hybrid RsbR fusion proteins displayed different fitness values that depended on the RsbR proteins from which they were derived. Our results here suggest that σB response dynamics can impact fitness, highlighting the physiological importance of the unusual stressosome-based general stress response system of B. subtilis. IMPORTANCE: The model bacterium Bacillus subtilis uses cytoplasmic multiprotein complexes, termed stressosomes, to activate the alternative sigma factor σB when facing environmental stresses. We have previously shown that genetically manipulating the complement of putative sensor proteins in stressosomes can alter the dynamics of the σB response in terms of its magnitude and timing. However, it is unknown whether these response dynamics impact the fitness of cells challenged by environmental stressors. Here, we examine the fitness of strains with different σB responses by competing strain pairs in exponential-phase co-cultures under environmental stress. We find that strains with different response dynamics show different competitive indices that differ by stressor. These results suggest that the dynamics of the σB response can affect the fitness of cells facing environmental stress, highlighting the relevance of different σB dynamics.


Assuntos
Bacillus subtilis , Fator sigma , Fator sigma/genética , Fator sigma/metabolismo , Bacillus subtilis/metabolismo , Cloreto de Sódio , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Fosfoproteínas , Etanol
13.
Proc Natl Acad Sci U S A ; 121(2): e2309670120, 2024 Jan 09.
Artigo em Inglês | MEDLINE | ID: mdl-38170755

RESUMO

Gene transcription is a fundamental cellular process carried out by RNA polymerase (RNAP). Transcription initiation is highly regulated, and in bacteria, transcription initiation is mediated by sigma (σ) factors. σ recruits RNAP to the promoter DNA region, located upstream of the transcription start site (TSS) and facilitates open complex formation, where double-stranded DNA is opened up into a transcription bubble and template strand DNA is positioned inside RNAP for initial RNA synthesis. During initial transcription, RNAP remains bound to σ and upstream DNA, presumably with an enlarging transcription bubble. The release of RNAP from upstream DNA is required for promoter escape and processive transcription elongation. Bacteria sigma factors can be broadly separated into two classes with the majority belonging to the σ70 class, represented by the σ70 that regulates housekeeping genes. σ54 forms a class on its own and regulates stress response genes. Extensive studies on σ70 have revealed the molecular mechanisms of the σ70 dependent process while how σ54 transitions from initial transcription to elongation is currently unknown. Here, we present a series of cryo-electron microscopy structures of the RNAP-σ54 initial transcribing complexes with progressively longer RNA, which reveal structural changes that lead to promoter escape. Our data show that initially, the transcription bubble enlarges, DNA strands scrunch, reducing the interactions between σ54 and DNA strands in the transcription bubble. RNA extension and further DNA scrunching help to release RNAP from σ54 and upstream DNA, enabling the transition to elongation.


Assuntos
Escherichia coli , Transcrição Gênica , Microscopia Crioeletrônica , Escherichia coli/genética , Escherichia coli/metabolismo , Regiões Promotoras Genéticas/genética , RNA Polimerases Dirigidas por DNA/metabolismo , DNA/metabolismo , RNA/metabolismo , Bactérias/metabolismo , Fator sigma/metabolismo , DNA Bacteriano/metabolismo
14.
J Gen Appl Microbiol ; 69(4): 215-228, 2024 Feb 02.
Artigo em Inglês | MEDLINE | ID: mdl-37380492

RESUMO

When Bacillus subtilis cells face environmental deterioration, such as exhaustion of nutrients and an increase in cell density, they form spores. It is known that phosphorylation of Spo0A and activation of σH are key events at the initiation of sporulation. However, the initiation of sporulation is an extremely complicated process, and the relationship between these two events remains to be elucidated. To determine the minimum requirements for triggering sporulation initiation, we attempted to induce cell sporulation at the log phase, regardless of nutrients and cell density. In rich media such as Luria-Bertani (LB) medium, the cells of B. subtilis do not sporulate efficiently, possibly because of excess nutrition. When the amount of xylose in the LB medium was limited, σH -dependent transcription of the strain, in which sigA was under the control of the xylose-inducible promoter, was induced, and the frequency of sporulation was elevated according to the decreased level of σA. We also employed a fusion of sad67, which codes for an active form of Spo0A, and the IPTG-inducible promoter. The combination of lowered σA expression and activated Spo0A allowed the cells in the log phase to stop growing and rush into spore development. This observation of enforced initiation of sporulation in the mutant strain was detected even in the presence of the wild-type strain, suggesting that only intracellular events initiate and fulfill spore development regardless of extracellular conditions. Under natural sporulation conditions, the amount of σA did not change drastically throughout growth. Mechanisms that sequester σA from the core RNA polymerase and help σH to become active exist, but this has not yet been elucidated.


Assuntos
Bacillus subtilis , Fatores de Transcrição , Bacillus subtilis/genética , Fatores de Transcrição/genética , Fator sigma/genética , Fator sigma/metabolismo , Xilose , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Esporos Bacterianos/genética , Regulação Bacteriana da Expressão Gênica , Transcrição Gênica
15.
Mol Microbiol ; 121(2): 167-195, 2024 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-37908155

RESUMO

Legionella pneumophila is a gram-negative bacteria found in natural and anthropogenic aquatic environments such as evaporative cooling towers, where it reproduces as an intracellular parasite of cohabiting protozoa. If L. pneumophila is aerosolized and inhaled by a susceptible person, bacteria may colonize their alveolar macrophages causing the opportunistic pneumonia Legionnaires' disease. L. pneumophila utilizes an elaborate regulatory network to control virulence processes such as the Dot/Icm Type IV secretion system and effector repertoire, responding to changing nutritional cues as their host becomes depleted. The bacteria subsequently differentiate to a transmissive state that can survive in the environment until a replacement host is encountered and colonized. In this review, we discuss the lifecycle of L. pneumophila and the molecular regulatory network that senses nutritional depletion via the stringent response, a link to stationary phase-like metabolic changes via alternative sigma factors, and two-component systems that are homologous to stress sensors in other pathogens, to regulate differentiation between the intracellular replicative phase and more transmissible states. Together, we highlight how this prototypic intracellular pathogen offers enormous potential in understanding how molecular mechanisms enable intracellular parasitism and pathogenicity.


Assuntos
Legionella pneumophila , Humanos , Legionella pneumophila/genética , Legionella pneumophila/metabolismo , Virulência , Fator sigma/metabolismo , Proteínas de Bactérias/metabolismo
16.
Bioorg Chem ; 143: 106983, 2024 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-38016396

RESUMO

RNA polymerase is an essential enzyme involved in bacterial transcription, playing a crucial role in RNA synthesis. However, it requires the association with sigma factors to initiate this process. In our previous work, we utilized a structure-based drug discovery approach to create benzoyl and benzyl benzoic acid compounds. These compounds were designed based on the amino acid residues within the key binding site of sigma factors, which are crucial for their interaction with RNA polymerase. By inhibiting bacterial transcription, these compounds exhibited notable antimicrobial activity, and we coined them as sigmacidins to highlight their resemblance to sigma factors and the benzoic acid structure. In this study, we further modified the compound scaffolds and developed a series of sulfonamidyl benzoic acid derivatives. These derivatives displayed potent antimicrobial activity, with minimum inhibitory concentrations (MICs) as low as 1 µg/mL, demonstrating their efficacy against bacteria. Furthermore, these compounds demonstrated low cytotoxicity, indicating their potential as safe antimicrobial agents. To ascertain their mechanism of action in interfering with bacterial transcription, we conducted biochemical and cellular assays. Overall, this study showcases the effectiveness of sulfonamidyl benzoic acid derivatives as antimicrobial agents by targeting protein-protein interactions involving RNA polymerase and sigma factors. Their strong antimicrobial activity and low cytotoxicity implicate their potential in combating antibiotic-resistant bacteria.


Assuntos
Antibacterianos , Anti-Infecciosos , Antibacterianos/farmacologia , Antibacterianos/química , Fator sigma/metabolismo , RNA Polimerases Dirigidas por DNA/metabolismo , Bactérias/metabolismo , Ácido Benzoico/farmacologia , Testes de Sensibilidade Microbiana
17.
Int J Biol Macromol ; 254(Pt 1): 127833, 2024 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-37918595

RESUMO

Vibrio species are motile gram-negative bacteria commonly found in aquatic environments. Vibrio species include pathogenic as well as non-pathogenic strains. Pathogenic Vibrio species have been reported in invertebrates and humans, whereas non-pathogenic strains are involved in symbiotic relationships with their eukaryotic hosts. These bacteria are also able to adapt to fluctuations in temperature, salinity, and pH, in addition to oxidative stress, and osmotic pressure in aquatic ecosystems. Moreover, they have also developed protective mechanisms against the immune systems of their hosts. Vibrio species accomplish adaptation to changing environments outside or inside the host by altering their gene expression profiles. To this end, several sigma factors specifically regulate gene expression, particularly under stressful environmental conditions. Moreover, other sigma factors are associated with biofilm formation and virulence as well. This review discusses different types of sigma and anti-sigma factors of Vibrio species involved in virulence and regulation of gene expression upon changes in environmental conditions. The evolutionary relationships between sigma factors with various physiological roles in Vibrio species are also discussed extensively.


Assuntos
Fator sigma , Vibrio , Humanos , Fator sigma/genética , Fator sigma/metabolismo , Ecossistema , Vibrio/metabolismo , Estresse Oxidativo , Virulência/genética , Regulação Bacteriana da Expressão Gênica
18.
mBio ; 15(1): e0203623, 2024 Jan 16.
Artigo em Inglês | MEDLINE | ID: mdl-38112466

RESUMO

IMPORTANCE: Hallmarks of the developmental cycle of the obligate intracellular pathogenic bacterium Chlamydia are the primary differentiation of the infectious elementary body (EB) into the proliferative reticulate body (RB) and the secondary differentiation of RBs back into EBs. The mechanisms regulating these transitions remain unclear. In this report, we developed an effective novel strategy termed dependence on plasmid-mediated expression (DOPE) that allows for the knockdown of essential genes in Chlamydia. We demonstrate that GrgA, a Chlamydia-specific transcription factor, is essential for the secondary differentiation and optimal growth of RBs. We also show that GrgA, a chromosome-encoded regulatory protein, controls the maintenance of the chlamydial virulence plasmid. Transcriptomic analysis further indicates that GrgA functions as a critical regulator of all three sigma factors that recognize different promoter sets at developmental stages. The DOPE strategy outlined here should provide a valuable tool for future studies examining chlamydial growth, development, and pathogenicity.


Assuntos
Infecções por Chlamydia , Chlamydia trachomatis , Humanos , Chlamydia trachomatis/metabolismo , Regulação Bacteriana da Expressão Gênica , Fatores de Transcrição/metabolismo , Fator sigma/genética , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo
19.
J Biol Chem ; 300(2): 105568, 2024 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-38103640

RESUMO

Upon Mg2+ starvation, a condition often associated with virulence, enterobacteria inhibit the ClpXP-dependent proteolysis of the master transcriptional regulator, σs, via IraM, a poorly understood antiadaptor that prevents RssB-dependent loading of σs onto ClpXP. This inhibition results in σs accumulation and expression of stress resistance genes. Here, we report on the structural analysis of RssB bound to IraM, which reveals that IraM induces two folding transitions within RssB, amplified via a segmented helical linker. These conformational changes result in an open, yet inhibited RssB structure in which IraM associates with both the C-terminal and N-terminal domains of RssB and prevents binding of σs to the 4-5-5 face of the N-terminal receiver domain. This work highlights the remarkable structural plasticity of RssB and reveals how a stress-specific RssB antagonist modulates a core stress response pathway that could be leveraged to control biofilm formation, virulence, and the development of antibiotic resistance.


Assuntos
Proteínas de Escherichia coli , Modelos Moleculares , Fatores de Transcrição , Endopeptidase Clp/metabolismo , Escherichia coli/química , Escherichia coli/metabolismo , Proteínas de Escherichia coli/química , Proteínas de Escherichia coli/metabolismo , Fosforilação , Ligação Proteica , Domínios Proteicos , Dobramento de Proteína , Estrutura Terciária de Proteína , Fator sigma/metabolismo , Fatores de Transcrição/química , Fatores de Transcrição/metabolismo
20.
Mol Microbiol ; 121(2): 243-259, 2024 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-38153189

RESUMO

The intracellular pathogen Legionella pneumophila translocates more than 300 effector proteins into its host cells. The expression levels of the genes encoding these effectors are orchestrated by an intricate regulatory network. Here, we introduce LelA, the first L. pneumophila LysR-type transcriptional regulator of effectors. Through bioinformatic and experimental analyses, we identified the LelA target regulatory element and demonstrated that it directly activates the expression of three L. pneumophila effectors (legL7, legL6, and legU1). We further found that the gene encoding LelA is positively regulated by the RpoS sigma factor, thus linking it to the known effector regulatory network. Examination of other species throughout the Legionella genus revealed that this regulatory element is found upstream of 34 genes encoding validated effectors, putative effectors, and hypothetical proteins. Moreover, ten of these genes were examined and found to be activated by the L. pneumophila LelA as well as by their orthologs in the corresponding species. LelA represents a novel type of Legionella effector regulator, which coordinates the expression of both adjacently and distantly located effector-encoding genes, thus forming small groups of co-regulated effectors.


Assuntos
Legionella pneumophila , Legionella , Legionella/genética , Legionella/metabolismo , Proteínas de Bactérias/metabolismo , Legionella pneumophila/metabolismo , Fator sigma/genética , Fator sigma/metabolismo , Sequências Reguladoras de Ácido Nucleico
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA
...