DksA is a conserved master regulator of stress response in Acinetobacter baumannii.
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ABSTRACT:
SUBMITTER: Maharjan RP
PROVIDER: S-EPMC10325922 | biostudies-literature | 2023 Jul
REPOSITORIES: biostudies-literature
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DksA is a conserved master regulator of stress response in Acinetobacter baumannii.
Maharjan Ram P RP Sullivan Geraldine J GJ Adams Felise G FG Shah Bhumika S BS Hawkey Jane J Delgado Natasha N Semenec Lucie L Dinh Hue H Li Liping L Short Francesca L FL Parkhill Julian J Paulsen Ian T IT Barquist Lars L Eijkelkamp Bart A BA Cain Amy K AK
Nucleic acids research 20230701 12
Coordination of bacterial stress response mechanisms is critical for long-term survival in harsh environments for successful host infection. The general and specific stress responses of well-studied Gram-negative pathogens like Escherichia coli are controlled by alternative sigma factors, archetypically RpoS. The deadly hospital pathogen Acinetobacter baumannii is notoriously resistant to environmental stresses, yet it lacks RpoS, and the molecular mechanisms driving this incredible stress toler ...[more]
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Project description:DksA with (p)ppGpp regulates a wide range of gene transcriptions during the stringent response. The aim of this study was to identify a DksA ortholog in Acinetobacter baumannii and clarify the roles of DksA in bacterial physiology and virulence. The ∆dksA mutant and its complemented strains were constructed using A. baumannii ATCC 17978. The AlS_0248 in A. baumannii ATCC 17978 was identified to dksA using sequence homology, protein structure prediction, and gene expression patterns under different culture conditions. The ∆dksA mutant strain showed a filamentous morphology compared with the wild-type (WT) strain. Bacterial growth was decreased in the ∆dksA mutant strain under static conditions. Surface motility was decreased in the ∆dksA mutant strain compared with the WT strain. In contrast, biofilm formation was increased and biofilm-associated genes, such as bfmR/S and csuC/D/E, were upregulated in the ∆dksA mutant strain. The ∆dksA mutant strain produced less autoinducers than the WT strain. The expression of abaI and abaR was significantly decreased in the ∆dksA mutant strain. Furthermore, the ∆dksA mutant strain showed less bacterial burden and milder histopathological changes in the lungs of mice than the WT strain. Mice survival was also significantly different between the ∆dksA mutant and WT strains. Conclusively, DksA is directly or indirectly involved in regulating a wide range of genes associated with bacterial physiology and virulence, which contributes to the pathogenesis of A. baumannii. Thus, DksA is a potential anti-virulence target for A. baumannii infection.
| S-EPMC8583241 | biostudies-literature
Project description:The stringent response regulators, (p)ppGpp and DksA, modulate various genes involved in physiological processes, virulence, and antimicrobial resistance in pathogenic bacteria. This study investigated the role of DksA in the antimicrobial susceptibility of Acinetobacter baumannii. The ∆dksA mutant (KM0248D) of A. baumannii ATCC 17978 and its complemented strain (KM0248C) were used, in addition to the ∆dksA mutant strain (NY0298D) of clinical 1656-2 strain. The microdilution assay was used to determine the minimum inhibitory concentrations (MICs) of antimicrobial agents. Quantitative real-time PCR was performed to analyze the expression of genes associated with efflux pumps. The KM0248D strain exhibited an increase of MICs to quinolones and tetracyclines, whereas KM0248D and NY0298D strains exhibited a decrease of MICs to aminoglycosides. The expression of genes associated with efflux pumps, including adeB, adeI/J, abeM, and/or tetA, was upregulated in both ∆dksA mutant strains. The deletion of dksA altered bacterial morphology in the clinical 1656-2 strain. In conclusion, DksA modulates the antimicrobial susceptibility of A. baumannii. The ∆dksA mutant strains of A. baumannii upregulate efflux pump gene expression, whereas (p)ppGpp-deficient mutants downregulate efflux pump gene expression. (p)ppGpp and DksA conduct opposite roles in the antimicrobial susceptibility of A. baumannii via efflux pump gene regulation.
| S-EPMC8698368 | biostudies-literature
Project description:Ability to redirect limiting cellular resources accurately is key to bacteria for surviving in harsh environments that they often encounter during their lifetime. DksA, a transcriptional initiating factor, plays critical roles in regulating stress responses and antibiotic tolerance. Acinetobacter baumannii has become a major healthcare threat and responsible for both nosocomial and community acquired deadly infections worldwide. Yet, little is known about the role of DksA in A. baumannii. Here we describe a highly pleiotropic nature of DksA that it helps redirect the key metabolic pathways associated with the respiration, energy production, protein biosynthesis. Inhibition of ribosomal RNAs, ATP production by DksA is detrimental to bacteria under bacteriostatic stresses such as copper and trimethoprim. By contrast, it is required for survival in stresses that generate reactive-oxygen species such as zinc and gentamycin. Bacteria lacking DksA exhibit increase sensitivity to human serum, promote biofilm formation and capsule production. Not only does DksA show diverse phenotypes in vitro, but it also impacted on virulence in a niche specific manner during in vivo Galleria mellonella and murine infections. Our data collectively provides the detailed insight into the role of DksA in stress protection and virulence in A. baumannii and layout a roadmap for similar findings in other clinically important pathogens.
2022-09-13 | GSE169081 | GEO
Project description:Acinetobacter baumannii is an emerging opportunistic pathogen that primarily infects critically ill patients in nosocomial settings. Because of its rapid acquisition of antibiotic resistance, infections caused by A. baumannii have become extremely difficult to treat, underlying the importance of identifying new antimicrobial targets for this pathogen. Manganese (Mn) is an essential nutrient metal required for a number of bacterial processes, including the response to oxidative stress. Here, we show that exogenous Mn can restore A. baumannii viability in the presence of reactive oxygen species (ROS). This restoration is not dependent on the high-affinity Nramp family Mn transporter, MumT, as a ΔmumT mutant is no more sensitive to hydrogen peroxide (H2O2) killing than wild-type A. baumannii However, mumR, which encodes the transcriptional regulator of mumT, is critical for growth and survival in the presence of H2O2, suggesting that MumR regulates additional genes that contribute to H2O2 resistance. RNA sequencing revealed a role for mumR in regulating the activity of a number of metabolic pathways, including two pathways, phenylacetate and gamma-aminobutyric acid catabolism, which were found to be important for resisting killing by H2O2 Finally, ΔmumR exhibited reduced fitness in a murine model of pneumonia, indicating that MumR-regulated gene products are crucial for protection against the host immune response. In summary, these results suggest that MumR facilitates resistance to the host immune response by activating a transcriptional program that is critical for surviving both Mn starvation and oxidative stress.
| S-EPMC7035938 | biostudies-literature
Project description:DksA controls transcription of genes associated with diverse stress responses, such as amino acid and carbon starvation, oxidative stress, and iron starvation. DksA binds within the secondary channel of RNA polymerase, extending its long coiled-coil domain towards the active site. The cellular expression of DksA remains constant due to a negative feedback autoregulation, raising the question of whether DksA activity is directly modulated during stress. Here, we show that Escherichia coli DksA is essential for survival in acidic conditions and that, while its cellular levels do not change significantly, DksA activity and binding to RNA polymerase are increased at lower pH, with a concomitant decrease in its stability. NMR data reveal pH-dependent structural changes centered at the interface of the N and C-terminal regions of DksA. Consistently, we show that a partial deletion of the N-terminal region and substitutions of a histidine 39 residue at the domain interface abolish pH sensitivity in vitro. Together, these data suggest that DksA responds to changes in pH by shifting between alternate conformations, in which competing interactions between the N- and C-terminal regions modify the protein activity.
| S-EPMC4370453 | biostudies-literature
Project description:Identification and validation is the first phase of target-based antimicrobial development. BfmR (RstA), a response regulator in a two-component signal transduction system (TCS) in Acinetobacter baumannii, is an intriguing potential antimicrobial target. A unique characteristic of BfmR is that its inhibition would have the dual benefit of significantly decreasing in vivo survival and increasing sensitivity to selected antimicrobials. Studies on the clinically relevant strain AB307-0294 have shown BfmR to be essential in vivo. Here, we demonstrate that this phenotype in strains AB307-0294 and AB908 is mediated, in part, by enabling growth in human ascites fluid and serum. Further, BfmR conferred resistance to complement-mediated bactericidal activity that was independent of capsular polysaccharide. Importantly, BfmR also increased resistance to the clinically important antimicrobials meropenem and colistin. BfmR was highly conserved among A. baumannii strains. The crystal structure of the receiver domain of BfmR was determined, lending insight into putative ligand binding sites. This enabled an in silico ligand binding analysis and a blind docking strategy to assess use as a potential druggable target. Predicted binding hot spots exist at the homodimer interface and the phosphorylation site. These data support pursuing the next step in the development process, which includes determining the degree of inhibition needed to impact growth/survival and the development a BfmR activity assay amenable to high-throughput screening for the identification of inhibitors. Such agents would represent a new class of antimicrobials active against A. baumannii which could be active against other Gram-negative bacilli that possess a TCS with shared homology. IMPORTANCE Increasing antibiotic resistance in bacteria, particularly Gram-negative bacilli, has significantly affected the ability of physicians to treat infections, with resultant increased morbidity, mortality, and health care costs. In fact, some strains of bacteria are resistant to all available antibiotics, such as Acinetobacter baumannii, which is the focus of this report. Therefore, the development of new antibiotics active against these resistant strains is urgently needed. In this study, BfmR is further validated as an intriguing target for a novel class of antibiotics. Successful inactivation of BfmR would confer the multiple benefits of a decreased ability of A. baumannii to survive in human body fluids, increased sensitivity to complement-mediated bactericidal activity and, importantly, increased sensitivity to other antibiotics. Structural studies support the potential for this "druggable" target, as they identify the potential for small-molecule binding at functionally relevant sites. Next-phase high-throughput screening studies utilizing BfmR are warranted.
| S-EPMC4888885 | biostudies-literature
Project description:A critical component of bacterial pathogenesis is the ability of an invading organism to sense and adapt to the harsh environment imposed by the host's immune system. This is especially important for opportunistic pathogens, such as Acinetobacter baumannii, a nutritionally versatile environmental organism that has recently gained attention as a life-threatening human pathogen. The emergence of A. baumannii is closely linked to antibiotic resistance, and many contemporary isolates are multidrug resistant (MDR). Unlike many other MDR pathogens, the molecular mechanisms underlying A. baumannii pathogenesis remain largely unknown. We report here the characterization of two recently identified virulence determinants, GigA and GigB, which comprise a signal transduction pathway required for surviving environmental stresses, causing infection and antibiotic resistance. Through transcriptome analysis, we show that GigA and GigB coordinately regulate the expression of many genes and are required for generating an appropriate transcriptional response during antibiotic exposure. Genetic and biochemical data demonstrate a direct link between GigA and GigB and the nitrogen phosphotransferase system (PTSNtr), establishing a novel connection between a novel stress response module and a well-conserved metabolic-sensing pathway. Based on the results presented here, we propose that GigA and GigB are master regulators of a global stress response in A. baumannii, and coupling this pathway with the PTSNtr allows A. baumannii to integrate cellular metabolic status with external environmental cues.IMPORTANCE Opportunistic pathogens, including Acinetobacter baumannii, encounter many harsh environments during the infection cycle, including antibiotic exposure and the hostile environment within a host. While the development of antibiotic resistance in A. baumannii has been well studied, how this organism senses and responds to environmental cues remain largely unknown. Herein, we investigate two previously identified virulence determinants, GigA and GigB, and report that they are required for in vitro stress resistance, likely comprising upstream elements of a global stress response pathway. Additional experiments identify a connection between GigA/GigB and a widely conserved metabolic-sensing pathway, the nitrogen phosphotransferase system. We propose that coordination of these two pathways allows A. baumannii to respond appropriately to changing environmental conditions, including those encountered during infection.
| S-EPMC5405210 | biostudies-literature
Project description:Abstract Indole is an important signal employed by many bacteria to modulate intraspecies signaling and interspecies or interkingdom communication. Our recent study revealed that indole plays a key role in regulating the physiology and virulence of Acinetobacter baumannii. However, it is not clear how A. baumannii perceives and responds to the indole signal in modulating biological functions. Here, we report that indole controls the physiology and virulence of A. baumannii through a previously uncharacterized response regulator designated as AbiR (A1S_1394), which contains a cheY-homologous receiver (REC) domain and a helix-turn-helix (HTH) DNA-binding domain. AbiR controls the same biological functions as the indole signal, and indole-deficient mutant phenotypes were rescued by in trans expression of AbiR. Intriguingly, unlike other response regulators that commonly interact with signal ligands through the REC domain, AbiR binds to indole with a high affinity via an unusual binding region, which is located between its REC and HTH domains. This interaction substantially enhances the activity of AbiR in promoter binding and in modulation of target gene expression. Taken together, our results present a widely conserved regulator that controls bacterial physiology and virulence by sensing the indole signal in a unique mechanism.
| S-EPMC10465187 | biostudies-literature
Project description:Acinetobacter baumannii is a leading cause of ventilator-associated pneumonia in intensive care units, and the increasing rates of antibiotic resistance make treating these infections challenging. Consequently, there is an urgent need to develop new antimicrobials to treat A. baumannii infections. One potential therapeutic option is to target bacterial systems involved in maintaining appropriate metal homeostasis, processes that are critical for the growth of pathogens within the host. The A. baumannii inner membrane zinc transporter ZnuABC is required for growth under low-zinc conditions and for A. baumannii pathogenesis. The expression of znuABC is regulated by the transcriptional repressor Zur. To investigate the role of Zur during the A. baumannii response to zinc limitation, a zur deletion mutant was generated, and transcriptional changes were analyzed using RNA sequencing. A number of Zur-regulated genes were identified that exhibit increased expression both when zur is absent and under low-zinc conditions, and Zur binds to predicted Zur box sequences of several genes affected by zinc levels or the zur mutation. Furthermore, the zur mutant is impaired for growth in the presence of both high and low zinc levels compared to wild-type A. baumannii. Finally, the zur mutant exhibits a defect in dissemination in a mouse model of A. baumannii pneumonia, establishing zinc sensing as a critical process during A. baumannii infection. These results define Zur-regulated genes within A. baumannii and demonstrate a requirement for Zur in the A. baumannii response to the various zinc levels experienced within the vertebrate host.
| S-EPMC4097591 | biostudies-literature
Project description:For the opportunistic pathogen Acinetobacter baumannii, desiccation tolerance is thought to contribute significantly to the persistence of these bacteria in the healthcare environment. We investigated the ability of A. baumannii to survive rapid drying, and found that some strains exhibited a profoundly desiccation-resistant phenotype, characterized by the ability of a large proportion of cells to survive on a dry surface for an extended period of time. However, this phenotype was only displayed during the stationary phase of growth. Most interestingly, we found that drying resistance could be lost after extended cultivation in liquid medium. Genome sequencing of isolates that became drying-sensitive identified mutations in bfmR, which encodes a two-component response regulator that is important for A. baumannii virulence. Additionally, BfmR was necessary for the expression of stress-related proteins during stationary phase, and one of these, KatE, was important for long-term drying survival. These results suggested that BfmR may control stress responses, and we demonstrated that the ΔbfmR mutant was more sensitive to hydrogen peroxide, nutrient starvation, and increased osmolarity. We also found that cross-protection against drying could be stimulated by either starvation, which required BfmR, or increased osmolarity. These results imply that BfmR plays a role in controlling stress responses in A. baumannii which help protect cells during desiccation, and they provide a regulatory link between this organism's ability to persist in the environment and pathogenicity.
| S-EPMC6181384 | biostudies-literature