The two‐component system <scp>TarR–TarS</scp> is regulated by <scp>c‐di‐GMP</scp>/<scp>FleQ</scp> and <scp>FliA</scp> and modulates antibiotic susceptibility in <i>Pseudomonas putida</i>

Xiao, Yujie; Nie, Liang; Chen, Haozhe; He, Meina; Liang, Qingyuan; Nie, Hailing; Chen, Wenli; Huang, Qiaoyun

doi:10.1111/1462-2920.15555

Cited by 6 publications

(2 citation statements)

References 62 publications

(84 reference statements)

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“…To identify the interacting partners of VfmH, an in vitro pull-down assay was performed, modified from the method described previously ( 54 ). In brief, histidine-tagged VfmH was overexpressed in BL21(DE3) cells for 4 h, and the cells were collected by centrifugation.…”

Section: Methodsmentioning

confidence: 99%

Dissecting the molecular dance: c-di-GMP, cAMP-CRP, and VfmH collaboration in pectate lyase regulation for Dickeya dadantii —unveiling the soft rot pathogen’s strategy

Banerjee,

Yuan,

Yang

2023

Microbiol Spectr

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Dickeya dadantii is a Gram-negative bacterium that causes soft rot diseases in a variety of plants. This pathogen secretes pectate lyase (Pel) enzymes to degrade the plant cell wall, a process controlled by the bacterial second messenger cyclic diguanylate monophosphate (c-di-GMP). VfmH is a Fis-family response regulator of the Vfm quorum sensing system, which has been previously reported to positively regulate pel transcription. Our results demonstrated that VfmH upregulated the production of Pel at low c-di-GMP levels, which was suppressed by c-di-GMP via deleting gene egcpB , encoding a phosphodiesterase that degrades c-di-GMP. Multiple sequence alignments between FleQ, a known c-di-GMP binding protein, and VfmH revealed several potential c-di-GMP binding motifs. We further validated these motifs via single amino acid substitutions. Indeed, derivatives of VfmH, including VfmH R195A , VfmH R279A , and VfmH R344A , abolished the c-di-GMP binding ability and the corresponding regulation in Pel. We found that VfmH exhibited ATPase activity, and this could be hindered by c-di-GMP binding. Interestingly, our results showed that VfmH interacted with CRP, suggesting that the cyclic AMP (cAMP) signaling pathway is also involved in the VfmH-mediated Pel regulation. Taken together, our study suggested that D. dadantii amalgamates both c-di-GMP and cAMP signaling networks along with the Vfm quorum sensing pathway to control Pel. IMPORTANCE Bacteria respond to environmental changes and adapt to host systems. The response regulator VfmH of the Vfm quorum sensing system regulates a crucial virulence factor, pectate lyase (Pel), in Dickeya dadantii . At high c-di-GMP concentrations, VfmH binds c-di-GMP, resulting in the loss of its activation property in the Pel and virulence regulation in D. dadantii . VfmH binds to c-di-GMP via three conserved arginine residues, and mutations of these residues eliminate the c-di-GMP-related phenotypes of VfmH in Pel synthesis. Our data also show that VfmH interacts with CRP to regulate pelD transcription, thus integrating cyclic AMP and c-di-GMP signaling pathways to control virulence in D. dadantii . We propose that VfmH is an important intermediate factor incorporating quorum sensing and nucleotide signaling pathways for the collective regulation of D. dadantii pathogenesis.

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Section: Methodsmentioning

confidence: 99%

Dissecting the molecular dance: c-di-GMP, cAMP-CRP, and VfmH collaboration in pectate lyase regulation for Dickeya dadantii —unveiling the soft rot pathogen’s strategy

Banerjee,

Yuan,

Yang

2023

Microbiol Spectr

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“…Upon activation, FliA also has a complex but demonstrated role in biofilm formation, as our lab [35] and others [36][37][38] have explored. In Pseudomonas putida, downstream targets of FliA, like the chemotaxis TarR-TarS two-component system and the second messenger cyclic-di-GMP, have been implicated as directly regulating antibiotic susceptibility to several antibiotics [39]. Most recently, FliA was identified among seven other protein "hubs" in the antibiotic resistance mechanism network of the gram-negative Stenotrophomonas maltophilia, which has an unusually high resistance for many antibiotics and heavy metals [40].…”

Section: Introductionmentioning

confidence: 99%

Constitutive Activation of RpoH and the Addition of L-arabinose Influence Antibiotic Sensitivity of PHL628 E. coli

Frizzell,

Taylor,

Ryno

2024

Antibiotics

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Antibiotics are used to combat the ever-present threat of infectious diseases, but bacteria are continually evolving an assortment of defenses that enable their survival against even the most potent treatments. While the demand for novel antibiotic agents is high, the discovery of a new agent is exceedingly rare. We chose to focus on understanding how different signal transduction pathways in the gram-negative bacterium Escherichia coli (E. coli) influence the sensitivity of the organism to antibiotics from three different classes: tetracycline, chloramphenicol, and levofloxacin. Using the PHL628 strain of E. coli, we exogenously overexpressed two transcription factors, FliA and RpoH.I54N (a constitutively active mutant), to determine their influence on the minimum inhibitory concentration (MIC) and minimum duration of killing (MDK) concentration for each of the studied antibiotics. We hypothesized that activating these pathways, which upregulate genes that respond to specific stressors, could mitigate bacterial response to antibiotic treatment. We also compared the exogenous overexpression of the constitutively active RpoH mutant to thermal heat shock that has feedback loops maintained. While FliA overexpression had no impact on MIC or antibiotic tolerance, RpoH.I54N overexpression reduced the MIC for tetracycline and chloramphenicol but had no independent impact on antibiotic tolerance. Thermal heat shock alone also did not affect MIC or antibiotic tolerance. L-arabinose, the small molecule used to induce expression in our system, unexpectedly independently increased the MICs for tetracycline (>2-fold) and levofloxacin (3-fold). Additionally, the combination of thermal heat shock and arabinose provided a synergistic, 5-fold increase in MIC for chloramphenicol. Arabinose increased the tolerance, as assessed by MDK99, for chloramphenicol (2-fold) and levofloxacin (4-fold). These experiments highlight the potential of the RpoH pathway to modulate antibiotic sensitivity and the emerging implication of arabinose in enhanced MIC and antibiotic tolerance.

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Cytoplasmic molecular chaperones in Pseudomonas species

et al. 2022

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The two‐component system TarR–TarS is regulated by c‐di‐GMP/FleQ and FliA and modulates antibiotic susceptibility in Pseudomonas putida

Cited by 6 publications

References 62 publications

Dissecting the molecular dance: c-di-GMP, cAMP-CRP, and VfmH collaboration in pectate lyase regulation for Dickeya dadantii —unveiling the soft rot pathogen’s strategy

Dissecting the molecular dance: c-di-GMP, cAMP-CRP, and VfmH collaboration in pectate lyase regulation for Dickeya dadantii —unveiling the soft rot pathogen’s strategy

Constitutive Activation of RpoH and the Addition of L-arabinose Influence Antibiotic Sensitivity of PHL628 E. coli

Cytoplasmic molecular chaperones in Pseudomonas species

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