Abstract
Pseudomonas aeruginosa can cause severe infections in humans. This bacterium often adopts a biofilm lifestyle that is hard to treat. In several previous studies, the PprA-PprB two-component system (TCS), which controls the expression of type IVb pili, BapA adhesin, and CupE fimbriae, was shown to be involved in biofilm formation (M. Romero, H. Silistre, L. Lovelock, V. J. Wright, K.-G. Chan, et al., Nucleic Acids Res 46:6823-6840, 2018, https://doi.org/10.1093/nar/gky324; S. de Bentzmann, C. Giraud, C. S. Bernard, V. Calderon, F. Ewald F, et al., PLoS Pathog 8:e1003052, 2012, https://doi.org/10.1371/journal.ppat.1003052). However, signals or environmental conditions that can trigger the PprA-PprB TCS are still unknown, and the molecular mechanisms of PprB-mediated biofilm formation are poorly characterized. Here, we report that carbon starvation stress (CSS) can induce the expression of pprB and genes in the PprB regulon. CSS-induced pprB transcription is mediated by the stress response sigma factor RpoS rather than the two-component sensor PprA. We also observed a strong negative regulation of PprB on the transcription of itself. Further experiments showed that PprB overexpression greatly enhanced cell-cell adhesion (CCA) and cell-surface adhesion (CSA) in P. aeruginosa Specifically, under the background of PprB overexpression, both the BapA adhesin and CupE fimbriae displayed positive effects on CCA and CSA, while the type IVb pili showed an unexpected negative effect on CCA and no effect on CSA. In addition, expression of the PprB regulon genes were significantly increased in 3-day colony biofilms, indicating a possible carbon limitation state. The CSS-RpoS-PprB-Bap/Flp/CupE pathway identified in this study provides a new perspective on the process of biofilm formation in carbon-limited environments.IMPORTANCE Typically, the determination of the external signals that can trigger a regulatory system is crucial to understand the regulatory logic and inward function of that system. The PprA-PprB two-component system was reported to be involved in biofilm formation in Pseudomonas aeruginosa, but the signals triggering this system are unknown. In this study, we found that carbon starvation stress (CSS) induces transcription of pprB and genes in the PprB regulon through an RpoS-dependent pathway. Increased PprB expression leads to enhanced cell-cell adhesion (CCA) and cell-surface adhesion (CSA) in P. aeruginosa Both CCA and CSA are largely dependent on the Bap secretion system and are moderately dependent on the CupE fimbriae. Our findings suggest that PprB reinforces the structure of biofilms under carbon-limited conditions, and the Bap secretion system and CupE fimbriae are two potential targets for biofilm treatment.
Highlights
IMPORTANCE Typically, the determination of the external signals that can trigger a regulatory system is crucial to understand the regulatory logic and inward function of that system
Despite the fact that bacteria cannot increase their biomass without carbon supplementation, we found that cells still divided into smaller daughter cells under carbon starvation stress (CSS), which may contribute to the increased CFU of the pprB mutant after 6-h carbon deprivation
With regard to the physiological role of PprB in bacteria, previous studies have mainly focused on the phenotypes of the pprB overexpression strains, which, compared with the wild-type strain, have shown increased cell membrane permeability and aminoglycoside sensitivity, decreased cellular cytotoxicity and virulence in flies, and better biofilm formation [22, 23]
Summary
IMPORTANCE Typically, the determination of the external signals that can trigger a regulatory system is crucial to understand the regulatory logic and inward function of that system. In one previous study, a hyperbiofilm phenotype that was independent of Pel and Psl was identified [22] Cells in this biofilm exhibited overexpression of PprB, decreased type III secretion, and increased drug susceptibility [22]. The pprB mutant strain was recently shown to form a significantly reduced biofilm in microfluidics systems [24] These results suggest that PprB and its downstream regulated proteins can dominate the formation of biofilm via a Pel- and Psl-independent pathway. The PprB regulon contains multiple open reading frames, including genes encoding type I secretion system components (bapA-bapD), CupE CU fimbriae, and type IVb pili, all of which are positively and directly regulated by PprB at the transcriptional level [22]. We further demonstrate the roles of type IVb pili, CupE fimbriae, and BapA adhesin in cell-cell adhesion (CCA) and cell-surface adhesion (CSA) by P. aeruginosa. The CSS-RpoS-PprB-BapA/Flp/CupE signaling pathway determined in this study provides a new perspective on the process of biofilm formation and may be helpful in directing biofilm treatment
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