Yersinia pseudotuberculosis BarA-UvrY Two-Component Regulatory System Represses Biofilms via CsrB

Abstract

The formation of biofilms by Yersinia pseudotuberculosis (Yptb) and Y. pestis requires the hmsHFRS genes, which direct production of a polysaccharide extracellular matrix (Hms-ECM). Despite possessing identical hmsHFRS sequences, Yptb produces much less Hms-ECM than Y. pestis. The regulatory influences that control Yptb Hms-ECM production and biofilm formation are not fully understood. In this study, negative regulators of biofilm production in Yptb were identified. Inactivation of the BarA/UvrY two-component system or the CsrB regulatory RNA increased binding of Congo Red dye, which correlates with extracellular polysaccharide production. These mutants also produced biofilms that were substantially more cohesive than the wild type strain. Disruption of uvrY was not sufficient for Yptb to cause proventricular blockage during infection of Xenopsylla cheopis fleas. However, this strain was less acutely toxic toward fleas than wild type Yptb. Flow cytometry measurements of lectin binding indicated that Yptb BarA/UvrY/CsrB mutants may produce higher levels of other carbohydrates in addition to poly-GlcNAc Hms-ECM. In an effort to characterize the relevant downstream targets of the BarA/UvrY system, we conducted a proteomic analysis to identify proteins with lower abundance in the csrB::Tn5 mutant strain. Urease subunit proteins were less abundant and urease enzymatic activity was lower, which likely reduced toxicity toward fleas. Loss of CsrB impacted expression of several potential regulatory proteins that may influence biofilms, including the RcsB regulator. Overexpression of CsrB did not alter the Congo-red binding phenotype of an rcsB::Tn5 mutant, suggesting that the effect of CsrB on biofilms may require RcsB. These results underscore the regulatory and compositional differences between Yptb and Y. pestis biofilms. By activating CsrB expression, the Yptb BarA/UvrY two-component system has pleiotropic effects that impact biofilm production and stability.

The phosphorelay BarA/SirA activates the non-cognate regulator RcsB in Salmonella enterica

“…By contrast, RcsB amounts were the same for both wild-type Salmonella and the csrB csrC double mutant in LB broth without NaCl (S7A Fig), which is in contrast to previous findings reported with Yersinia pseudotuberculosis [33], and this was also the case for RcsD amounts (S7B Fig). Cumulatively, the results in this section strongly suggest that when wild-type Salmonella experiences low osmolarity, BarA activates RcsB-dependent transcription by reducing RcsC abundance through the action of the SirA-dependent CsrB and CsrC regulatory RNAs.…”

“…CsrB and CsrC act by sequestering CsrA, an RNA-binding protein that binds to the 5' untranslated region (UTR) of hundreds of transcripts, typically decreasing their translation [29][30][31][32]. That BarA activation of RcsB may be mediated by CsrB and CsrC in Salmonella is also supported by the phenotype of a csrB mutant Yersinia pseudotuberculosis grown on solid media, which produced 8-fold less RcsB protein than the wild-type strain [33].…”

Regulation of colony morphology and biofilm formation in Shewanella algae

“…Surprisingly, while the phosphorelay activity of BarA was not required for S. algae biofilm formation on the abiotic surface, phenotypic differences in biofilm formation patterns were recorded upon the expression of mutants in two or all three amino acids in the phosphorylation cascade. Notably, control of biofilm cohesiveness by BarA has been reported in Y. pseudotuberculosis (Schachterle et al, 2018). A limitation of our study is the strong plasmid effect, which altered dramatically the biofilm phenotypes under investigation, as well as the intrinsic difficulty of performing gene manipulation in this strain.…”

“…Interruption of the hybrid sensor histidine kinase barA caused significant inhibition of biofilm formation. A study in Yersinia pseudotuberculosis showed a barA mutant to increase CR binding while surface-associated biofilm in multiwell plates was not significantly different from that of the WT (Schachterle et al, 2018). Surprisingly, while the phosphorelay activity of BarA was not required for S. algae biofilm formation on the abiotic surface, phenotypic differences in biofilm formation patterns were recorded upon the expression of mutants in two or all three amino acids in the phosphorylation cascade.…”

Viscoelastic response ofEscherichia colibiofilms to genetically altered expression of extracellular matrix components

“…We also tested colony morphology and binding of the diazo dye, Congo red, to visualize differences in curli and PGA production by these strains. [58][59] Colonies of wt, pga and crl strains were red in color (Panel D) indicative of binding of Congo red to both PGA and crl, while the cla and epsstrains did not bind the dye. To compare the viscosity of bacterial suspensions and biofilms we measured viscosity curves (Fig.…”