Two-component signal transduction systems (TCS) are often used by bacteria to rapidly assess and respond to environmental changes. ChvG/ChvI is a TCS conserved in α-proteobacteria and known for regulating expression of genes related to exopolysaccharide production, virulence and growth. The sensor kinase ChvG autophosphorylates upon yet unknown signals and phosphorylates the response regulator ChvI to activate transcription. Recent studies in Caulobacter crescentus showed that chv mutants are sensitive to vancomycin treatment and fail to grow in synthetic minimal media. In this work, we identified the osmotic imbalance as the main cause of growth impairment in synthetic minimal media. We also determined the ChvI regulon and confirmed that ChvI regulates cell envelope architecture at different levels by controlling outer membrane, peptidoglycan assembly/recycling and inner membrane proteins. Furthermore, we identified genes with osmoregulatory properties and confirmed that osmotic upshift is a signal triggering ChvG-dependent phosphorylation of ChvI. In addition, we challenged chv mutants with other cell envelope related stress and found that targeting with antibiotics the transpeptidation of peptidoglycan during cell elongation impairs growth of the mutant. Moreover, these antibiotics activate expression of the chvIG-hprK operon in ChvI-dependent and independent ways. Finally, we observed that the sensor kinase ChvG fused to a fluorescent protein relocates from a patchy-spotty distribution to distinctive foci after transition from complex to synthetic minimal media. Interestingly, this pattern of (re)location has been described for proteins involved in cell growth control and peptidoglycan synthesis upon osmotic shock. Overall, our data support that the ChvGI TCS is mainly used to maintain cell envelope homeostasis by monitoring osmotic imbalances and damages in the peptidoglycan layer.

Two-component systems (TCS) are often used by bacteria to rapidly assess and respond to environmental changes. The ChvG/ChvI (ChvGI) TCS conserved in α-proteobacteria is known for regulating expression of genes related to exopolysaccharide production, virulence and growth. The sensor kinase ChvG autophosphorylates upon yet unknown signals and phosphorylates the response regulator ChvI to regulate transcription. Recent studies in Caulobacter crescentus showed that chv mutants are sensitive to vancomycin treatment and fail to grow in synthetic minimal media. In this work, we identified the osmotic imbalance as the main cause of growth impairment in synthetic minimal media. We also determined the ChvI regulon and found that ChvI regulates cell envelope architecture by controlling outer membrane, peptidoglycan assembly/recycling and inner membrane proteins. In addition, we found that ChvI phosphorylation is also activated upon antibiotic treatment with vancomycin. We also challenged chv mutants with other cell envelope related stress and found that treatment with antibiotics targeting transpeptidation of peptidoglycan during cell elongation impairs growth of the mutant. Finally, we observed that the sensor kinase ChvG relocates from a patchy-spotty distribution to distinctive foci after transition from complex to synthetic minimal media. Interestingly, this pattern of (re)location has been described for proteins involved in cell growth control and peptidoglycan synthesis upon osmotic shock. Overall, our data support that the ChvGI TCS is mainly used to monitor and respond to osmotic imbalances and damages in the peptidoglycan layer to maintain cell envelope homeostasis.

Summary Serratia sp. strain ATCC 39006 (S39006) can float in aqueous environments due to natural production of gas vesicles (GVs). Expression of genes for GV morphogenesis is stimulated in low oxygen conditions, thereby enabling migration to the air–liquid interface. Quorum sensing (via SmaI and SmaR) and transcriptional and post‐transcriptional regulators, including RbsR and RsmA, respectively, connect the control of cell buoyancy, motility and secondary metabolism. Here, we define a new pleiotropic regulator found in screens of GV mutants. A mutation in the gene trkH , encoding a potassium transporter, caused upregulation of GV formation, flotation, and the prodigiosin antibiotic, and downregulation of flagellar motility. Pressure nephelometry revealed that the mutation in trkH affected cell turgor pressure. Our results show that osmotic change is an important physiological parameter modulating cell buoyancy and antimicrobial production in S39006, in response to environmental potassium levels.

Summary Serratia sp. ATCC 39006 produces intracellular gas vesicles to enable upward flotation in water columns. It also uses flagellar rotation to swim through liquid and swarm across semi‐solid surfaces. Flotation and motility can be co‐regulated with production of a β‐lactam antibiotic (carbapenem carboxylate) and a linear tripyrrole red antibiotic, prodigiosin. Production of gas vesicles, carbapenem and prodigiosin antibiotics, and motility are controlled by master transcriptional and post‐transcriptional regulators, including the SmaI/SmaR‐based quorum sensing system and the mRNA binding protein, RsmA. Recently, the ribose operon repressor, RbsR, was also defined as a pleiotropic regulator of flotation and virulence factor elaboration in this strain. Here, we report the discovery of a new global regulator (FloR; a DeoR family transcription factor) that modulates flotation through control of gas vesicle morphogenesis. The floR mutation is highly pleiotropic, down‐regulating production of gas vesicles, carbapenem and prodigiosin antibiotics, and infection in Caenorhabditis elegans, but up‐regulating flagellar motility. Detailed proteomic analysis using TMT peptide labelling and LC–MS/MS revealed that FloR is a physiological master regulator that operates through subordinate pleiotropic regulators including Rap, RpoS, RsmA, PigU, PstS and PigT.