The LacI–Family Transcription Factor, RbsR, Is a Pleiotropic Regulator of Motility, Virulence, Siderophore and Antibiotic Production, Gas Vesicle Morphogenesis and Flotation in Serratia

Abstract

Gas vesicles (GVs) are proteinaceous, gas-filled organelles used by some bacteria to enable upward movement into favorable air/liquid interfaces in aquatic environments. Serratia sp. ATCC39006 (S39006) was the first enterobacterium discovered to produce GVs naturally. The regulation of GV assembly in this host is complex and part of a wider regulatory network affecting various phenotypes, including antibiotic biosynthesis. To identify new regulators of GVs, a comprehensive mutant library containing 71,000 insertion mutants was generated by random transposon mutagenesis and 311 putative GV-defective mutants identified. Three of these mutants were found to have a transposon inserted in a LacI family transcription regulator gene (rbsR) of the putative ribose operon. Each of these rbsR mutants was GV-defective; no GVs were visible by phase contrast microscopy (PCM) or transmission electron microscopy (TEM). GV deficiency was caused by the reduction of gvpA1 and gvrA transcription (the first genes of the two contiguous operons in the GV gene locus). Our results also showed that a mutation in rbsR was highly pleiotropic; the production of two secondary metabolites (carbapenem and prodigiosin antibiotics) was abolished. Interestingly, the intrinsic resistance to the carbapenem antibiotic was not affected by the rbsR mutation. In addition, the production of a siderophore, cellulase and plant virulence was reduced in the mutant, whereas it exhibited increased swimming and swarming motility. The RbsR protein was predicted to bind to regions upstream of at least 18 genes in S39006 including rbsD (the first gene of the ribose operon) and gvrA. Electrophoretic mobility shift assays (EMSA) confirmed that RbsR bound to DNA sequences upstream of rbsD, but not gvrA. The results of this study indicate that RbsR is a global regulator that affects the modulation of GV biogenesis, but also with complex pleiotropic physiological impacts in S39006.

Environmental potassium regulates bacterial flotation, antibiotic production and turgor pressure in Serratia through the TrkH transporter

“…Previous studies on S39006 GV production screened for translucent mutants in the prodigiosin negative strain NWA19 ( Δ pigC ) (Ramsay et al, ; Lee et al, ). We focused this screen on transconjugants that appeared more opaque than NWA19 to try to identify novel negative regulators of GV production.…”

“…Cell buoyancy control is physiologically connected with motility and antimicrobial production through global pleiotropic regulators in S39006 (Ramsay et al, ; Lee et al, ). We noticed that AQY107A showed moderately impaired flagellar motility when compared with WT, while ectopic expression of wild‐type trkH restored swimming motility in the mutant (Supporting Information Fig.…”

“…S6 and S7). Interestingly, high phosphate concentrations and different carbon sources such as ribose and gluconate also have negative impacts on prodigiosin production in S39006 (Fineran et al, 2005a; Lee et al, ). Previous work in Serratia marcescens also showed that carbon sources, such as glucose and maltose, have negative impacts on prodigiosin expression (Haddix et al, ).…”

“…Other transcriptional and post‐transcriptional regulators also control GV gene expression (Ramsay et al, ; Tashiro et al, ; Lee et al, ). At low cell densities a LuxR‐family quorum sensing transcription factor (SmaR) inhibits gvpA1‐gvpY expression through direct binding to its promoter and, indirectly, through repression of gvrA‐gvrC expression.…”

“…However, when cell population density rises the autoinducer N‐butanoyl‐L‐homoserine lactone (BHL; produced by SmaI) accumulates and binds to SmaR to de‐repress transcription of the GV operons (Ramsay et al, ; Tashiro et al, ). In addition to quorum sensing, the mRNA‐binding protein, RsmA (the homologue of the Escherichia coli CsrA protein) and the ribose operon repressor, RbsR, are also involved in gene regulation for GV production in S39006 (Ramsay et al, ; Lee et al, ). RsmA and RbsR are positive regulators of the gvrA‐gvrC operon and connect cell buoyancy regulation with carbon metabolism.…”