Temperature-responsive sensing regulates biocontrol factor expression in Pseudomonas fluorescens CHA0

Abstract

In the plant-beneficial, root-colonizing strain Pseudomonas fluorescens CHA0, the Gac/Rsm signal transduction pathway positively regulates the synthesis of biocontrol factors (mostly antifungal secondary metabolites) and contributes to oxidative stress response via the stress sigma factor RpoS. The backbone of this pathway consists of the GacS/GacA two-component system, which activates the expression of three small regulatory RNAs (RsmX, RsmY, RsmZ) and thereby counters translational repression exerted by the RsmA and RsmE proteins on target mRNAs encoding biocontrol factors. We found that the expression of typical biocontrol factors, that is, antibiotic compounds and hydrogen cyanide (involving the phlA and hcnA genes), was significantly lower at 35 1C than at 30 1C. The expression of the rpoS gene was affected in parallel. This temperature control depended on RetS, a sensor kinase acting as an antagonist of the GacS/GacA system. An additional sensor kinase, LadS, which activated the GacS/GacA system, apparently did not contribute to thermosensitivity. Mutations in gacS or gacA were epistatic to (that is, they overruled) mutations in retS or ladS for expression of the small RNAs RsmXYZ. These data are consistent with a model according to which RetS-GacS and LadS-GacS interactions shape the output of the Gac/ Rsm pathway and the environmental temperature influences the RetS-GacS interaction in P. fluorescens CHA0.

Phenotypic Switching in Pseudomonas brassicacearum Involves GacS- and GacA-Dependent Rsm Small RNAs

“…3) in strain NFM421 showed differential expression patterns, suggesting that in addition to the GacS-GacA system further regulators may be involved in regulating transcription of rsmX, rsmY, and rsmZ. In P. aeruginosa and P. fluorescens CHA0, two sensor kinases, LadS and RetS, inversely influence the output of the GacS sensor (18,40). Two other regulators, PsrA and HptB, have recently been shown to modulate rsmZ and rsmY expression, respectively.…”

Context-dependent symbioses and their potential roles in wildlife diseases

“…They form natural mutualisms with many animals and plants, in which the hosts gain bacterial protection from pathogens [10,47]. For example, Pseudomonas fluorescens produces antibiotics that can benefit its plant hosts, but expression of the genes for antibiotic production is downregulated at 358C compared with 308C [48]. Pseudomonas sp.…”

GacA-Controlled Activation of Promoters for Small RNA Genes in Pseudomonas fluorescens

“…Electroporation of bacterial cells with plasmid DNA was done as described previously (15). Conditions for amplifying PCR fragments have been detailed elsewhere (26). Primers are listed in Table S1 in the supplemental material.…”

“…Chromosomal insertion of the mutated rsmY-lacZ fusions was achieved by recruiting them from the relevant plasmids on 3.5-kb EcoRI-XhoI fragments, which were blunted and cloned into the mini-Tn7 vector pME6182 digested with SmaI. The constructs obtained (pME7682 to pME7685, pME7689, pME7694 to pME7697, and pME7699) and pUX-BF13 were introduced into recipient strains by coelectroporation, allowing transposition of the constructs into the chromosomal Tn7 attachment site (26,59).…”

“…For pME497-dependent mobilization (57) of suicide plasmids (that is, pME3087 derivatives) from E. coli to P. fluorescens, chloramphenicol (Cm) at 10 g ml Ϫ1 and Tc were used to select for the recipient having integrated the suicide plasmid. Enrichment for Tc-sensitive strains, from which the suicide plasmid had been excised, was performed as previously described (26). Routine incubation temperatures were 30°C for P. fluorescens and 37°C for E. coli.…”