Functional identification of the prnABCD operon and its regulation in Serratia plymuthica

Abstract

The antibiotic pyrrolnitrin (PRN) is a tryptophan-derived secondary metabolite that plays an important role in the biocontrol of plant diseases due to its broad-spectrum of antimicrobial activities. The PRN biosynthetic gene cluster remains to be characterised in Serratia plymuthica, though it is highly conserved in PRN-producing bacteria. To better understand PRN biosynthesis and its regulation in Serratia, the prnABCD operon from S. plymuthica G3 was cloned, sequenced and expressed in Escherichia coli DH5α. Furthermore, an engineered strain prnind which is a conditional mutant of G3 prnABCD under the control of the Ptac promoter was constructed. This mutant was able to overproduce PRN with isopropylthiogalactoside (IPTG) induction by overexpressing prnABCD, whilst behaving as a conditional mutant of G3 prnABCD in the absence of IPTG. These results confirmed that prnABCD is responsible for PRN biosynthesis in strain G3. Further experiments involving lux-/dsRed-based promoter fusions, combined with site-directed mutagenesis of the putative σ extended -10 region in the prnA promoter, and liquid chromatography-mass spectrometry (LC-MS) analysis extended our previous knowledge about G3, revealing that quorum sensing (QS) regulates PRN biosynthesis through cross talk with RpoS, which may directly activated prnABCD transcription. These findings suggest that PRN in S. plymuthica G3 is produced in a tightly controlled manner, and has diverse functions, such as modulation of cell motility, in addition to antimicrobial activities. Meanwhile, the construction of inducible mutants could be a powerful tool to improve PRN production, beyond its potential use for the investigation of the biological function of PRN.

Screening of the High-Rhizosphere Competent Limoniastrum monopetalum’ Culturable Endophyte Microbiota Allows the Recovery of Multifaceted and Versatile Biocontrol Agents

“…Several other reports documented our reported genera from various crops: As an example, endophytes Bacillus licheniformis , Klebsiella sp., Pantoea sp., Bacillus sp. and Bacillus cereus isolated from maize roots [51], Bacillus sp., and Enterobacter sp., isolated from corn roots [53] and Serratia plymuthica strain G3 isolated from wheat stems [54].…”

Expanding the application of tryptophan: Industrial biomanufacturing of tryptophan derivatives

“…The prnABCD operon was cloned from plymuthica G3 and expressed in E. coli DH5α, the mutant was able to overproduce pyrrolnitrin with isopropyl β-D-thiogalactoside (IPTG) induction by overexpressing prnABCD ( Liu et al, 2018 ). In the wild-type strain, the amount of pyrrolnitrin secreted is small ( Pawar et al, 2019 ).…”

Serratia inhibens sp. nov., a new antifungal species isolated from potato (Solanum tuberosum)

“…Several species and strains of Serratia have been isolated from hospitals [1] and other environments; notably, water [2] and plant surfaces [3–7] have been reported to host Serratia species. Many members of the genus Serratia have been described to produce secondary metabolites including the red pigment prodigiosin [8–10], and antimicrobial compounds [4, 5, 8, 11–17]. Serratia plymuthica strains in particular, have been reported to produce secondary metabolites that could be used in biocontrol, for example strain RVH1 produces three types of zeamine [15], strain A153 synthesizes oocydins, andrimide, zeamine, and pyrrolnitrin [4], strain 4RX5 produces oocydin [5], strain 4RX13 synthesizes sodorifen [11, 12] and strain PRI-2C produces pyrrolnitrin [18].…”