RpoS is a pleiotropic regulator of motility, biofilm formation, exoenzymes, siderophore and prodigiosin production, and trade-off during prolonged stationary phase in Serratia marcescens.

Han Qin,Ying Liu,Xiyue Cao,Dairong Qiao,Hui Xu,Jia Jiang,Yi Cao,Weishao Lian,Marie-Joelle Virolle

doi:10.1371/journal.pone.0232549

Abstract

Prodigiosin is an important secondary metabolite produced by Serratia marcescens. It can help strains resist stresses from other microorganisms and environmental factors to achieve self-preservation. Prodigiosin is also a promising secondary metabolite due to its pharmacological characteristics. However, pigmentless S. marcescens mutants always emerge after prolonged starvation, which might be a way for the bacteria to adapt to starvation conditions, but it could be a major problem in the industrial application of S. marcescens. To identify the molecular mechanisms of loss of prodigiosin production, two mutants were isolated after 16 days of prolonged incubation of wild-type (WT) S. marcescens 1912768R; one mutant (named 1912768WR) exhibited reduced production of prodigiosin, and a second mutant (named 1912768W) was totally defective. Comparative genomic analysis revealed that the two mutants had either mutations or deletions in rpoS. Knockout of rpoS in S. marcescens 1912768R had pleiotropic effects. Complementation of rpoS in the ΔrpoS mutant further confirmed that RpoS was a positive regulator of prodigiosin production and that its regulatory role in prodigiosin biosynthesis was opposite that in Serratia sp. ATCC 39006, which had a different type of pig cluster; further, rpoS from Serratia sp. ATCC 39006 and other strains complemented the prodigiosin defect of the ΔrpoS mutant, suggesting that the pig promoters are more important than the genes in the regulation of prodigiosin production. Deletion of rpoS strongly impaired the resistance of S. marcescens to stresses but increased membrane permeability for nutritional competence; competition assays in rich and minimum media showed that the ΔrpoS mutant outcompeted its isogenic WT strain. All these data support the idea that RpoS is pleiotropic and that the loss of prodigiosin biosynthesis in S. marcescens 1912768R during prolonged incubation is due to a mutation in rpoS, which appears to be a self-preservation and nutritional competence (SPANC) trade-off.

Highlights

Serratia marcescens, a gram-negative bacterium belonging to the Enterobacteriaceae family, is found in a wide range of ecological niches in the environment [1]
Selective advantage for nonpigmented mutants during prolonged stationary phase To verify whether prolonged incubation would enrich the de novo emergence of prodigiosindefective individuals, we performed a series of long-term incubations of the WT strain 1912768R in lysogeny broth (LB) medium
We identified that RpoS was a positive regulator of prodigiosin biosynthesis in S. marcescens 1912768R; mutation of rpoS introduced pleiotropic phenotypes

Summary

Introduction

A gram-negative bacterium belonging to the Enterobacteriaceae family, is found in a wide range of ecological niches in the environment [1]. It is known for its ability to synthesize the striking red pigment prodigiosin, which is of great interest for its antimicrobial as well as immunosuppressant and other bioactive properties [2,3,4]. S. marcescens is a model strain for studying the regulation of prodigiosin production and other secondary metabolites, such as carbapenem and serratamolide [5,6,7]. Prodigiosin is synthesized by the prodigiosin biosynthesis gene cluster designated pig [6]. The stationary phase sigma factor rpoS or σs is a transcriptional factor and considered to be a global regulator that controls approximately 23% of the genes in the E. coli genome [19]. The effect of RpoS on the stress resistance and fitness of S. marcescens is unknown and remains to be explored

Methods

Results

Conclusion