OmpR and RcsB abolish temporal and spatial changes in expression of flhD in Escherichia coli Biofilm

Abstract

BackgroundBiofilms are communities of bacteria that are characterized by specific phenotypes, including an increased resistance towards anti-microbials and the host immune system. This calls for the development of novel biofilm prevention and treatment options to combat infectious disease. In Escherichia coli, numerous global regulators have been implicated in the control of biofilm associated cell surface organelles. These include the flagellar regulator FlhD/FlhC, the osmoregulator EnvZ/OmpR, and the colanic acid activator RcsCDB. Using flow cell technology and fluorescence microscopy, we determined the temporal expression from flhD::gfp, ompR::gfp, and rcsB::gfp in E. coli biofilm, as well as the impact of the negative regulation of flhD by OmpR and RcsB. Spatial gene expression was investigated from flhD::gfp.ResultsThe temporal gene expression profile for flhD yielded an early peak at 12 h, a minimum of expression at 35 h, and a second increase in expression towards 51 h of biofilm development. In contrast, the ompR profile showed a peak at 35 h. A mutation in ompR abolished time dependence of flhD expression after the initial growth period of 12 h. Intriguingly, rcsB expression did not correlate inversely with flhD expression, yet a mutation in rcsB abolished time dependence of flhD expression as well. Spatially, expression of flhD was highest in the outermost layer of the biofilm in the parent strain. In ompR and rcsB mutants, flhD was expressed throughout the biofilm. Mutations in both, ompR and rcsB increased flhD expression throughout all temporal and spatial experiments. This increase was paralleled by reductions in biofilm amounts at four tested time points.ConclusionOur data lead to the conclusion that FlhD/FlhC and its regulation by OmpR and RcsB may be our first target mechanism for the development of novel biofilm prevention and treatment techniques.

Involvement of Two-Component Signaling on Bacterial Motility and Biofilm Development

“…This hypothesis is supported by a study by Samanta and coworkers, who performed temporal and spatial expression studies with the flhD promoter from the E. coli K-12 strain AJW678 that did not contain an IS element (25). The temporal transcription pattern of flhD was highest at 12 h and lowest at 35 h and increased again toward 51 h. The temporal profile for ompR was the inverse of that for flhD, and transcription of rcsB increased steadily throughout biofilm growth.…”

“…The spatial transcription of ompR was highest at the attachment surface and decreased with increasing distance from the surface, whereas transcription of rcsB increased toward the edge of the biofilm (131). Temporal and spatial patterns of flhD expression were abolished in ompR and rcsB mutants (25).…”

“…In addition, a knockout mutation in ompR exhibited increased flhDC transcription in a growth phaseindependent manner (24). An ompR mutant also exhibited a lack of temporal expression from the flhD promoter during biofilm development (25).…”

Deletion analysis of RcsC reveals a novel signalling pathway controlling poly-N-acetylglucosamine synthesis and biofilm formation in Escherichia coli

“…Remarkably, the OmpR protein has been implicated as an important regulator of biofilm formation in E. coli and in other bacteria such as Yersinia (Brzostek et al, 2012;Samanta et al, 2013).…”

Spontaneous mutations in the flhD operon generate motility heterogeneity in Escherichia coli biofilm

“…Three examples of adaptive gene expression leading to motility heterogeneity in a biofilm come from Bacillus subtilis [4], Escherichia coli K-12 [5], and Salmonella enterica [6–8]. In B. subtilis , genes of motility, matrix production, and sporulation are expressed in a spatiotemporal order [4].…”