The Response Regulator RcsB Activates Expression of Mat Fimbriae in Meningitic Escherichia coli

Abstract

The common colonization factor of Escherichia coli, the Mat (also termed ECP) fimbria, functions to advance biofilm formation on inert surfaces as well as bacterial adherence to epithelial cells and subsequent colonization. We used global mini-Tn5 transposon mutagenesis to identify novel regulators of biofilm formation by the meningitic E. coli isolate IHE 3034. Of the 4,418 transformants, we found 17 that were impaired in biofilm formation. Most of these mutants were affected in lipopolysaccharide synthesis and were reduced in growth but not in Mat fimbria expression. In contrast, two mutants grew well but did not express Mat fimbria. The insertions in these two mutants were located at different sites of the rcsB gene, which encodes a DNA-binding response regulator of the Rcs response regulon. The mutations abrogated temperature-dependent biofilm formation by IHE 3034, and the phenotype correlated with loss of mat expression. The defect in biofilm formation in the rcsB mutant was reversed upon complementation with rcsB as well as by overexpression of structural mat genes but not by overexpression of the fimbria-specific activator gene matA. Monitoring of the mat operon promoter activity with chromosomal reporter fusions showed that the RcsB protein and an RcsAB box in the mat regulatory region, but not RcsC, RcsD, AckA, and Pta, are essential for initiation of mat transcription. Gel retardation assays showed that RcsB specifically binds to the mat promoter DNA, which enables its function in promoting biofilm formation by E. coli. Bacterial growth in biofilm provides many advantages compared to a planktonic lifestyle. In a biofilm, bacteria gain resistance to antimicrobial agents and defense systems of the host as well as better protection from predation and other environmental assaults, such as hydrodynamic shear forces and desiccation (9,20). This protective effect probably explains why bacterial biofilm growth is common in many environmental, clinical, and industrial settings and why it complicates the elimination of biofilmassociated bacteria. Following initial attachment to a surface, bacterial cells undergo profound physiological and metabolic adjustments along the transition from a single-cell state into a communal lifestyle. This complex process of biofilm development requires integration of a variety of environmental, cell-to-cell, and intracellular signals into an interplay of regulatory networks (25,50).Escherichia coli is a commensal bacterium in the gastrointestinal tract as well as a pathogen of intra-and extraintestinal body sites. E. coli expresses an arsenal of surface appendages that confer tissue-adhesive and biofilm formation capacity upon the bacterium (27, 28). Meningitis-associated and temperature-regulated (Mat) fimbriae (or E. coli common pili [ECP]) promote biofilm growth on inert surfaces (34) and adhesion to human epithelial cell lines in vitro (4,32,53,55). Available data also indicate that the fimbria is expressed in vivo in the human body and in the intestine of infant mouse (32, 5...

Interaction of the RcsB Response Regulator with Auxiliary Transcription Regulators in Escherichia coli

“…2A). The data are in accordance with previous results showing that MatA (EcpR) activates the mat operon and that RcsB is required for mat operon expression (22,23). Taken together, the results confirm that both proteins, MatA and RcsB, are required for activation of the matA CFT073 promoter, and they indicate that a MatA-RcsB heterodimer activates the mat promoter.…”

“…The moderate difference in activation of Pmat by the RcsB-D56 mutants and by wild-type RcsB (2468 units) suggests that transcriptional activation of the matA CFT073 promoter by MatARcsB heterodimers is not (or is only weakly) dependent on RcsB phosphorylation. These results are in accordance with expression studies in NMEC strain IHE3034, where deletion of the Rcs phosphorelay genes rcsC and rcsD did not affect mat expression (22).…”

“…One of the main elements in this control is the availability of the interaction partners. Expression of all of the corresponding genes is repressed by the nucleoid-associated and global repressor protein H-NS (22,47,54,55) (Fig. 7).…”

“…However, no target loci of DctR (25) are known, whereas MatA (EcpR) activates the mat (ecp) operon in NMEC, enterohemorrhagic E. coli, and other E. coli but not in E. coli K12 (20,22,23). To analyze the relevance of the MatA-RcsB and DctR-RcsB protein interaction in gene regulation, we first needed to identify target genes that may serve as reporters for DctR and MatA, respectively.…”

The fimbriae activator MatA switches off motility in Escherichia coli by repression of the flagellar master operon flhDC

“…Our previous results and this report demonstrate that MatA exerts a dual regulatory function on the choice of a planktonic/sessile lifestyle, and further support the existence of an antagonistic control of attachment and motility in E. coli. Our ongoing work indicates that MatA is tightly integrated into two separate regulatory circuits, H-NS-MatA-flhDC and RcsB-MatA-flhDC, both of which control expression of the flagellum (T. A. Lehti and others, unpublished results; Lehti et al, 2012).…”