A transcriptome study of the QseEF two-component system and the QseG membrane protein in enterohaemorrhagic Escherichia coli O157 : H7

Abstract

QseE is a sensor kinase that responds to epinephrine, sulfate and phosphate. QseE constitutes a two-component signalling system together with the QseF sigma(54)-dependent response regulator. Encoded within the same operon as qseEF is the qseG gene, which encodes a membrane protein involved in the translocation of a type III secretion effector protein of enterohaemorrhagic Escherichia coli (EHEC) into epithelial cells. The qseEGF genes also form an operon with the glnB gene, which encodes the E. coli nitrogen sensor PII protein. Here we report a transcriptome analysis comparing qseE, qseF andqseG single mutants with the wild-type strain. This study revealed that the proteins encoded by these genes play a modest but significant role in iron uptake. Although QseEFG regulate genes involved in nitrogen utilization, these proteins do not play a notable role in nitrogen metabolism. In addition, QseEFG regulate transcription of the rcsBC and phoPQ two-component systems, linking several signal transduction pathways. The similarity of the microarray profiles of these mutants also indicates that these proteins work together. These data indicate that QseEFG are involved in the regulation of virulence and metabolism in EHEC.

Fig. 1.

Venn diagrams showing the number of overlapping up- or downregulated genes between the qseE, F and G mutant strains compared with the WT.

Fig. 2.

qseEFGglnB affects nitrogen gene regulation. (a) Heat map showing genes involved in nitrogen regulation. These genes are primarily downregulated in qseE, qseG and qseF. (b) Growth curves showing the growth of each mutant in minimal media and media into which glutamine was titrated. In both cases, there was no significant difference in growth between the WT EHEC and the qseE, qseG and qseF mutant strains.

Fig. 3.

(a) Real-time RT-PCR showing the regulation of iron-utilization genes in the qseE, qseG and qseF mutant backgrounds. Significant downregulation of fepE and entB expression was seen. (b) Colony growth of mutant and complemented strains on media containing either 350 μM DPD or 350 μM DPD with 8 μM haemin. Each strain was equally affected by DPD addition and able to recover equally to the WT when haemin was added. (c) Growth of mutant strains versus WT EHEC in DMEM. No growth defect was seen in any of the mutant strains. (d) Growth of mutant strains versus WT EHEC in DMEM containing 350 μM DPD. The qseE and qseG mutants grew slightly more slowly than the WT, and this was complemented by the addition of the gene expressed on a plasmid. *P ≤0.05 using Student's t test.

Fig. 4.

QseEFG cross-talks with additional signalling systems. Real-time RT-PCR analysis was used to investigate the transcriptional regulation exerted by QseEFG on other two-component systems. Overall, expression of phoP and phoQ was downregulated in qseE, qseF and qseG mutants, while rcsB and rcsC were upregulated in the mutant strains. *P ≤0.05; **P ≤0.005 using Student's t test.