Abstract
The locus of enterocyte effacement (LEE) is essential for virulence of enterohaemorrhagic Escherichia coli (EHEC) and enteropathogenic E. coli (EPEC). The 41 genes of the LEE encode type III secretion system proteins and three associated regulators: Ler, GrlA and GrlR. Ler is a positive regulator for most of the LEE operons, including grlRA. GrlA controls the expression of ler, ehxCABD and flhDC operons. GrlR binds to GrlA and suppresses its function. Here we report the crystal structure of GrlR-GrlAΔ (aa 1-106) complex (2:1) and its functional characterization. We show that GrlR interacts with the Helix-Turn-Helix motif of GrlA. Moreover, GrlA binds to the promoter DNA fragments of ler, ehxCABD and flhDC, and GrlR outcompetes with these promoter DNA sequences for the Helix-Turn-Helix motif of GrlA. These findings provide mechanistic insight into a regulatory module for the virulence of EPEC and EHEC, two important pathogens that cause devastating diseases.
Highlights
The locus of enterocyte effacement (LEE) is essential for virulence of enterohaemorrhagic Escherichia coli (EHEC) and enteropathogenic E. coli (EPEC)
This histonelike nucleoid-structuring (H-NS)-mediated repression, can be countered by Ler (LEE-encoded regulator), a protein encoded by the first gene in the LEE1 operon that functions as a positive regulator of other LEE operons (LEE2–LEE5)[4]
The expression of the LEE1 operon is strictly dependent on binding of the integration host factor protein immediately upstream to the LEE1 promoter[4], and activation of LEE1 expression is promoted by two redundant positive regulators, PerC5 and GrlA
Summary
The locus of enterocyte effacement (LEE) is essential for virulence of enterohaemorrhagic Escherichia coli (EHEC) and enteropathogenic E. coli (EPEC). The crystal structure of the complex revealed that GrlR interacts with the HTH motif region of GrlAD (Fig. 3a,b and Supplementary Fig. S2a,b). We further verified the role of the key residues of GrlR in mediating GrlA binding, which shows that alanine substitution at D35 and E60 of GrlR could abolish its binding to MBP-GrlAD (Fig. 5b and Supplementary Fig. S5d).
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