TheE. colitranscription factor GrlA is regulated by subcellular compartmentalization and activated in response to mechanical stimuli

Abstract

EnterohemorrhagicEscherichia coli(EHEC) is a foodborne pathogen that colonizes the gastrointestinal tract and has evolved intricate mechanisms to sense and respond to the host environment. Upon the sensation of chemical and physical cues specific to the host’s intestinal environment, locus of enterocyte effacement (LEE)-encoded virulence genes are activated and promote intestinal colonization. The LEE transcriptional activator GrlA mediates EHEC’s response to mechanical cues characteristic of the intestinal niche, including adhesive force that results from bacterial adherence to epithelial cells and fluid shear that results from intestinal motility and transit. GrlA expression and release from its inhibitor GrlR was not sufficient to induce virulence gene transcription; mechanical stimuli were required for GrlA activation. The exact mechanism of GrlA activation, however, remained unknown. We isolated GrlA mutants that activate LEE transcription, independent of applied mechanical stimuli. In nonstimulated EHEC, wild-type GrlA associates with cardiolipin membrane domains via a patch of basic C-terminal residues, and this membrane sequestration is disrupted in EHEC that expresses constitutively active GrlA mutants. GrlA transitions from an inactive, membrane-associated state and relocalizes to the cytoplasm in response to mechanical stimuli, allowing GrlA to bind and activate the LEE1 promoter. GrlA expression and its relocalization in response to mechanical stimuli are required for optimal virulence regulation and colonization of the host intestinal tract during infection. These data suggest a posttranslational regulatory mechanism of the mechanosensor GrlA, whereby virulence gene expression can be rapidly fine-tuned in response to the highly dynamic spatiotemporal mechanical profile of the gastrointestinal tract.