Sec24 interaction is essential for localization and virulence‐associated function of the bacterial effector protein NleA

Abstract

Summary Enteropathogenic and enterohaemorrhagic Escherichia coli (EPEC and EHEC) are food‐borne pathogens that cause severe diarrhoeal disease in humans. Citrobacter rodentium is a related mouse pathogen that serves as a small animal model for EPEC and EHEC infections. EPEC, EHEC and C. rodentium translocate bacterial virulence proteins directly into host cells via a type III secretion system (T3SS). Non‐LEE‐encoded effector A (NleA) is a T3SS effector that is common to EPEC, EHEC and C. rodentium and is required for bacterial virulence. NleA localizes to the host cell secretory pathway and inhibits vesicle trafficking by interacting with the Sec24 subunit of mammalian coatamer protein II complex (COPII). Mammalian cells express four paralogues of Sec24 (Sec24A–D), which mediate selection of cargo proteins for transport and possess distinct, but overlapping cargo specificities. Here, we show that NleA binds Sec24A–D with two distinct mechanisms. An NleA protein variant with greatly diminished interaction with all Sec24 paralogues does not properly localize, does not inhibit COPII‐mediated vesicle budding, and does not confer virulence in the mouse infection model. Together, this work provides strong evidence that the interaction and inhibition of COPII by NleA is an important aspect of EPEC‐ and EHEC‐mediated disease.

Bacteria Fighting Back: How Pathogens Target and Subvert the Host Innate Immune System

“…The first step of cargo packaging and vesicle budding from the ER is regulated by NleA/EspI, which acts to prevent normal COPII mediated vesicle trafficking, thereby inhibiting ER to Golgi transport (Figure 2) (Kim et al, 2007). Although this mechanism of regulation is not fully understood, it is clear that NleA/EspI binds to Sec23/24, a heterodimeric subunit of the CopII coat, in a Sar1 and lipid-dependent manner and decreases the rate of GTP hydrolysis by Sar1 (Thanabalasuriar et al, 2012). This interaction has been shown to stabilize COPII, which presumably leads to a defect in recycling and thus an abnormal buildup of COPII coated vesicles that slows proper sorting of anterograde trafficked vesicular cargo.…”

Recent Advances in Understanding Enteric Pathogenic Escherichia coli

“…EIEC/Shigella escape submucosal macrophages by induction of macrophage cell death followed by basolateral invasion of colonocytes and lateral spread. tory functions, Nle effectors such as EspJ have antiphagocytic properties (130), while NleA alters host protein secretion (131) and tight junction integrity (132) and inhibits vesicle trafficking (133). NleH is capable of modulating apoptotic response (134).…”

Tight Junction Disruption Induced by Type 3 Secretion System Effectors Injected by Enteropathogenic and Enterohemorrhagic Escherichia coli

“…The NleA mutants NleAI414A/I416A and NleAI414A/I416A/ΔIIQ transfected in HeLa cells do not co-localized with the giantin Golgi marker as the native NleA does. Additionally, NleAI414A/I416A/ΔIIQ mutant showed a diminished binding to all four mammalian Sec24 paralogs (Thanabalasuriar et al, 2012). …”

“…Mice infected with a strain of C. rodentium engineered to express NleAI414A/I416A/ΔIIQ instead of native NleA was completely avirulent in mice, providing strong evidence that Sec24 interaction with NleA is critical for bacteria virulence (Thanabalasuriar et al, 2012). While the same susceptible C3H/HeOuJ mice infected with C. rodentium showed virtually no localized staining of claudin-3 in the TJs.…”