Tyrosine phosphorylation controls cortactin binding to two enterohaemorrhagic Escherichia coli effectors: Tir and EspFu/TccP

Abstract

SummaryEnterohaemorrhagic Escherichia coli (EHEC) is an important food-borne pathogen that, upon infection, causes destruction of the microvilli brush border of intestinal cells. EHEC is able to recruit several host cell proteins and induce actin accumulation beneath its adherence site, forming a pedestal-like structure upon which the bacterium is firmly attached. Injection of bacterial effectors into the host cells is required to trigger the recruitment and activation of proteins, such as cortactin, neural Wiskott-Aldrich syndrome protein (N-WASP) and Arp2/3 complex, directly involved in the actin polymerization process. We found that cortactin, an actin-binding protein, has a pivotal role during pedestal formation by EHEC. Cortactin was found to bind directly to two important virulence factors of EHEC, Tir and EspF u, which are translocated into the host cells during infection. Binding of cortactin to these effectors is dependent upon tyrosine phosphorylation and a balance between tyrosine phosphorylation and dephosphorylation of cortactin is required to regulate pedestal formation by EHEC.

Cortactin Recruitment by Enterohemorrhagic Escherichia coli O157:H7 during Infection In Vitro and Ex Vivo

“…For example, Listeria monocytogenes and Shigella flexneri exploit cortactin for invasion of mammalian host cells (reviewed in reference 25). In addition, cortactin is reported to bind the N terminus of Tir (8) and to play a role in triggering actin polymerization at the site of EHEC adhesion to eukaryotic cells (7). Here, we have studied the recruitment of cortactin during EHEC infection of cultured cells and human IVOC.…”

“…In vitro, cortactin, via its SH3 domain, reportedly binds directly to the N terminus of Tir (8), which contains a polyproline region (amino acids 17 to 23) that is a putative SH3 domain-binding site. We therefore investigated if the N terminus of Tir and the polyproline region contained within it have a role in cortactin recruitment under adherent bacteria.…”

“…Recently, Cantarelli et al have shown that cortactin can bind the N terminus of Tir and the proline-rich repeats of TccP (8). The fact that cortactin can activate N-WASP by itself, is able to bind Tir, and can interact with TccP led to the hypothesis that it might be the adaptor protein connecting Tir and TccP during infection (8). Alternatively, cortactin may contribute to an alternative pathway of actin assembly, such as the inefficient TccP-independent actin polymerization triggered by EHEC.…”

Membrane-deforming Proteins Play Distinct Roles in Actin Pedestal Biogenesis by Enterohemorrhagic Escherichia coli

“…Finally, it is important to note that the proline-rich sequences within EspF U are additionally recognized by the SH3 domains of IRSp53 and IRTKS, I-BAR proteins that mediate the recruitment of EspF U to the EHEC transmembrane receptor, Tir (13,14). EspF U has also been reported to bind the SH3 domain of cortactin (46), another actin-associated factor. Thus, with numerous potential host cell targets, the mechanisms by which the EspF U repeat region organizes a complex signaling platform during actin pedestal formation are only beginning to be elucidated.…”

Control of Gliotactin localization and levels by tyrosine phosphorylation and endocytosis is necessary for survival of polarized epithelia

“…S1A). The former is a mutant version of Gliotactin that should produce a constitutive state of dephosphorylation (GliFF), and the latter is a form that should mimic phosphorylation and be in a constitutive state of phosphorylation (GliDD) (Cantarelli et al, 2007;Cuevas et al, 2001;Huh et al, 2004;Meyer et al, 2003). As a control, we generated a wild-type Gliotactin construct (GliWT), and all three constructs were placed under the control of the GAL4 upstream activating sequence (UAS) sequence (Brand and Perrimon, 1993).…”