Abstract
The Shigella type III effector IpaA contains three binding sites for the focal adhesion protein vinculin (VBSs), which are involved in bacterial invasion of host cells. Here, we report that IpaA VBS3 unexpectedly binds to talin. The 2.5Å resolution crystal structure of IpaA VBS3 in complex with the talin H1-H4 helices shows a tightly folded α-helical bundle, which is in contrast to the bundle unraveling upon vinculin interaction. High-affinity binding to talin H1-H4 requires a core of hydrophobic residues and electrostatic interactions conserved in talin VBS H46. Remarkably, IpaA VBS3 localizes to filopodial distal adhesions enriched in talin, but not vinculin. In addition, IpaA VBS3 binding to talin was required for filopodial adhesions and efficient capture of Shigella. These results point to the functional diversity of VBSs and support a specific role for talin binding by a subset of VBSs in the formation of filopodial adhesions.
Highlights
The cytoskeletal linkers talin and vinculin play important and distinct roles in integrin-mediated cell adhesion (Atherton et al, 2016; Klapholz and Brown, 2017)
Our results suggest that binding of IpaA VBS3 to a semistretched talin conformer stabilizes filopodial adhesions and stimulates bacterial capture
A significant decrease in the percentage of foci forming talin coat structures was observed for the ipaA mutant strains expressing IpaA DVBS1–2 or IpaA DVBS3 compared with full-length IpaA, with 19% ± 0.7% and 25% ± 5% relative to 52% ± 6.7%, respectively (Figure 1E). These results indicate that talin recruitment could occur via IpaA VBS3 or via IpaA VBS1–2, possibly through vinculin-talin interactions
Summary
The cytoskeletal linkers talin and vinculin play important and distinct roles in integrin-mediated cell adhesion (Atherton et al, 2016; Klapholz and Brown, 2017). Talin consists of an amino-terminal FERM domain connected to a large rod domain containing vinculin binding sites (VBSs) buried in helical bundles (Calderwood et al, 2013; Lagarrigue et al, 2016; Yan et al, 2015). Stretching of the talin molecule due to actomyosin-dependent contractility leads to the unveiling of VBSs buried in helical bundles along the talin rod domain (Yan et al, 2015). Talin VBSs activate vinculin that further tethers actin filaments to adjust adhesion strength in response to substrate stiffness (Gingras et al, 2005; Yan et al, 2015). In vitro measurements on single molecules indicate that unveiling of VBSs in various talin bundles occurs with a defined hierarchy, suggesting different talin activation states
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