The Eukaryotic Linear Motif resource for
Functional Sites in Proteins
Accession:
Functional site class:
Vinculin binding site
Functional site description:
Vinculin was initially discovered in focal adhesions and adherens junctions as a component that provides a structural and functional link between F-actin and cell adhesion molecules of the cadherin and integrin families. Vinculin is activated by relieving its closed state by severing its head-tail hydrophobic interaction when a vinculin activator protein like Talin, α actinin or the bacterial proteins IpaA or sca4 bind to it. These proteins all interact with the N-terminal head domain of vinculin (Vh1) through an amphipathic α-helical region known as the vinculin binding site (VBS) and induce a helix-bundle conversion in vinculin’s head domain allowing vinculin to bind to its other partners and reorganize the actin cytoskeleton in adherens junctions and focal adhesions. Vinculin is essential for embryonic development and also has tumor suppressor functions. Its over-expression impairs cell motility and tumor cell metastasis whereas its suppression augments cell motility and tumorigenesis.

ELM Description:
The Talin-class of vinculin binding site (VBS), also present in bacterial vinculin activators, is formed by a set of hydrophobic residues on one face of an amphipathic helix which can otherwise be placed back into a folded helical bundle domain. Upon interaction with the head domain of vinculin (Vh), the α-helix is inserted between helices α1 and α2 of Vh and the buried residues are exposed. Binding occurs largely through van der Waals interactions, whereby the hydrophobic face of VBS interacts with the hydrophobic core of the N-terminal helical bundle of Vh. Sequence alignment of different VBS shows a consensus pattern of nine hydrophobic positions in a span of nineteen residues. Position 5 is usually Ala and is always a small side chain. Position 15 is also tightly restricted to Leu-Met-Val. All other hydrophobic positions tolerate greater variation. The current regular expression matches the known high affinity (low nanomolar) binders and some but not all the peptides that were confirmed as binders in a SPOT array peptide analysis (Gingras,2005). While the SPOT preferences are broadly in agreement with the known VBS motifs, it is likely that array values reflect lower affinity binding than is required in a cytoskeletal complex subject to mechanical force.

In addition, the current regular expression matches the α-helices in the crystal structures 4DJ9 and 1ZW2 that correspond to the predicted helices 50 and 58 in talin1 that were shown to bind to Vh1 in the SPOT array, demonstrating the binding. However, it is uncertain whether they represent true instances since they cover the binding sites for integrin and F-actin, respectively.
Pattern: [VMIL][MILFYPA][^P][TASKHC][AVSC][^P][^P][ILVM][^P][^P][^P][LMTVI][^P][^P][LMV][ILVMA][^P][^P][AIVLMT]
Pattern Probability: 5.538e-07
Present in taxon: Eukaryota
o See 12 Instances for LIG_Vh1_VBS_1
o Abstract
Vinculin was initially identified as a protein highly enriched in cell-to-cell contact and underlying substratum. It has a crucial role in the maintenance and regulation of cell adhesion and migration. In the cytosol, vinculin exists in an inactivated/auto inhibited state and becomes activated when recruited to cell–cell and cell–matrix adherens-type junctions and it anchors these adhesion complexes to the actin cytoskeleton by binding to talin in integrin complexes or to α-actinin in cadherin junctions. Vinculin is made up of a globular head domain (Vh) linked to a tail domain (Vt) by a short proline rich sequence. An intramolecular interaction between head and tail domains masks numerous ligand binding sites in all three regions (Ziegler,2006). The interactions between head and tail domains are so strong that vinculin activation generally requires two or more ligands (combinatorial activation pathway) to relieve the intramolecular head–tail interaction though, in some cases, exposure to a high affinity vinculin binding site present in talin or α-actinin might be sufficient (Bays,2017).

The interaction between the head domain of vinculin and some of its interacting proteins such as talin, α-actinin, IpaA of Shigella flexneri and Rickettsial sca4 antigen induces a marked conformational change in Vh, creating a novel helical bundle structure that displaces Vt from Vh (Izard,2004). These vinculin activators are characterized by vinculin binding sites (VBSs). VBSs are buried amphipathic α-helices of about 19 residues in length that reside within four- or five-helix bundle domains (Goult,2013). The key interacting residues are buried inside the core of the helical bundles and are released from their buried locales upon interaction with the Vh domain. VBSs activate vinculin by burying within and inducing helix bundle conversion in the N-terminal four-helix subdomain of Vh, which displaces Vt from a distance and exposes its cryptic F-actin binding sites. Eleven potential vinculin binding sites have been described in the talin rod domain, most of them showing low affinity and were not able to induce helix bundle conversion (Gingras,2005). The functional VBS helices must bind with low nanomolar affinity as the complex is dependent on force-induced activation of the talin rod domain (del Rio,2009). The VBS of α-actinin binds to Vh1 in an inverted orientation compared to the binding of talin’s VBSs and provokes distinct structural changes in vinculin (Bois,2005). Thus, vinculin’s Vh1 domain acts as an actin filament-linking molecular switch that undergoes distinct structural changes provoked by talin and α-actinin binding in focal adhesions versus adherens junctions, respectively.

Bacterial pathogens like Shigella flexneri and Rickettsia spp. have developed protein structures that resemble the architecture of the VBSs in talin with the same biological function and co-localization with talin that can induce actin polymerization without the need of integrin activation (Hamiaux,2006, Park,2011). The IpaA, invasin protein of Shigella harbors three tandem high affinity VBS motifs which are readily available and act in a redundant manner to bind and activate vinculin. The IpaA VBSs efficiently mimic the talin VBS-induced vinculin activation and actin filament hijack by IpaA is required for maximal entry of Shigella into host cells. It can simultaneously bind three Vh1 molecules and thus stabilize the focal adhesion-like complexes seen at pathogen entry sites (Park,2011).

Surface cell antigen 4 (sca4) is a protein from R. rickettsii that contains two VBSs, sca4-VBS-N and sca4-VBS-C, where the former resembles the binding of the talin or IpaA VBSs, while the later contains a Pro at position +14 causing a kink in the helix structure. The crystal structure of sca4-VBS-C with vinculin shows that the position of the α-helix α1 in the Vh1 domain of vinculin has a dramatic movement compared to its corresponding position in other structures containing non-kinked helices (Park,2011), therefore exemplifying a third variant Vh1 binding-motif.
o 9 selected references:

o 12 GO-Terms:

o 12 Instances for LIG_Vh1_VBS_1
(click table headers for sorting; Notes column: =Number of Switches, =Number of Interactions)
Acc., Gene-, NameStartEndSubsequenceLogic#Ev.OrganismNotes
P18010 ipaA
IPAA_SHIFL
492 510 IFEASKKVTNSLSNLISLIG TP 4 Shigella flexneri [new]
P54939 TLN1
TLN1_CHICK
856 874 LLSAAKILADATAKMVEAAK TP 3 Gallus gallus (Chicken) [new]
P26039 Tln1
TLN1_MOUSE
2345 2363 ILEAAKSIAAATSALVKAAS FP 4 Mus musculus (House mouse) [new]
P26039 Tln1
TLN1_MOUSE
1950 1968 LIECARRVSEKVSHVLAALQ TP 3 Mus musculus (House mouse) [new]
P26039 Tln1
TLN1_MOUSE
1633 1651 LAGHSRTVSDSIKKLITSMR TP 4 Mus musculus (House mouse) [new]
P26039 Tln1
TLN1_MOUSE
856 874 LLSAAKILADATAKMVEAAK TP 3 Mus musculus (House mouse) [new]
P26039 Tln1
TLN1_MOUSE
608 626 LLQAAKGLAGAVSELLRSAQ TP 3 Mus musculus (House mouse) [new]
P54939 TLN1
TLN1_CHICK
1949 1967 LIESARKVSEKVSHVLAALQ TP 5 Gallus gallus (Chicken) [new]
B0BXR4 RrIowa_0797
B0BXR4_RICRO
413 431 LLNAATALSGSMQYLLNYVN TP 3 Rickettsia rickettsii str. Iowa [new]
P18010 ipaA
IPAA_SHIFL
612 630 IYKAAKDVTTSLSKVLKNIN TP 7 Shigella flexneri [new]
P18010 ipaA
IPAA_SHIFL
566 584 IYEKAKEVSSALSKVLSKID TP 6 Shigella flexneri [new]
Q9Y490 TLN1
TLN1_HUMAN
608 626 LLQAAKGLAGAVSELLRSAQ TP 2 Homo sapiens (Human) [new]
Please cite: ELM 2016-data update and new functionality of the eukaryotic linear motif resource. (PMID:26615199)

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