scholarly journals Structural basis of αE-catenin–F-actin catch bond behavior

2020 ◽  
Author(s):  
Xiao-Ping Xu ◽  
Sabine Pokutta ◽  
Miguel Torres ◽  
Mark F. Swift ◽  
Dorit Hanein ◽  
...  
Keyword(s):  
Bound States ◽  
Mechanical Load ◽  
Bound State ◽  
Actin Binding ◽  
Structural Basis ◽  
Catch Bond ◽  
Bond Behavior ◽  
Cell Matrix ◽  
Actin Binding Domain ◽  
Cell Cell

ABSTRACTCell-cell and cell-matrix junctions transmit mechanical forces during tissue morphogenesis and homeostasis. α-Catenin links cell-cell adhesion complexes to the actin cytoskeleton, and mechanical load strengthens its binding to F-actin in a direction-sensitive manner. This so-called catch bond behavior is described by a model in which force promotes a transition between weak and strong actin-bound states. We describe the cryo-electron microscopy structure of the F-actin-bound αE-catenin actin-binding domain, which in solution forms a 5-helix bundle. Upon binding to actin, the first helix of the bundle dissociates and the remaining four helices and connecting loops rearrange to form the interface with actin. Deletion of the N-terminal helix produces strong actin binding in the absence of force. Our analysis explains how mechanical force applied to αE-catenin or its homolog vinculin favors the strongly bound state, and the dependence of catch bond strength on the direction of applied force.

scholarly journals Structural basis of αE-catenin–F-actin catch bond behavior

eLife ◽  
2020 ◽  
Vol 9 ◽  
Author(s):  
Xiao-Ping Xu ◽  
Sabine Pokutta ◽  
Megan Torres ◽  
Mark F Swift ◽  
Dorit Hanein ◽  
...  
Keyword(s):  
Bound States ◽  
Mechanical Load ◽  
Optical Trap ◽  
Bound State ◽  
Actin Binding ◽  
Structural Basis ◽  
Catch Bond ◽  
Strong State ◽  
Cell Matrix ◽  
Cell Cell

Cell-cell and cell-matrix junctions transmit mechanical forces during tissue morphogenesis and homeostasis. α-Catenin links cell-cell adhesion complexes to the actin cytoskeleton, and mechanical load strengthens its binding to F-actin in a direction-sensitive manner. Specifically, optical trap experiments revealed that force promotes a transition between weak and strong actin-bound states. Here, we describe the cryo-electron microscopy structure of the F-actin-bound αE-catenin actin-binding domain, which in solution forms a five-helix bundle. In the actin-bound structure, the first helix of the bundle dissociates and the remaining four helices and connecting loops rearrange to form the interface with actin. Deletion of the first helix produces strong actin binding in the absence of force, suggesting that the actin-bound structure corresponds to the strong state. Our analysis explains how mechanical force applied to αE-catenin or its homolog vinculin favors the strongly bound state, and the dependence of catch bond strength on the direction of applied force.

scholarly journals Ponsin/SH3P12: An l-Afadin– and Vinculin-binding Protein Localized at Cell–Cell and Cell–Matrix Adherens Junctions

1999 ◽  
Vol 144 (5) ◽  
pp. 1001-1018 ◽  
Author(s):  
Kenji Mandai ◽  
Hiroyuki Nakanishi ◽  
Ayako Satoh ◽  
Kenichi Takahashi ◽  
Keiko Satoh ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Binding Protein ◽  
Actin Binding ◽  
Rich Region ◽  
Sh3 Domains ◽  
Cell Matrix ◽  
Actin Binding Domain ◽  
Splicing Variants ◽  
Proline Rich Region ◽  
Cell Cell

We recently isolated a novel actin filament (F-actin)–binding protein, afadin, that has two isoforms, l- and s-afadins. l-Afadin is ubiquitously expressed and specifically localized at zonula adherens (ZA) in epithelial cells and at cell–cell adherens junction (AJ) in nonepithelial cells, whereas s-afadin is abundantly expressed in neural tissue. l-Afadin has one PDZ domain, three proline-rich regions, and one F-actin–binding domain, whereas s-afadin lacks the third proline-rich region and the F-actin–binding domain. To understand the molecular mechanism of the specific localization of l-afadin at ZA in epithelial cells and at cell–cell AJ in nonepithelial cells, we attempted here to identify an l-afadin–binding protein(s) and isolated a protein, named ponsin. Ponsin had many splicing variants and the primary structures of two of them were determined. Both the two variants had three Src homology 3 (SH3) domains and turned out to be splicing variants of SH3P12. The third proline-rich region of l-afadin bound to the region of ponsin containing the second and third SH3 domains. Ponsin was ubiquitously expressed and localized at ZA in epithelial cells, at cell–cell AJ in nonepithelial cells, and at cell–matrix AJ in both types of cells. Ponsin furthermore directly bound vinculin, an F-actin–binding protein localized at ZA in epithelial cells, at cell–cell AJ in nonepithelial cells, and at cell–matrix AJ in both types of cells. Vinculin has one proline-rich region where two proline-rich sequences are located. The proline-rich region bound to the region of ponsin containing the first and second SH3 domains. l-Afadin and vinculin bound to ponsin in a competitive manner and these three proteins hardly formed a ternary complex. These results indicate that ponsin is an l-afadin– and vinculin-binding protein localized at ZA in epithelial cells, at cell–cell AJ in nonepithelial cells, and at cell–matrix AJ in both types of cells.

scholarly journals The Cryogenic Electron Microscopy Structure of the Cell Adhesion Regulator Metavinculin Reveals an Isoform-Specific Kinked Helix in Its Cytoskeleton Binding Domain

2021 ◽  
Vol 22 (2) ◽  
pp. 645
Author(s):  
Erumbi S. Rangarajan ◽  
Tina Izard
Keyword(s):  
Electron Microscopy ◽  
Cell Adhesion ◽  
Binding Domain ◽  
Cytoskeletal Proteins ◽  
Cell Junctions ◽  
Actin Binding ◽  
Cell Matrix ◽  
Actin Binding Domain ◽  
Α Helix ◽  
Cell Cell

Vinculin and its heart-specific splice variant metavinculin are key regulators of cell adhesion processes. These membrane-bound cytoskeletal proteins regulate the cell shape by binding to several other proteins at cell–cell and cell–matrix junctions. Vinculin and metavinculin link integrin adhesion molecules to the filamentous actin network. Loss of both proteins prevents cell adhesion and cell spreading and reduces the formation of stress fibers, focal adhesions, or lamellipodia extensions. The binding of talin at cell–matrix junctions or of α-catenin at cell–cell junctions activates vinculin and metavinculin by releasing their autoinhibitory head–tail interaction. Once activated, vinculin and metavinculin bind F-actin via their five-helix bundle tail domains. Unlike vinculin, metavinculin has a 68-amino-acid insertion before the second α-helix of this five-helix F-actin–binding domain. Here, we present the full-length cryogenic electron microscopy structure of metavinculin that captures the dynamics of its individual domains and unveiled a hallmark structural feature, namely a kinked isoform-specific α-helix in its F-actin-binding domain. Our identified conformational landscape of metavinculin suggests a structural priming mechanism that is consistent with the cell adhesion functions of metavinculin in response to mechanical and cellular cues. Our findings expand our understanding of metavinculin function in the heart with implications for the etiologies of cardiomyopathies.

scholarly journals αE-catenin actin-binding domain alters actin filament conformation and regulates binding of nucleation and disassembly factors

2013 ◽  
Vol 24 (23) ◽  
pp. 3710-3720 ◽  
Author(s):  
Scott D. Hansen ◽  
Adam V. Kwiatkowski ◽  
Chung-Yueh Ouyang ◽  
HongJun Liu ◽  
Sabine Pokutta ◽  
...  
Keyword(s):  
Actin Filament ◽  
Actin Filaments ◽  
Binding Domain ◽  
Actin Binding ◽  
Filamentous Actin ◽  
Filament Structure ◽  
Actin Binding Domain ◽  
Filament Bundles ◽  
Underlying Mechanisms ◽  
Cell Cell

The actin-binding protein αE-catenin may contribute to transitions between cell migration and cell–cell adhesion that depend on remodeling the actin cytoskeleton, but the underlying mechanisms are unknown. We show that the αE-catenin actin-binding domain (ABD) binds cooperatively to individual actin filaments and that binding is accompanied by a conformational change in the actin protomer that affects filament structure. αE-catenin ABD binding limits barbed-end growth, especially in actin filament bundles. αE-catenin ABD inhibits actin filament branching by the Arp2/3 complex and severing by cofilin, both of which contact regions of the actin protomer that are structurally altered by αE-catenin ABD binding. In epithelial cells, there is little correlation between the distribution of αE-catenin and the Arp2/3 complex at developing cell–cell contacts. Our results indicate that αE-catenin binding to filamentous actin favors assembly of unbranched filament bundles that are protected from severing over more dynamic, branched filament arrays.

scholarly journals Characterization of an F-actin-binding domain in the cytoskeletal protein vinculin.

1994 ◽  
Vol 126 (5) ◽  
pp. 1231-1240 ◽  
Author(s):  
A R Menkel ◽  
M Kroemker ◽  
P Bubeck ◽  
M Ronsiek ◽  
G Nikolai ◽  
...  
Keyword(s):  
Actin Filaments ◽  
Direct Interaction ◽  
Binding Domain ◽  
Actin Binding ◽  
Filamentous Actin ◽  
Permeabilized Cells ◽  
Cell Matrix ◽  
Actin Binding Domain ◽  
Major Structural Component

Vinculin, a major structural component of vertebrate cell-cell and cell-matrix adherens junctions, has been found to interact with several other junctional components. In this report, we have identified and characterized a binding site for filamentous actin. These results included studies with gizzard vinculin, its proteolytic head and tail fragments, and recombinant proteins containing various gizzard vinculin sequences fused to the maltose binding protein (MBP) of Escherichia coli. In cosedimentation assays, only the vinculin tail sequence mediated a direct interaction with actin filaments. The binding was saturable, with a dissociation constant value in the micromolar range. Experiments with deletion clones localized the actin-binding domain to a region confined by residues 893-1016 in the 170-residue-long carboxyterminal segment, while the proline-rich hinge connecting the globular head to the rodlike tail was not required for this interaction. In fixed and permeabilized cells (cell models), as well as after microinjection, proteins containing the actin-binding domain specifically decorated stress fibers and the cortical network of fibroblasts and epithelial cells, as well as of brush border type microvilli. These results corroborated the sedimentation experiments. Our data support and extend previous work showing that vinculin binds directly to actin filaments. They are consistent with a model suggesting that in adhesive cells, the NH2-terminal head piece of vinculin directs this molecule to the focal contact sites, while its tail segment causes bundling of the actin filament ends into the characteristic spear tip-shaped structures.

scholarly journals Vinculin-dependent actin bundling regulates cell migration and traction forces

2015 ◽  
Vol 465 (3) ◽  
pp. 383-393 ◽  
Author(s):  
Karry M. Jannie ◽  
Shawn M. Ellerbroek ◽  
Dennis W. Zhou ◽  
Sophia Chen ◽  
David J. Crompton ◽  
...  
Keyword(s):  
Cell Migration ◽  
Traction Force ◽  
Force Generation ◽  
Actin Binding ◽  
Traction Forces ◽  
Cell Matrix ◽  
Cell Cell

Vinculin transduces force and orchestrates mechanical signalling at cell–cell and cell–matrix adhesions. Cells expressing a mutant vinculin deficient in actin binding and bundling display migration and traction force defects. Vinculin binding to actin is critical for cell migration and force generation.

scholarly journals Author response: Structural basis of αE-catenin–F-actin catch bond behavior

2020 ◽  
Author(s):  
Xiao-Ping Xu ◽  
Sabine Pokutta ◽  
Megan Torres ◽  
Mark F Swift ◽  
Dorit Hanein ◽  
...  
Keyword(s):  
Author Response ◽  
Structural Basis ◽  
Catch Bond ◽  
Bond Behavior

scholarly journals Structural basis of the filamin A actin-binding domain interaction with F-actin

2018 ◽  
Vol 25 (10) ◽  
pp. 918-927 ◽  
Author(s):  
Daniel V. Iwamoto ◽  
Andrew Huehn ◽  
Bertrand Simon ◽  
Clotilde Huet-Calderwood ◽  
Massimiliano Baldassarre ◽  
...  
Keyword(s):  
Binding Domain ◽  
Actin Binding ◽  
Structural Basis ◽  
Filamin A ◽  
Domain Interaction ◽  
Actin Binding Domain

Membrane-Bound Transcription Factor Site-1 Protease in PF429242 Bound State: Computational Kinetics and Dynamics of Reversible Binding

Drug Research ◽  
2019 ◽  
Vol 69 (12) ◽  
pp. 643-649 ◽  
Author(s):  
Omotuyi I. Olaposi ◽  
Nash Oyekanmi ◽  
Ayodeji A. Ojo ◽  
Gabriel O. Eniafe
Keyword(s):  
Transcription Factor ◽  
Bound States ◽  
Homology Model ◽  
Bound State ◽  
Catalytic Triad ◽  
Dynamics Simulation ◽  
Structural Basis ◽  
Viral Glycoprotein ◽  
Reversible Binding ◽  
Membrane Bound

AbstractMembrane-bound transcription factor site-1 protease (S1P) is an emerging clinical target due to its roles in lipogenesis, lysosomal biogenesis, unfolded protein response and viral glycoprotein processing. In this study, homology model of S1P was created in order to understand the structural basis for S1P inhibition by PF429242 using molecular docking, molecular dynamics simulation and in silico kinetics studies. PF429242 was docked (GlideScorePF429242=−5.20 kcal/mol) into the catalytic triad (D218, H249 and S414) and validated (R2=0.5686). The reversible binding kinetic parameter (Koff/Kon) was estimated at=7.28E-03 M with fully bound and apo-states interspersed by 3 transient ligand-bound states with unique binding signatures; water plays a major role in PF429242 dissociation from the catalytic site. Communication between key catalytic triad residues is altered in the presence of PF429242. In apo-S1P state, S414–S307/V216-D218 is the preferred route but in PF429242-bound state, S414-S417/V216-D218 is preferred. Communication between S414 and H249 is also shortened in PF429242 bound state; here, only L410 is required unlike apo-state, which requires P418, V256 and F252. Ligand binding did not alter the communication route between S414 and H249 as both recruited D244 and G220. In conclusion, PF429242 binds tightly but reversible to S1P and the details of this interaction has been presented to guide future efforts at developing novel inhibitors. Site-1-protease; PF429242; Kon/Koff; Network analysis

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