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 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 Substrate rigidity modulates traction forces and stoichiometry of cell matrix adhesions.

2021 ◽  
Author(s):  
Hayri E Balcioglu ◽  
Rolf Harkes ◽  
Erik Danen ◽  
Thomas Schmidt
Keyword(s):  
Intracellular Signaling ◽  
Traction Force ◽  
Effective Stiffness ◽  
Traction Forces ◽  
Superresolution Microscopy ◽  
Cell Matrix ◽  
Force Increase ◽  
Distinct Cell ◽  
Associated Proteins ◽  
Relative Change

In cell matrix adhesions, integrin receptors and associated proteins provide a dynamic coupling of the extracellular matrix (ECM) to the cytoskeleton. This allows bidirectional transmission of forces between the ECM and the cytoskeleton, which tunes intracellular signaling cascades that control survival, proliferation, differentiation, and motility. The quantitative relationships between recruitment of distinct cell matrix adhesion proteins and local cellular traction forces are not known. Here, we applied quantitative superresolution microscopy to cell matrix adhesions formed on fibronectin-stamped elastomeric pillars and developed an approach to relate the number of talin, vinculin, paxillin, and focal adhesion kinase (FAK) molecules to the local cellular traction force. We find that FAK recruitment does not show an association with traction-force application whereas a ~60 pN force increase is associated with the recruitment of one talin, two vinculin, and two paxillin molecules on a substrate of effective stiffness of 47 kPa. On a substrate with a four-fold lower effective stiffness the stoichiometry of talin:vinculin:paxillin changes to 2:12:6 for the same ~60 pN traction force. The relative change in force-related vinculin recruitment indicates a stiffness-dependent switch in vinculin function in cell matrix adhesions. Our results reveal a substrate-stiffness-dependent modulation of the relation between cellular traction-force and the molecular stoichiometry of cell-matrix adhesions.

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 Contact inhibition of locomotion determines cell–cell and cell–substrate forces in tissues

2016 ◽  
Vol 113 (10) ◽  
pp. 2660-2665 ◽  
Author(s):  
Juliane Zimmermann ◽  
Brian A. Camley ◽  
Wouter-Jan Rappel ◽  
Herbert Levine
Keyword(s):  
Cell Density ◽  
Mdck Cells ◽  
Traction Force ◽  
Contact Inhibition ◽  
Traction Forces ◽  
Cell Substrate ◽  
Small Clusters ◽  
Cellular Forces ◽  
Darby Canine Kidney ◽  
Cell Cell

Cells organized in tissues exert forces on their neighbors and their environment. Those cellular forces determine tissue homeostasis as well as reorganization during embryonic development and wound healing. To understand how cellular forces are generated and how they can influence the tissue state, we develop a particle-based simulation model for adhesive cell clusters and monolayers. Cells are contractile, exert forces on their substrate and on each other, and interact through contact inhibition of locomotion (CIL), meaning that cell–cell contacts suppress force transduction to the substrate and propulsion forces align away from neighbors. Our model captures the traction force patterns of small clusters of nonmotile cells and larger sheets of motile Madin–Darby canine kidney (MDCK) cells. In agreement with observations in a spreading MDCK colony, the cell density in the center increases as cells divide and the tissue grows. A feedback between cell density, CIL, and cell–cell adhesion gives rise to a linear relationship between cell density and intercellular tensile stress and forces the tissue into a nonmotile state characterized by a broad distribution of traction forces. Our model also captures the experimentally observed tissue flow around circular obstacles, and CIL accounts for traction forces at the edge.

scholarly journals The Interplay Between Cell-Cell and Cell-Matrix Forces Regulates Cell Migration Dynamics

2019 ◽  
Vol 117 (10) ◽  
pp. 1795-1804 ◽  
Author(s):  
Apratim Bajpai ◽  
Jie Tong ◽  
Weiyi Qian ◽  
Yansong Peng ◽  
Weiqiang Chen
Keyword(s):  
Cell Migration ◽  
Cell Matrix ◽  
Migration Dynamics ◽  
Cell Cell

scholarly journals RAP1-RAC1 Signaling Has an Important Role in Adhesion and Migration in HNSCC

2020 ◽  
Vol 99 (8) ◽  
pp. 959-968 ◽  
Author(s):  
M. Liu ◽  
R. Banerjee ◽  
C. Rossa ◽  
N.J. D’Silva
Keyword(s):  
Cell Migration ◽  
Cell Adhesion ◽  
Matrix Adhesion ◽  
Cell Matrix ◽  
And Migration ◽  
Rap1 Activation ◽  
Cell Matrix Adhesion ◽  
Α5β1 Integrin ◽  
Hnscc Cell ◽  
Cell Cell

Cell-cell adhesion is a key mechanism to control tissue integrity and migration. In head and neck squamous cell carcinoma (HNSCC), cell migration facilitates distant metastases and is correlated with poor prognosis. RAP1, a ras-like protein, has an important role in the progression of HNSCC. RAC1 is an integrin-linked, ras-like protein that promotes cell migration. Here we show that loss of cell-cell adhesion is correlated with inactivation of RAP1 confirmed by 2 different biochemical approaches. RAP1 activation is required for cell-matrix adhesion confirmed by adhesion to fibronectin-coated plates with cells that have biochemically activated RAP1. This effect is reversed when RAP1 is inactivated. In addition, RAP1GTP-mediated adhesion is only facilitated through α5β1 integrin complex and is not a function of either α5 or β1 integrin alone. Moreover, the inside-out signaling of RAP1 activation is coordinated with RAC1 activation. These findings show that RAP1 has a prominent role in cell-matrix adhesion via extracellular matrix molecule fibronectin-induced α5β1 integrin and supports a critical role for the RAP1/RAC1 signaling axis in HNSCC cell migration.

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 Regulation of SMC traction forces in human aortic thoracic aneurysms

2021 ◽  
Author(s):  
Claudie Petit ◽  
Ali-Akbar Karkhaneh Yousefi ◽  
Olfa Ben Moussa ◽  
Jean-Baptiste Michel ◽  
Alain Guignandon ◽  
...  
Keyword(s):  
Traction Force ◽  
Aortic Aneurysms ◽  
Force Generation ◽  
Dynamic Force ◽  
Generation Process ◽  
Traction Forces ◽  
Thoracic Aortic Aneurysms ◽  
And Gender ◽  
Thoracic Aneurysms ◽  
Future Work

AbstractSmooth muscle cells (SMCs) usually express a contractile phenotype in the healthy aorta. However, aortic SMCs have the ability to undergo profound changes in phenotype in response to changes in their extracellular environment, as occurs in ascending thoracic aortic aneurysms (ATAA). Accordingly, there is a pressing need to quantify the mechanobiological effects of these changes at single cell level. To address this need, we applied Traction Force Microscopy (TFM) on 759 cells coming from three primary healthy (AoPrim) human SMC lineages and three primary aneurysmal (AnevPrim) human SMC lineages, from age and gender matched donors. We measured the basal traction forces applied by each of these cells onto compliant hydrogels of different stiffness (4, 8, 12, 25 kPa). Although the range of force generation by SMCs suggested some heterogeneity, we observed that: 1. the traction forces were significantly larger on substrates of larger stiffness; 2. traction forces in AnevPrim were significantly higher than in AoPrim cells. We modelled computationally the dynamic force generation process in SMCs using the motor-clutch model and found that it accounts well for the stiffness-dependent traction forces. The existence of larger traction forces in the AnevPrim SMCs were related to the larger size of cells in these lineages. We conclude that phenotype changes occurring in ATAA, which were previously known to reduce the expression of elongated and contractile SMCs (rendering SMCs less responsive to vasoactive agents), tend also to induce stronger SMCs. Future work aims at understanding the causes of this alteration process in aortic aneurysms.

scholarly journals Nexilin: A Novel Actin Filament-binding Protein Localized at Cell–Matrix Adherens Junction

1998 ◽  
Vol 143 (5) ◽  
pp. 1227-1238 ◽  
Author(s):  
Toshihisa Ohtsuka ◽  
Hiroyuki Nakanishi ◽  
Wataru Ikeda ◽  
Ayako Satoh ◽  
Yumiko Momose ◽  
...  
Keyword(s):  
Molecular Weight ◽  
Rat Brain ◽  
Actin Filament ◽  
Binding Protein ◽  
Adherens Junction ◽  
Actin Binding ◽  
Calculated Molecular Weight ◽  
Cell Matrix ◽  
Focal Contacts ◽  
Cell Cell

We isolated two novel actin filament (F-actin)–binding proteins from rat brain and rat 3Y1 fibroblast. They were splicing variants, and we named brain big one b-nexilin and fibroblast small one s-nexilin. b-Nexilin purified from rat brain was a protein of 656 amino acids (aa) with a calculated molecular weight of 78,392, whereas s-nexilin, encoded by the cDNA isolated from rat 3Y1 cells by the reverse transcriptase-PCR method, was a protein of 606 aa with a calculated molecular weight of 71,942. b-Nexilin had two F-actin– binding domains (ABDs) at the NH2-terminal and middle regions, whereas s-nexilin had one ABD at the middle region because 64 aa residues were deleted and 14 aa residues were inserted in the first NH2-terminal ABD of b-nexilin, and thereby the first ABD lost its activity. b- and s-nexilins bound along the sides of F-actin, but only b-nexilin showed F-actin cross-linking activity. b-Nexilin was mainly expressed in brain and testis, whereas s-nexilin was mainly expressed in testis, spleen, and fibroblasts, such as rat 3Y1 and mouse Swiss 3T3 cells, but neither b- nor s-nexilin was detected in liver, kidney, or cultured epithelial cells. An immunofluorescence microscopic study revealed that s-nexilin was colocalized with vinculin, talin, and paxillin at cell– matrix adherens junction (AJ) and focal contacts, but not at cell–cell AJ, in 3Y1 cells. Overexpressed b- and s-nexilins were localized at focal contacts but not at cell–cell AJ. These results indicate that nexilin is a novel F-actin–binding protein localized at cell–matrix AJ.

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