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Biophysical Journal Dec 2023Transmission of cell-generated (i.e., endogenous) tension at cell-cell contacts is crucial for tissue shape changes during morphogenesis and adult tissue repair in...
Transmission of cell-generated (i.e., endogenous) tension at cell-cell contacts is crucial for tissue shape changes during morphogenesis and adult tissue repair in tissues such as epithelia. E-cadherin-based adhesions at cell-cell contacts are the primary means by which endogenous tension is transmitted between cells. The E-cadherin-β-catenin-α-catenin complex mechanically couples to the actin cytoskeleton (and thereby the cell's contractile machinery) both directly and indirectly. However, the key adhesion constituents required for substantial endogenous force transmission at these adhesions in cell-cell contacts are unclear. Due to the role of α-catenin as a mechanotransducer that recruits vinculin at cell-cell contacts, we expected α-catenin to be essential for sustaining normal levels of force transmission. Instead, using the traction force imbalance method to determine the inter-cellular force at a single cell-cell contact between cell pairs, we found that it is vinculin that is essential for sustaining normal levels of endogenous force transmission, with absence of vinculin decreasing the inter-cellular tension by over 50%. Our results constrain the potential mechanical pathways of force transmission at cell-cell contacts and suggest that vinculin can transmit forces at E-cadherin adhesions independent of α-catenin, possibly through β-catenin. Furthermore, we tested the ability of lateral cell-cell contacts to withstand external stretch and found that both vinculin and α-catenin are essential to maintain cell-cell contact stability under external forces.
Topics: alpha Catenin; Vinculin; beta Catenin; Cadherins; Cell Adhesion; Actins
PubMed: 38350000
DOI: 10.1016/j.bpj.2023.10.029 -
BioEssays : News and Reviews in... Sep 1998Vinculin and alpha-catenin are two functionally related proteins of adherens junctions, structures in which cells make contacts to neighboring cells or to the... (Comparative Study)
Comparative Study Review
Vinculin and alpha-catenin are two functionally related proteins of adherens junctions, structures in which cells make contacts to neighboring cells or to the extracellular matrix. At these sites, the actin cytoskeleton of animal cells is anchored to the plasma membrane. Junction assembly and disassembly are coordinated in processes as different as mitosis, cell movement and tissue formation. Since adherens junctions are assembled from a large number of proteins, these molecules have to be coordinately activated and spatially regulated. Vinculin and alpha-catenin have been characterized as tumor suppressors, suggesting that they have a regulatory function in addition to their structural role. Several possible modes of vinculin and alpha-catenin regulation are discussed here, as the published data favor the concept that no single model fully explains the complexity of adherens junctions. Most probably, cells select from a variety of possibilities to solve the problem of making specific contacts.
Topics: Actins; Amino Acid Sequence; Animals; Cell Adhesion; Cell Membrane; Cell Transformation, Neoplastic; Cells, Cultured; Cytoskeletal Proteins; Cytoskeleton; Genes, Tumor Suppressor; Intercellular Junctions; Ligands; Models, Biological; Molecular Sequence Data; Protein Structure, Secondary; Structure-Activity Relationship; Vinculin; alpha Catenin
PubMed: 9819562
DOI: 10.1002/(SICI)1521-1878(199809)20:9<733::AID-BIES6>3.0.CO;2-H -
Fish & Shellfish Immunology Apr 2023Myeloid differentiation factor 88 (MyD88) is the canonical adaptor for inflammatory signaling pathways downstream from members of the Toll-like receptor (TLR) and...
Myeloid differentiation factor 88 (MyD88) is the canonical adaptor for inflammatory signaling pathways downstream from members of the Toll-like receptor (TLR) and interleukin-1 (IL-1) receptor families, which activates the NF-κB signaling pathway and regulates immune and inflammatory responses. In this study, we found that Vinculin B (Vclb) is an inhibitor in the NF-κB signaling pathway, and its inhibitory effect was enhanced by LPS induction. Furthermore, Vclb inhibits NF-κB activation by targeting MyD88, thereby suppressing the production of inflammatory cytokines. Mechanistically, Vclb inhibits the NF-κB signaling pathway by targeting MyD88 ubiquitin-proteasome pathway. In summary, our study reveals that Vclb inhibits NF-κB signaling activation and mediates innate immunity in teleosts via the ubiquitin-proteasome pathway of MyD88.
Topics: Animals; NF-kappa B; Myeloid Differentiation Factor 88; Vinculin; Proteasome Endopeptidase Complex; Signal Transduction; Perciformes; Ubiquitins
PubMed: 36931481
DOI: 10.1016/j.fsi.2023.108683 -
Biophysical Journal Apr 2018Cell migration is a complex process, requiring coordination of many subcellular processes including membrane protrusion, adhesion, and contractility. For efficient cell...
Cell migration is a complex process, requiring coordination of many subcellular processes including membrane protrusion, adhesion, and contractility. For efficient cell migration, cells must concurrently control both transmission of large forces through adhesion structures and translocation of the cell body via adhesion turnover. Although mechanical regulation of protein dynamics has been proposed to play a major role in force transmission during cell migration, the key proteins and their exact roles are not completely understood. Vinculin is an adhesion protein that mediates force-sensitive processes, such as adhesion assembly under cytoskeletal load. Here, we elucidate the mechanical regulation of vinculin dynamics. Specifically, we paired measurements of vinculin loads using a Förster resonance energy transfer-based tension sensor and vinculin dynamics using fluorescence recovery after photobleaching to measure force-sensitive protein dynamics in living cells. We find that vinculin adopts a variety of mechanical states at adhesions, and the relationship between vinculin load and vinculin dynamics can be altered by the inhibition of vinculin binding to talin or actin or reduction of cytoskeletal contractility. Furthermore, the force-stabilized state of vinculin required for the stabilization of membrane protrusions is unnecessary for random migration, but is required for directional migration along a substrate-bound cue. These data show that the force-sensitive dynamics of vinculin impact force transmission and enable the mechanical integration of subcellular processes. These results suggest that the regulation of force-sensitive protein dynamics may have an underappreciated role in many cellular processes.
Topics: Actomyosin; Animals; Biomechanical Phenomena; Cell Line; Cell Movement; Cell Survival; Focal Adhesions; Mechanical Phenomena; Mice; Talin; Vinculin; rho-Associated Kinases
PubMed: 29642037
DOI: 10.1016/j.bpj.2018.02.019 -
Science (New York, N.Y.) Aug 2017Vinculin is an actin-binding protein thought to reinforce cell-cell and cell-matrix adhesions. However, how mechanical load affects the vinculin-F-actin bond is unclear....
Vinculin is an actin-binding protein thought to reinforce cell-cell and cell-matrix adhesions. However, how mechanical load affects the vinculin-F-actin bond is unclear. Using a single-molecule optical trap assay, we found that vinculin forms a force-dependent catch bond with F-actin through its tail domain, but with lifetimes that depend strongly on the direction of the applied force. Force toward the pointed (-) end of the actin filament resulted in a bond that was maximally stable at 8 piconewtons, with a mean lifetime (12 seconds) 10 times as long as the mean lifetime when force was applied toward the barbed (+) end. A computational model of lamellipodial actin dynamics suggests that the directionality of the vinculin-F-actin bond could establish long-range order in the actin cytoskeleton. The directional and force-stabilized binding of vinculin to F-actin may be a mechanism by which adhesion complexes maintain front-rear asymmetry in migrating cells.
Topics: Actin Cytoskeleton; Actins; Cell Movement; Computer Simulation; Models, Chemical; Optical Tweezers; Protein Binding; Pseudopodia; Single Molecule Imaging; Vinculin
PubMed: 28818948
DOI: 10.1126/science.aan2556 -
Developmental Dynamics : An Official... Jul 2023Drosophila Singed (mammalian Fascin) is an actin-binding protein that is known mainly for bundling parallel actin filaments. Among many functions of Singed, it is...
INTRODUCTION
Drosophila Singed (mammalian Fascin) is an actin-binding protein that is known mainly for bundling parallel actin filaments. Among many functions of Singed, it is required for cell motility for both Drosophila and mammalian systems. Increased Fascin-1 levels positively correlate with greater metastasis and poor prognosis in human cancer. Border cell cluster, forms and migrates during Drosophila egg chamber development, shows higher expression of Singed compared with other follicle cells. Interestingly, loss of singed in border cells does not lead to any effect other than delay.
RESULT
In this work, we have screened many actin-binding proteins in search of functional redundancy with Singed for border cell migration. We have found that Vinculin works with Singed to regulate border cell migration, albeit mildly. Although Vinculin is known for anchoring F-actin to the membrane, knockdown of both singed and vinculin leads to a reduced level of F-actin and changes in protrusion characteristics in border cells. We have also observed that they may act together to control microvilli length of brush border membrane vesicles and the shape of egg chambers in Drosophila.
CONCLUSIONS
We may conclude that singed and vinculin work together to control F-actin and these interactions are consistent across multiple platforms.
Topics: Animals; Actin Cytoskeleton; Actins; Cell Movement; Drosophila; Vinculin
PubMed: 36912821
DOI: 10.1002/dvdy.585 -
Biochemical Society Transactions Apr 2008Integrins are alphabeta heterodimeric receptors that mediate attachment of cells to the extracellular matrix and therefore play important roles in cell adhesion,... (Review)
Review
Integrins are alphabeta heterodimeric receptors that mediate attachment of cells to the extracellular matrix and therefore play important roles in cell adhesion, migration, proliferation and survival. Among the cytoskeletal proteins that interact directly with the beta-chain cytoplasmic domain, talin has emerged as playing a critical role in integrin activation and linkage to the actin cytoskeleton. Talin (2541 amino acids) is an elongated (60 nm) flexible antiparallel dimer, with a small globular head connected to an extended rod. The talin head contains a FERM (4.1/ezrin/radixin/moesin) domain (residues 86-400) with binding sites for several beta integrin cytodomains and the talin rod contains a second lower-affinity integrin-binding site, a highly conserved C-terminal actin-binding site and also several binding sites for vinculin. We have determined previously the crystal structures of two domains from the talin rod, spanning residues 482-789. Talin-(482-655), which contains a VBS (vinculin-binding site), folds into a five-helix bundle whereas talin-(656-789) is a four-helix bundle. We have also reported the crystal structure of the N-terminal vinculin head domain in complex with an activated form of talin. In the present paper, we consider how binding sites buried within the folded helical bundles of talin and alpha-actinin form interactions with vinculin.
Topics: Actinin; Cell Communication; Cytoskeleton; Humans; Integrins; Models, Molecular; Protein Conformation; Protein Structure, Tertiary; Talin; Vinculin
PubMed: 18363566
DOI: 10.1042/BST0360235 -
Biophysical Journal Mar 2020Vinculin is a universal adaptor protein that transiently reinforces the mechanical stability of adhesion complexes. It stabilizes mechanical connections that cells...
Vinculin is a universal adaptor protein that transiently reinforces the mechanical stability of adhesion complexes. It stabilizes mechanical connections that cells establish between the actomyosin cytoskeleton and the extracellular matrix via integrins or to neighboring cells via cadherins, yet little is known regarding its mechanical design. Vinculin binding sites (VBSs) from different nonhomologous actin-binding proteins use conserved helical motifs to associate with the vinculin head domain. We studied the mechanical stability of such complexes by pulling VBS peptides derived from talin, α-actinin, and Shigella IpaA out of the vinculin head domain. Experimental data from atomic force microscopy single-molecule force spectroscopy and steered molecular dynamics (SMD) simulations both revealed greater mechanical stability of the complex for shear-like than for zipper-like pulling configurations. This suggests that reinforcement occurs along preferential force directions, thus stabilizing those cytoskeletal filament architectures that result in shear-like pulling geometries. Large force-induced conformational changes in the vinculin head domain, as well as protein-specific fine-tuning of the VBS sequence, including sequence inversion, allow for an even more nuanced force response.
Topics: Binding Sites; Models, Molecular; Protein Binding; Talin; Vinculin
PubMed: 32109366
DOI: 10.1016/j.bpj.2019.12.042 -
Journal of Cellular Biochemistry Jun 2014Vinculin is a talin-binding protein that promotes integrin-mediated cell adhesion, but the mechanisms are not understood. Because talin is a direct activator of...
Vinculin is a talin-binding protein that promotes integrin-mediated cell adhesion, but the mechanisms are not understood. Because talin is a direct activator of integrins, we asked whether and how vinculin regulates the formation of integrin: talin complexes. We report that VD1 (aa 1-258) and its talin-binding mutant, VD1A50I, bind directly and equally to several β integrin cytoplasmic tails (βCT). Results from competition assays show that VD1, but not VD1A50I, inhibits the interaction of talin (Tn) and talin rod (TnR), but not talin head (TnH) with β3CT. The inhibition observed could be the result of VD1 binding to one or more of the 11 vinculin binding sites (VBSs) in the TnR domain. Our studies demonstrate that VD1 binding to amino acids 482-911, a VBS rich region, in TnR perturbs the interaction of rod with β3CT. The integrin activation assays done using CHOA5 cells show that activated vinculin enhances αIIbβ3 integrin activation and that the effect is dependent on talin. The TnR domain however shows no integrin activation unlike TnH that shows enhanced integrin activation. The overall results indicate that activated vinculin promotes talin-mediated integrin activation by binding to accessible VBSs in TnR and thus displacing the TnR from the β3 subunit. The study presented, defines a novel direct interaction of VD1 with β3CT and provides an attractive explanation for vinculin's ability to potentiate integrin-mediated cell adhesion through directly binding to both TnR and the integrin cytoplasmic tail.
Topics: Animals; Binding Sites; CHO Cells; Cricetinae; Cricetulus; Flow Cytometry; HEK293 Cells; Humans; Immunoblotting; Integrin beta3; Kinetics; Luminescent Proteins; Models, Biological; Models, Molecular; Mutation; Protein Binding; Protein Isoforms; Protein Structure, Tertiary; Talin; Vinculin
PubMed: 24446374
DOI: 10.1002/jcb.24772 -
PloS One 2017The formation of multicellular tissues during development is governed by mechanical forces that drive cell shape and tissue architecture. Protein complexes at sites of...
The formation of multicellular tissues during development is governed by mechanical forces that drive cell shape and tissue architecture. Protein complexes at sites of adhesion to the extracellular matrix (ECM) and cell neighbors, not only transmit these mechanical forces, but also allow cells to respond to changes in force by inducing biochemical feedback pathways. Such force-induced signaling processes are termed mechanotransduction. Vinculin is a central protein in mechanotransduction that in both integrin-mediated cell-ECM and cadherin-mediated cell-cell adhesions mediates force-induced cytoskeletal remodeling and adhesion strengthening. Vinculin was found to be important for the integrity and remodeling of epithelial tissues in cell culture models and could therefore be expected to be of broad importance in epithelial morphogenesis in vivo. Besides a function in mouse heart development, however, the importance of vinculin in morphogenesis of other vertebrate tissues has remained unclear. To investigate this further, we knocked out vinculin functioning in zebrafish, which contain two fully functional isoforms designated as vinculin A and vinculin B that both show high sequence conservation with higher vertebrates. Using TALEN and CRISPR-Cas gene editing technology we generated vinculin-deficient zebrafish. While single vinculin A mutants are viable and able to reproduce, additional loss of zygotic vinculin B was lethal after embryonic stages. Remarkably, vinculin-deficient embryos do not show major developmental defects, apart from mild pericardial edemas. These results lead to the conclusion that vinculin is not of broad importance for the development and morphogenesis of zebrafish tissues.
Topics: Animals; Cell Adhesion; Dogs; Embryonic Development; Extracellular Matrix; Gene Expression Regulation, Developmental; Gene Knockout Techniques; HEK293 Cells; Humans; Madin Darby Canine Kidney Cells; Mechanotransduction, Cellular; Vinculin; Zebrafish; Zebrafish Proteins
PubMed: 28767718
DOI: 10.1371/journal.pone.0182278