Heterogeneity of Focal Adhesions and Focal Contacts in Motile Fibroblasts

2018 ◽  
pp. 205-218 ◽  
Author(s):  
Aleena Gladkikh ◽  
Anastasia Kovaleva ◽  
Anna Tvorogova ◽  
Ivan A. Vorobjev
Keyword(s):  
Focal Adhesions ◽  
Focal Contacts

scholarly journals Cell motion as a stochastic process controlled by focal contacts dynamics

2019 ◽  
Author(s):  
Simon Lo Vecchio ◽  
Raghavan Thiagarajan ◽  
David Caballero ◽  
Vincent Vigon ◽  
Laurent Navoret ◽  
...  
Keyword(s):  
Control Cell ◽  
Focal Adhesions ◽  
Contact Dynamics ◽  
Driving Mechanism ◽  
Focal Contact ◽  
Focal Contacts ◽  
Consistent Model ◽  
Cell Motion

SUMMARYDirected cell motion is essential in physiological and pathological processes such as morphogenesis, wound healing and cancer spreading. Chemotaxis has often been proposed as the driving mechanism, even though evidence of long-range gradients is often lacking in vivo. By patterning adhesive regions in space, we control cell shape and the associated potential to move along one direction in another mode of migration coined ratchetaxis. We report that focal contacts distributions collectively dictate cell directionality, and bias is non-linearly increased by gap distance between adhesive regions. Focal contact dynamics on micro-patterns allow to integrate these phenomena in a consistent model where each focal contact can be translated into a force with known amplitude and direction, leading to quantitative predictions for cell motion in every condition. Altogether, our study shows how local and minutes timescale dynamics of focal adhesions and their distribution lead to long term cellular motion with simple geometric rules.

Cell-substrate contacts in cultured chick embryonic cells: an interference reflection study

1982 ◽  
Vol 58 (1) ◽  
pp. 165-183
Author(s):  
G.W. Ireland ◽  
C.D. Stern
Keyword(s):  
Focal Adhesions ◽  
Peripheral Zone ◽  
Embryonic Cells ◽  
Extracellular Material ◽  
Cell Contacts ◽  
Specialized Cell ◽  
Conspicuous Feature ◽  
Focal Contacts ◽  
Cell Substrate ◽  
Epithelial Sheets

Cell-substrate contacts in explants of different regions of early chick tissues were investigated using the technique of interference reflection microscopy. All the explants spread as epithelial sheets. During initial spreading a peripheral zone of 2–3 cells formed broad contacts with the substrate. In spread explants some cells in the centre made broad substrate contacts. A mat of extracellular material containing fibronectin was found under the explants. Focal contacts and focal adhesions increased in number during culture, and stress fibres were associated with them. These changes in cell contacts appeared more quickly in some tissues than in others. After 24 h, explants of hypoblast and definitive endoblast could easily be distinguished but by 7 days they were very similar. In the absence of serum, specialized cell contacts developed more quickly; in higher concentrations of serum, more slowly. Confrontations between explants were also examined. The most conspicuous feature was that cells in invading explants normally underlapped invaded cells. Invasion from above by an unspread explant could occur even if the invaded explant had formed many focal adhesions.

The behaviour of fibroblasts migrating from chick heart explants: changes in adhesion, locomotion and growth, and in the distribution of actomyosin and fibronectin

1979 ◽  
Vol 39 (1) ◽  
pp. 149-165
Author(s):  
J.R. Couchman ◽  
D.A. Rees
Keyword(s):  
Growth Rate ◽  
Focal Adhesions ◽  
High Rate ◽  
Chick Embryos ◽  
Focal Contacts ◽  
Parallel Increase ◽  
Fibronectin Expression ◽  
Chick Heart ◽  
High Degree

Fibroblasts migrating from heart explants of chick embryos at first have a high rate of locomotion but lack focal contacts or adhesions and also lack substantial actin-containing bundles. A meshwork of 7-nm filaments is present particularly in submembranous regions and is proposed to be directed towards efficient locomotion whilst maintaining a high degree of spreading. Also during the first 48 h there is little production of extracellular fibronectin and the growth rate is low. Later, these fibroblasts develop focal contacts and focal adhesions together with actomyosin bundles, with a parallel increase in fibronectin expression. We propose that progressive immobilization by the development of focal adhesions and actomyosin structures occurs to set these cells up for growth.

scholarly journals Probing mechanical principles of focal contacts in cell–matrix adhesion with a coupled stochastic–elastic modelling framework

2011 ◽  
Vol 8 (62) ◽  
pp. 1217-1232 ◽  
Author(s):  
Huajian Gao ◽  
Jin Qian ◽  
Bin Chen
Keyword(s):  
Numerical Models ◽  
Focal Adhesions ◽  
Point Of View ◽  
Matrix Adhesion ◽  
Modelling Framework ◽  
Cell Matrix ◽  
Focal Contacts ◽  
Wide Range ◽  
Sense And Respond ◽  
Cell Matrix Adhesion

Cell–matrix adhesion depends on the collective behaviours of clusters of receptor–ligand bonds called focal contacts between cell and extracellular matrix. While the behaviour of a single molecular bond is governed by statistical mechanics at the molecular scale, continuum mechanics should be valid at a larger scale. This paper presents an overview of a series of recent theoretical studies aimed at probing the basic mechanical principles of focal contacts in cell–matrix adhesion via stochastic–elastic models in which stochastic descriptions of molecular bonds and elastic descriptions of interfacial traction–separation are unified in a single modelling framework. The intention here is to illustrate these principles using simple analytical and numerical models. The aim of the discussions is to provide possible clues to the following questions: why does the size of focal adhesions (FAs) fall into a narrow range around the micrometre scale? How can cells sense and respond to substrates of varied stiffness via FAs? How do the magnitude and orientation of mechanical forces affect the binding dynamics of FAs? The effects of cluster size, cell–matrix elastic modulus, loading direction and cytoskeletal pretension on the lifetime of FA clusters have been investigated by theoretical arguments as well as Monte Carlo numerical simulations, with results showing that intermediate adhesion size, stiff substrate, cytoskeleton stiffening, low-angle pulling and moderate cytoskeletal pretension are factors that contribute to stable FAs. From a mechanistic point of view, these results provide possible explanations for a wide range of experimental observations and suggest multiple mechanisms by which cells can actively control adhesion and de-adhesion via cytoskeletal contractile machinery in response to mechanical properties of their surroundings.

scholarly journals A New Paxillin-binding Protein, PAG3/Papα/KIAA0400, Bearing an ADP-Ribosylation Factor GTPase-activating Protein Activity, Is Involved in Paxillin Recruitment to Focal Adhesions and Cell Migration

2000 ◽  
Vol 11 (4) ◽  
pp. 1315-1327 ◽  
Author(s):  
Akiko Kondo ◽  
Shigeru Hashimoto ◽  
Hajime Yano ◽  
Kuniaki Nagayama ◽  
Yuichi Mazaki ◽  
...  
Keyword(s):  
Subcellular Localization ◽  
Binding Protein ◽  
Focal Adhesions ◽  
Cell Periphery ◽  
Gtpase Activating Protein ◽  
Protein Activity ◽  
Adp Ribosylation ◽  
Focal Contacts ◽  
Adp Ribosylation Factor

Paxillin acts as an adaptor molecule in integrin signaling. Paxillin is localized to focal contacts but seems to also exist in a relatively large cytoplasmic pool. Here, we report the identification of a new paxillin-binding protein, PAG3 (paxillin-associated protein with ADP-ribosylation factor [ARF] GTPase-activating protein [GAP] activity, number 3), which is involved in regulation of the subcellular localization of paxillin. PAG3 bound to all paxillin isoforms and was induced during monocyte maturation, at which time paxillin expression is also increased and integrins are activated. PAG3 was diffusely distributed in the cytoplasm in premature monocytes but became localized at cell periphery in mature monocytes, a fraction of which then colocalized with paxillin. PAG3, on the other hand, did not accumulate at focal adhesion plaques, suggesting that PAG3 is not an integrin assembly protein. PAG3 was identical to KIAA0400/Papα, which was previously identified as a Pyk2-binding protein bearing a GAP activity toward several ARFs in vitro. Mammalian ARFs fall into three classes, and we showed that all classes could affect subcellular localization of paxillin. We also examined possible interaction of PAG3 with ARFs and showed evidence that at least one of them, ARF6, seems to be an intracellular substrate for GAP activity of PAG3. Moreover, overexpression of PAG3, but not its GAP-inactive mutant, inhibited paxillin recruitment to focal contacts and hampered cell migratory activities, whereas cell adhesion activities were almost unaffected. Therefore, our results demonstrate that paxillin recruitment to focal adhesions is not mediated by simple cytoplasmic diffusion; rather, PAG3 appears to be involved in this process, possibly through its GAP activity toward ARF proteins. Our result thus delineates a new aspect of regulation of cell migratory activities.

scholarly journals The mechanisms and dynamics of αvβ3 integrin clustering in living cells

2005 ◽  
Vol 171 (2) ◽  
pp. 383-392 ◽  
Author(s):  
Caroline Cluzel ◽  
Frédéric Saltel ◽  
Jost Lussi ◽  
Frédérique Paulhe ◽  
Beat A. Imhof ◽  
...  
Keyword(s):  
Ternary Complex ◽  
De Novo ◽  
Focal Adhesions ◽  
Αvβ3 Integrin ◽  
Open Conformation ◽  
Focal Contacts ◽  
Integrin Clustering ◽  
Αvβ3 Integrins ◽  
Mutational Activation ◽  
Integrin Family

During cell migration, the physical link between the extracellular substrate and the actin cytoskeleton mediated by receptors of the integrin family is constantly modified. We analyzed the mechanisms that regulate the clustering and incorporation of activated αvβ3 integrins into focal adhesions. Manganese (Mn2+) or mutational activation of integrins induced the formation of de novo F-actin–independent integrin clusters. These clusters recruited talin, but not other focal adhesion adapters, and overexpression of the integrin-binding head domain of talin increased clustering. Integrin clustering required immobilized ligand and was prevented by the sequestration of phosphoinositole-4,5-bisphosphate (PI(4,5)P2). Fluorescence recovery after photobleaching analysis of Mn2+-induced integrin clusters revealed increased integrin turnover compared with mature focal contacts, whereas stabilization of the open conformation of the integrin ectodomain by mutagenesis reduced integrin turnover in focal contacts. Thus, integrin clustering requires the formation of the ternary complex consisting of activated integrins, immobilized ligands, talin, and PI(4,5)P2. The dynamic remodeling of this ternary complex controls cell motility.

scholarly journals Ultrastructural localization of plasma membrane-associated urokinase-type plasminogen activator at focal contacts.

1988 ◽  
Vol 106 (1) ◽  
pp. 87-95 ◽  
Author(s):  
J Pöllänen ◽  
K Hedman ◽  
L S Nielsen ◽  
K Danø ◽  
A Vaheri
Keyword(s):  
Plasma Membrane ◽  
Plasminogen Activator ◽  
Focal Adhesions ◽  
Lung Fibroblasts ◽  
Migration And Invasion ◽  
Cell Contact ◽  
Focal Contacts ◽  
Type Plasminogen Activator ◽  
Urokinase Type Plasminogen Activator ◽  
Adhesion Plaques

We have recently shown that urokinase-type plasminogen activator (u-PA) and plasminogen activator inhibitor type 1 are both found extracellularly beneath cultured human skin fibroblasts and HT-1080 sarcoma cells, but in distinct localizations. Here, the ultrastructural distribution of u-PA was studied using immunoferritin electron microscopy. In HT-1080 cells, u-PA on the extracellular aspect of the plasma membrane was detected at sites of direct contact of the cell with the growth substratum beneath all parts of the ventral cell surface. The ferritin-labeled adhesion plaques, which were enriched in submembraneous microfilaments, were frequently seen at the leading lamellae of the cells as well as in lamellipodia and microspikes. Besides the cell-substratum adhesion plaques, ferritin label was detected at cell-cell contact sites. Double-label immunofluorescence showed a striking colocalization of u-PA and vinculin in both HT-1080 cells and WI-38 lung fibroblasts, which is consistent with u-PA being a focal contact component. The u-PA-containing focal contacts of WI-38 cells had no direct codistribution with fibronectin fibrils. In WI-38 cells made stationary by cultivation in a medium containing 0.5% FCS, vinculin plaques became highly elongated and more centrally located, whereas u-PA immunolabel disappeared from such focal adhesions. These findings show that plasma membrane-associated u-PA is an intrinsic component of focal contacts, where, we propose, it enables directional proteolysis for cell migration and invasion.

Molecular diversity of cell-matrix adhesions

1999 ◽  
Vol 112 (11) ◽  
pp. 1655-1669 ◽  
Author(s):  
E. Zamir ◽  
B.Z. Katz ◽  
S. Aota ◽  
K.M. Yamada ◽  
B. Geiger ◽  
...  
Keyword(s):  
Kinase Inhibitor ◽  
Focal Adhesions ◽  
Automatic Identification ◽  
Molecular Heterogeneity ◽  
Cell Matrix ◽  
Adhesion Sites ◽  
Actomyosin Contractility ◽  
Focal Contacts ◽  
Quantitative Immunofluorescence ◽  
Ratio Imaging

In this study we have examined for molecular heterogeneity of cell-matrix adhesions and the involvement of actomyosin contractility in the selective recruitment of different plaque proteins. For this purpose, we have developed a novel microscopic approach for molecular morphometry, based on automatic identification of matrix adhesions, followed by quantitative immunofluorescence and morphometric analysis. Particularly informative was fluorescence ratio imaging, comparing the local labeling intensities of different plaque molecules, including vinculin, paxillin, tensin and phosphotyrosine-containing proteins. Ratio imaging revealed considerable molecular heterogeneity between and within adhesion sites. Most striking were the differences between focal contacts, which are vinculin- and paxillin-rich and contain high levels of phosphotyrosine, and fibrillar adhesions, which are tensin-rich and contain little or no phosphotyrosine. Ratio imaging also revealed considerable variability in the molecular substructure of individual focal contacts, pointing to a non-uniform distribution of phosphotyrosine and the different plaque constituents. Studying the quantitative relationships between the various components of the submembrane plaque indicated that the levels of vinculin, paxillin and phosphotyrosine in adhesion sites are positively correlated with each other and negatively correlated with the levels of tensin. Tyrosine phosphorylation of focal contacts was highly sensitive to cellular contractility, and was diminished within 5 minutes after treatment with the kinase inhibitor H-7, an inhibitor of actomyosin contractility. This was followed by the loss of paxillin and vinculin from the focal adhesions. Tensin-rich fibrillar adhesions were relatively insensitive to H-7 treatment. These findings suggest a role for contractility in the generation of matrix adhesion diversity.

Syndecan-4 core protein is sufficient for the assembly of focal adhesions and actin stress fibers

1999 ◽  
Vol 112 (20) ◽  
pp. 3433-3441 ◽  
Author(s):  
F. Echtermeyer ◽  
P.C. Baciu ◽  
S. Saoncella ◽  
Y. Ge ◽  
P.F. Goetinck
Keyword(s):  
Core Protein ◽  
Chinese Hamster Ovary Cells ◽  
Chinese Hamster ◽  
Focal Adhesions ◽  
Stress Fibers ◽  
Actin Stress Fiber ◽  
The Core ◽  
Focal Contacts ◽  
Mutant Cells ◽  
Actin Stress Fibers

The formation of focal adhesions and actin stress fibers on fibronectin is dependent on signaling through (β)1 integrins and the heparan sulfate proteoglycan syndecan-4, and we have analyzed the requirement of the glycosaminoglycan chains of syndecan-4 during these events. Chinese hamster ovary cells with mutations in key enzymes of the glycanation process do not synthesize glycosaminoglycan chains and are unable to assemble actin stress fibers and focal contacts when cultured on fibronectin. Transfection of the mutant cells with a cDNA that encodes the core protein of chicken syndecan-4 leads to the production of unglycanated core protein. The overexpression of syndecan-4 core protein in these mutant cells increases cell spreading and is sufficient for these cells to assemble actin stress fibers and focal adhesions similar to wild-type cells seeded on fibronectin and vitronectin matrices. Syndecan-4 core protein colocalizes to focal contacts in mutant cells that have been transfected with the syndecan-4 core protein cDNA. These data indicate an essential role for the core protein of syndecan-4 in the generation of signals leading to actin stress fiber and focal contact assembly.

scholarly journals pp60c-src and related tyrosine kinases: a role in the assembly and reorganization of matrix adhesions

2001 ◽  
Vol 114 (12) ◽  
pp. 2279-2289 ◽  
Author(s):  
Tova Volberg ◽  
Lewis Romer ◽  
Eli Zamir ◽  
Benjamin Geiger
Keyword(s):  
Tyrosine Kinases ◽  
Mutant Cell ◽  
Focal Adhesions ◽  
Src Family Kinases ◽  
Wild Type ◽  
Altered Expression ◽  
Cell Matrix ◽  
Focal Contacts ◽  
Cell Matrix Adhesion

Activation of tyrosine kinases during integrin-mediated cell-matrix adhesion is involved both in the regulation of focal contact assembly and in the initiation of signaling processes at the cell-matrix adhesive interface. In order to determine the role of pp60c-src and related kinases in these processes, we have compared the dynamic reorganization of phosphotyrosine, vinculin, focal adhesion kinase and tensin in cells with altered expression of Src-family kinases. Both null cells for pp60c-src and triple knockout cells for pp60c-src, pp59fyn, and pp62c-yes exhibited decreased phosphotyrosine levels in focal contacts when compared with wild-type cells. pp60c-src-null cells also exhibited faster assembly of cell-matrix adhesions and a more exuberant recruitment of FAK to these sites. Tensin, which normally segregates into fibrillar adhesions was localized in large focal contacts in the two mutant cell lines, suggesting involvement of pp60c-src in the segregation of focal contacts and fibrillar adhesions. Moreover, treatment of wild-type cells with tyrphostin AG1007, which inhibits both pp60c-src and FAK activity, induced accumulation of tensin in peripheral focal adhesions. These findings demonstrate that Src family kinases, and pp60c-src in particular, have a central role in regulating protein dynamics at cell-matrix interfaces, both during early stages of interaction and in mature focal contacts.

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