Substrate Stiffness as a Regulator in Catalyzing Integrin-Mediated Cell Adhesion Nucleation: A Mechanochemical Model

2013 ◽  
Vol 459 ◽  
pp. 555-559
Author(s):  
Xiao Ling Peng
Keyword(s):  
Cell Adhesion ◽  
Energy Input ◽  
Experimental Studies ◽  
Substrate Stiffness ◽  
Dependent Manner ◽  
Integrin Activation ◽  
A Value ◽  
Integrin Clustering ◽  
Mechanochemical Model

We provide here a simplified mechanochemical model to describe the role of substrate stiffness in mediating the chemical reactions between integrins on cell membrane and ligands immobilized on the substrate. By taking into account the energy input for integrin activation on a compliant substrate, Our simulation shows that integrin activation and the downstream integrin clustering can be regulated by substrate stiffness in a value-dependent manner, which is consistent with previous experimental studies.

Integrin mechanotransduction stimulates caveolin-1 phosphorylation and recruitment of Csk to mediate actin reorganization

2005 ◽  
Vol 288 (2) ◽  
pp. H936-H945 ◽  
Author(s):  
C. Radel ◽  
V. Rizzo
Keyword(s):  
Shear Stress ◽  
Dependent Manner ◽  
Laminar Shear Stress ◽  
Integrin Activation ◽  
Actin Reorganization ◽  
Aortic Endothelial Cells ◽  
Caveolin 1 ◽  
Domain Peptide ◽  
Mlc Phosphorylation

To identify the role of caveolin-1 in integrin mechanotransduction, we exposed bovine aortic endothelial cells to 10 dyn/cm2 of laminar shear stress. Caveolin-1 was acutely and transiently phosphorylated with shear, occurring downstream of β1-integrin activation as the β1-integrin blocking antibody JB1A was inhibitory. In manipulating Src family kinase (SFK) activity with knockdown of Csk or type 1 protein phosphatase (PP1) treatment, we observed coordinate increase and decrease in shear-induced caveolin-1 phosphorylation, respectively. Hence, shear-stimulated caveolin-1 phosphorylation is regulated by SFKs. Shear-induced recruitment and phosphorylation of caveolin-1 occurred at β1-integrin sites in a β1-integrin- and SFK-dependent manner. Csk, described to interact with pY14-caveolin-1 and integrins, bound to an increased pool of phosphorylated caveolin-1 after shear corresponding with elevated Csk at β1-integrin sites. Like caveolin-1, treatment with JB1A and PP1 attenuated shear-induced Csk association with β1-integrins. Csk function was assayed with transfection of a caveolin-1 phosphorylation domain peptide. The peptide attenuated shear-induced association of Csk at β1-integrin sites, as well as colocalization of Csk with paxillin and phosphorylated caveolin-1. Because integrin and Csk activity regulate cytoskeletal reorganization, we evaluated the role of this mechanism in shear-induced myosin light chain (MLC) phosphorylation. Knockdown of Csk expression was sufficient to reduce MLC diphosphorylation due to shear. Disruption of Csk-integrin association by peptide treatment was also inhibitory of the MLC diphosphorylation response. Together these data indicate that integrin activation with shear stress results in SFK-regulated caveolin-1 phosphorylation that, in turn, mediates Csk association at integrin sites, where it plays a role in downstream, shear-stimulated MLC diphosphorylation.

Mechanosensing machinery for cells under low substratum rigidity

2008 ◽  
Vol 295 (6) ◽  
pp. C1579-C1589 ◽  
Author(s):  
Wei-Chun Wei ◽  
Hsi-Hui Lin ◽  
Meng-Ru Shen ◽  
Ming-Jer Tang
Keyword(s):  
Physical Environment ◽  
Resonance Energy Transfer ◽  
Collagen Gel ◽  
Resonance Energy ◽  
Dependent Manner ◽  
Mechanical Stimuli ◽  
Integrin Activation ◽  
Integrin Clustering ◽  
Dose Dependent ◽  
Insight Into

Mechanical stimuli are essential during development and tumorigenesis. However, how cells sense their physical environment under low rigidity is still unknown. Here we show that low rigidity of collagen gel downregulates β1-integrin activation, clustering, and focal adhesion kinase (FAK) Y397 phosphorylation, which is mediated by delayed raft formation. Moreover, overexpression of autoclustered β1-integrin (V737N), but not constitutively active β1-integrin (G429N), rescues FAKY397 phosphorylation level suppressed by low substratum rigidity. Using fluorescence resonance energy transfer to assess β1-integrin clustering, we have found that substratum rigidity between 58 and 386 Pa triggers β1-integrin clustering in a dose-dependent manner, which is highly dependent on actin filaments but not microtubules. Furthermore, augmentation of β1-integrin clustering enhances the interaction between β1-integrin, FAK, and talin. Our results indicate that contact with collagen fibrils is not sufficient for integrin activation. However, substratum rigidity is required for integrin clustering and activation. Together, our findings provide new insight into the mechanosensing machinery and the mode of action for epithelial cells in response to their physical environment under low rigidity.

scholarly journals Increased extracellular pressure enhances cancer cell integrin-binding affinity through phosphorylation of β1-integrin at threonine 788/789

2009 ◽  
Vol 296 (1) ◽  
pp. C193-C204 ◽  
Author(s):  
David H. Craig ◽  
Christopher P. Gayer ◽  
Keri L. Schaubert ◽  
Yanzhang Wei ◽  
Jinhua Li ◽  
...  
Keyword(s):  
Cell Adhesion ◽  
Binding Affinity ◽  
Cancer Cell ◽  
Elevated Pressure ◽  
Colon Cancer Cells ◽  
Wild Type ◽  
Cancer Cell Adhesion ◽  
Integrin Clustering ◽  
Murine Fibroblasts

Increased extracellular pressure stimulates β1-integrin-dependent cancer cell adhesion. We asked whether pressure-induced adhesion is mediated by changes in β1-integrin binding affinity or avidity and whether these changes are phosphorylation dependent. We evaluated integrin affinity and clustering in human SW620 colon cancer cells by measuring differences in binding between soluble Arg-Gly-Asp (RGD)-Fc ligands and RGD-Fc-F(ab′)2 multimeric complexes under ambient and 15-mmHg increased pressures. Phosphorylation of β1-integrin S785 and T788/9 residues in SW620 and primary malignant colonocytes was assessed in parallel. We further used GD25-β1-integrin-null murine fibroblasts stably transfected with either wild-type β1A-integrin, S785A, TT788/9AA, or T788D mutants to investigate the role of β1-integrin site-specific phosphorylation. SW620 binding of RGD-Fc-F(ab′)2 multimeric complexes, but not soluble RGD-Fc ligands, was sensitive to integrin clustering. RGD-Fc ligand binding was significantly increased under elevated pressure, suggesting that pressure modulates β1-integrin affinity. Pressure stimulated both β1-integrin S785 and T788/9 phosphorylation. GD25-β1A-integrin wild-type and S785A cells displayed an increase in adhesion to fibronectin under elevated pressure, an effect absent in β1-integrin-null and TT788/9AA cells. T788D substitution significantly elevated basal cell adhesion but displayed no further increase under pressure. These results suggest pressure-induced cell adhesion is mediated by β1-integrin T788/9 phosphorylation-dependent changes in integrin binding affinity.

scholarly journals Radil controls neutrophil adhesion and motility through β2-integrin activation

2012 ◽  
Vol 23 (24) ◽  
pp. 4751-4765 ◽  
Author(s):  
Lunhua Liu ◽  
Wulin Aerbajinai ◽  
Syed M. Ahmed ◽  
Griffin P. Rodgers ◽  
Stephane Angers ◽  
...  
Keyword(s):  
Cell Adhesion ◽  
Inflammatory Responses ◽  
Neutrophil Adhesion ◽  
Dependent Manner ◽  
Neutrophil Chemotaxis ◽  
Integrin Activation ◽  
Chemotactic Migration ◽  
Long Tail ◽  
Β2 Integrin ◽  
Inhibitory Antibodies

Integrin activation is required to facilitate multiple adhesion-dependent functions of neutrophils, such as chemotaxis, which is critical for inflammatory responses to injury and pathogens. However, little is known about the mechanisms that mediate integrin activation in neutrophils. We show that Radil, a novel Rap1 effector, regulates β1- and β2-integrin activation and controls neutrophil chemotaxis. On activation and chemotactic migration of neutrophils, Radil quickly translocates from the cytoplasm to the plasma membrane in a Rap1a-GTP–dependent manner. Cells overexpressing Radil show a substantial increase in cell adhesion, as well as in integrin/focal adhesion kinase (FAK) activation, and exhibit an elongated morphology, with severe tail retraction defects. This phenotype is effectively rescued by treatment with either β2-integrin inhibitory antibodies or FAK inhibitors. Conversely, knockdown of Radil causes severe inhibition of cell adhesion, β2-integrin activation, and chemotaxis. Furthermore, we found that inhibition of Rap activity by RapGAP coexpression inhibits Radil-mediated integrin and FAK activation, decreases cell adhesion, and abrogates the long-tail phenotype of Radil cells. Overall, these studies establish that Radil regulates neutrophil adhesion and motility by linking Rap1 to β2-integrin activation.

scholarly journals Modulation of RGD sequence motifs regulates disintegrin recognition of αIIbβ3 and α5β1 integrin complexes

1998 ◽  
Vol 335 (2) ◽  
pp. 247-257 ◽  
Author(s):  
Salman RAHMAN ◽  
Alex AITKEN ◽  
Geraldine FLYNN ◽  
Caroline FORMSTONE ◽  
Geoffrey F. SAVIDGE
Keyword(s):  
Cell Adhesion ◽  
Inhibitory Activity ◽  
K562 Cell ◽  
Platelet Adhesion ◽  
K562 Cells ◽  
Dependent Manner ◽  
Sequence Motifs ◽  
Wild Type ◽  
Rgd Sequence

Several recent studies have demonstrated that the amino acid residues flanking the RGD sequence of high-affinity ligands modulate their specificity of interaction with integrin complexes. The present study has addressed the role of the residues flanking the RGD sequence in regulating the recognition by disintegrin of the αIIbβ3 and α5β1 complexes by construction of a panel of recombinant molecules of Elegantin (the platelet aggregation inhibitor from the venom of Trimerasurus elegans) expressing specific RGD sequence motifs. Wild-type Elegantin (ARGDNP) and several variants including Eleg. AM (ARGDMP), Eleg. PM (PRGDMP) and Eleg. PN (PRGDNP) were expressed as glutathione S-transferase (GST) fusion proteins in Escherichia coli. The inhibitory efficacies of the panel of Elegantin variants were analysed in platelet adhesion assays with substrates immobilized with fibrinogen and fibronectin. Elegantin molecules containing an Ala residue N-terminal to the RGD sequence (wild-type Elegantin and Eleg. AM) showed strong inhibitory activity towards αIIbβ3-dependent platelet adhesion on fibronectin, whereas a Pro residue in this position (Eleg. PM and Kistrin, the inhibitor from the venom of Calloselasma rhodostoma) engendered lower activity. The decreased activity could not be attributed to a decrease in the affinity of the disintegrin for the αIIbβ3 complex because both Eleg. AM and Eleg. PM had similar Kd (app) values. In contrast, Elegantin molecules into which a Met residue was introduced in place of the Asn residue C-terminal to the RGD sequence showed 10–13-fold elevated inhibitory activity towards platelet adhesion on fibrinogen and this was maintained with either a Pro or Ala residue N-terminal to the RGD sequence. In experiments with the α5β1 complex on K562 cells, the inhibitory efficacies of the panel of Elegantin molecules were analysed under two different cation conditions. First, in the presence of Ca2+/Mg2+, K562 cell adhesion on fibronectin was inhibited equally well by Elegantin and Eleg. AM but inhibited poorly by Eleg. PM and Kistrin. In contrast with platelets, the decreased inhibitory efficacy of the PRGDMP disintegrins was due to poor recognition of the α5β1 complex. In the presence of Mn2+ cation, K562 cell adhesion on fibrinogen was observed in an α5β1-dependent manner. Under these conditions both PRGD and ARGD containing disintegrins were strong inhibitors of K562 cell adhesion on fibrinogen and this was due to a markedly improved recognition of the α5β1 complex by the PRGD molecules. These observations demonstrate the pivotal role of the amino acids flanking the RGD sequence for disintegrin recognition of integrin complexes and highlight the subtle nature by which integrin-ligand binding specificity can be modulated by both cation and adhesive motif.

scholarly journals Suppression of integrin activation by the membrane-distal sequence of the integrin alphaIIb cytoplasmic tail

2004 ◽  
Vol 379 (2) ◽  
pp. 317-323 ◽  
Author(s):  
Jun YAMANOUCHI ◽  
Takaaki HATO ◽  
Tatsushiro TAMURA ◽  
Shigeru FUJITA
Keyword(s):  
Distal Region ◽  
Metabolic Energy ◽  
Single Amino Acid ◽  
Dependent Manner ◽  
Integrin Activation ◽  
Α Subunit ◽  
Ample Evidence ◽  
Cytoplasmic Tails ◽  
Energy Dependent

Integrin cytoplasmic tails regulate integrin activation including an increase in integrin affinity for ligands. Although there is ample evidence that the membrane-proximal regions of the α and β tails interact with each other to maintain integrins in a low-affinity state, little is known about the role of the membrane-distal region of the α tail in regulation of integrin activation. We report a critical sequence for regulation of integrin activation in the membranedistal region of the αIIb tail. Alanine substitution of the RPP residues in the αIIb tail rendered αIIbβ3 constitutively active in a metabolic energy-dependent manner. Although an αIIb/α6Aβ3 chimaeric integrin, in which the αIIb tail was replaced by the α6A tail, was in an energy-dependent active state to bind soluble ligands, introduction of the RPP sequence into the α6A tail inhibited binding of an activation-dependent antibody PAC1. In αIIb/α6Aβ3, deleting the TSDA sequence from the α6A tail or single amino acid substitutions of the TSDA residues inhibited αIIb/α6Aβ3 activation and replacing the membrane-distal region of the αIIb tail with TSDA rendered αIIbβ3 active, suggesting a stimulatory role of TSDA in energy-dependent integrin activation. However, adding TSDA to the αIIb tail containing the RPP sequence of the membrane-distal region failed to activate αIIbβ3. These results suggest that the RPP sequence after the GFFKR motif of the αIIb tail suppresses energy-dependent αIIbβ3 activation. These findings provide a molecular basis for the regulation of energy-dependent integrin activation by α subunit tails.

scholarly journals DOWN REGULATION OF u-PA BY A NUTRIENT MIXTURE IN HEMANGIOMA (EOMA) CELLS BY INDUCING CASPASE-DEPENDENT APOPTOSIS

2018 ◽  
Vol 40 (2) ◽  
pp. 90-94
Author(s):  
M W Roomi ◽  
B Bhanap ◽  
A Niedzwiecki ◽  
M Rath
Keyword(s):  
Morphological Changes ◽  
Experimental Studies ◽  
Positive Control ◽  
Surgical Removal ◽  
Dependent Manner ◽  
Antitumor Effects ◽  
Tea Extract ◽  
Dose Dependent

Hemangiomas are the most common congenital vascular and benign tumor in infants and children. Most hemangiomas do not cause major symptoms to require intervention, however, the larger hemangiomas have tendency to bleed and may require surgical removal. Experimental studies have demonstrated the role of urokinase plasminogen activator (u-PA), especially cell surface u-PA, as an initiator of extra-cellular matrix proteolysis and associated tumor cell invasion. Aim: To examine, whether the antitumor effects of a specific nutrient mixture are due to induction of apoptosis by inhibition of u-PA. Materials and Methods: A nutrient mixture containing lysine, proline, ascorbic acid, and green tea extract which has showed anticancer activity against a number of cancer cell lines was used as an experimental composition. EOMA cells were grown in appropriate media with antibiotics in 24well tissue culture plates. At near confluence, the cells were treated with nutrition mixture at 10, 100, 1000 µg/ml in triplicate. Analysis of u-PA activity was carried out by fibrin zymography. Morphological changes and caspase activation associated with apoptosis induction was checked by H&E staining and Live Green caspase assay, respectively. Apoptosis inducing anticancer drug camptothecin (10 µM) was used as positive control. Results: The nutrition mixture exhibited dose response toxicity with maximum toxicity 55% (p < 0.001) at 1000 µg/ml. EOMA cells expressed u-PA, which was inhibited by nutrition mixture in a dose-dependent manner. The caspase analysis revealed a dose dependent increase in apoptosis of EOMA hemangioma cells, with an increasing apoptosis observed at 100 µg/ml, and maximum at 1000 µg/ml. Cells treated with nutrition mixture showed significantly more apoptotic changes than the control or camptothecin-treated cells. Conclusion: These results suggest that NM may induce apoptosis of hemangioma cells in vitro thus warranting further investigation.

scholarly journals Regulation of urokinase receptor function and pericellular proteolysis by the integrin α5β1

2009 ◽  
Vol 101 (05) ◽  
pp. 954-962 ◽  
Author(s):  
Rosemary Bass ◽  
Vincent Ellis
Keyword(s):  
Cell Adhesion ◽  
Cell Types ◽  
Dependent Manner ◽  
Integrin Α5β1 ◽  
Upa Receptor ◽  
Cellular Localisation ◽  
Dependent Cell ◽  
Invasive Cell ◽  
Pericellular Proteolysis

SummaryInteractions between the uPA receptor (uPAR) and various inte-grins, including α5β1, are known to modulate integrin-dependent cell adhesion, and we have shown that the integrin-associated tetraspanin protein CD82 down-regulates uPAR-dependent plasminogen activation by affecting α5β1 cellular localisation. Here we have investigated whether overexpression of α5β1 directly affects uPAR-dependent pericellular proteolysis. CHO cells overexpressing α5β1 were found to activate plasminogen at a rate up to 18-fold faster than B2CHO cells which are α5-deficient. This effect was dependent on the activation state of α5β1, as it was maximal in the presence of Mn2+. To determine the role of uPAR-α5β1 interactions in this effect, we determined the adhesion of these cells to immobilised soluble uPAR (suPAR). Neither cell-type was found to adhere to suPAR, but both cell types were found to adhere to an anti-uPAR monoclonal antibody in a uPAR- and integrin-dependent manner. This adhesion was 10-fold greater in the absence of α5β1, possibly implicating the involvement of non-α5-integrins. Soluble forms of the various components were used to investigate the molecular basis of these effects, but no direct interactions could be demonstrated between α5β1 and either uPAR, uPA or uPA-uPAR complex. This suggests that assembly of these components on the plasma membrane is required to influence uPAR function, increasing uPAR-dependent pericellular proteolysis and decreasing uPAR-dependent cell adhesion. These interactions may be modified by other integrins, suggesting a complex interplay between uPAR and integrins on the cell surface with the potential to regulate invasive cell migration.

scholarly journals The F1 loop of the talin head domain acts as a gatekeeper in integrin activation and clustering

2020 ◽  
Vol 133 (19) ◽  
pp. jcs239202 ◽  
Author(s):  
Sampo Kukkurainen ◽  
Latifeh Azizi ◽  
Pingfeng Zhang ◽  
Marie-Claude Jacquier ◽  
Mo Baikoghli ◽  
...  
Keyword(s):  
Cell Adhesion ◽  
Cytoplasmic Tail ◽  
First Person ◽  
Integrin Activation ◽  
Inner Membrane ◽  
Flexible Loop ◽  
Head Domain ◽  
Integrin Clustering ◽  
Membrane Bound ◽  
Β3 Integrin

ABSTRACTIntegrin activation and clustering by talin are early steps of cell adhesion. Membrane-bound talin head domain and kindlin bind to the β integrin cytoplasmic tail, cooperating to activate the heterodimeric integrin, and the talin head domain induces integrin clustering in the presence of Mn2+. Here we show that kindlin-1 can replace Mn2+ to mediate β3 integrin clustering induced by the talin head, but not that induced by the F2–F3 fragment of talin. Integrin clustering mediated by kindlin-1 and the talin head was lost upon deletion of the flexible loop within the talin head F1 subdomain. Further mutagenesis identified hydrophobic and acidic motifs in the F1 loop responsible for β3 integrin clustering. Modeling, computational and cysteine crosslinking studies showed direct and catalytic interactions of the acidic F1 loop motif with the juxtamembrane domains of α- and β3-integrins, in order to activate the β3 integrin heterodimer, further detailing the mechanism by which the talin–kindlin complex activates and clusters integrins. Moreover, the F1 loop interaction with the β3 integrin tail required the newly identified compact FERM fold of the talin head, which positions the F1 loop next to the inner membrane clasp of the talin-bound integrin heterodimer.This article has an associated First Person interview with the first author of the paper.

Integrin Activation

2001 ◽  
Vol 86 (07) ◽  
pp. 316-323 ◽  
Author(s):  
D. G. Woodside ◽  
S. Liu ◽  
M. H. Ginsberg
Keyword(s):  
Cell Adhesion ◽  
Cell Surface ◽  
Ligand Binding ◽  
Cellular Adhesion ◽  
Structural Basis ◽  
Surface Adhesion ◽  
Integrin Activation ◽  
Ligand Activation ◽  
Dependent Cell

SummaryIntegrins are cell surface adhesion receptors that participate in a variety of important processes throughout the vasculature. Here we summarize some recent findings on the regulation of integrin mediated cellular adhesion. Particular emphasis is placed on the regulation of integrin affinity for ligand (activation), although this is just one mechanism by which regulation of integrin-dependent cell adhesion can occur. Also discussed are recent observations on the structural basis of integrin activation, the role of the cytoplasmic domain in integrin affinity regulation, and potential mechanisms by which activation signals are propagated from integrin cytoplasmic domains to the extracellular ligand binding domain.

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