The heel and toe of the cell's foot: A multifaceted approach for understanding the structure and dynamics of focal adhesions

Wolfenson, Haguy; Henis, Yoav I.; Geiger, Benjamin; Bershadsky, Alexander D.

doi:10.1002/cm.20410

Cited by 115 publications

(111 citation statements)

References 132 publications

(175 reference statements)

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“…Actin stress fibers are contractile actomyosin structures that play an important role in regulating cell morphology, motility and adhesion (Tojkander et al, 2012). Focal adhesions consist of protein complexes, which connect the cytoskeleton to the extracellular matrix (Wolfenson et al, 2009). We propose that compromised focal adhesion and stress fiber formation in HMI-1a3-treated cells leads to decreased cell adhesion and contractility, which are manifested by cell elongation.…”

Section: Discussionmentioning

confidence: 97%

Evidence for a role of MRCK in mediating HeLa cell elongation induced by the C1 domain ligand HMI-1a3

Talman

Gateva

Ahti

et al. 2014

European Journal of Pharmaceutical Sciences

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Diacylglycerol (DAG) is a central mediator of signaling pathways that regulate cell proliferation, survival and apoptosis. Therefore, C1 domain, the DAG binding site within protein kinase C (PKC) and other DAG effector proteins, is considered a potential cancer drug target. Derivatives of 5-(hydroxymethyl)isophthalic acid are a novel group of C1 domain ligands with antiproliferative and differentiation-inducing effects. Our previous work showed that these isophthalate derivatives exhibit antiproliferative and elongation-inducing effects in HeLa human cervical cancer cells. In this study we further characterized the effects of bis(3-trifluoromethylbenzyl) 5-(hydroxymethyl)isophthalate (HMI-1a3) on HeLa cell proliferation and morphology. HMI-1a3-induced cell elongation was accompanied with loss of focal adhesions and actin stress fibers, and exposure to HMI-1a3 induced a prominent relocation of cofilin-1 into the nucleus regardless of cell phenotype. The antiproliferative and morphological responses to HMI-1a3 were not modified by coexposure to pharmacological inhibition or activation of PKC, or by RNAi knock-down of specific PKC isoforms, suggesting that the effects of HMI-1a3 were not mediated by PKC. Genome-wide gene expression microarray and gene set enrichment analysis suggested that, among others, HMI-1a3 induces changes in small GTPasemediated signaling pathways. Our experiments revealed that the isophthalates bind also to the C1 domains of β2-chimaerin, protein kinase D (PKD) and MRCK, which are potential mediators of small GTPase signaling and cytoskeletal reorganization. Pharmacological inhibition of MRCK, but not that of PKD attenuated HMI1a3-induced cell elongation, suggesting that MRCK participates in mediating the effects of HMI-1a3 on HeLa cell morphology.

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Section: Discussionmentioning

confidence: 97%

Evidence for a role of MRCK in mediating HeLa cell elongation induced by the C1 domain ligand HMI-1a3

Talman

Gateva

Ahti

et al. 2014

European Journal of Pharmaceutical Sciences

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“…Our analysis of subadhesion, multidimensional traction stress introduces additional factors to consider. These distinct mechanical loads within a single focal adhesion could contribute to the spatial structure of adhesions, for example by differentially regulating protein binding, conformational changes, and force-induced signaling (35). Tension in the distal end could promote protein unfolding and interactions (e.g., between vinculin, talin, and actin) to mediate a molecular clutch (36)(37)(38)(39) or phosphorylation (40), whereas shearing and compression in the proximal end may induce the rapid protein exchanges that mediate focal adhesion elongation (41)(42)(43)(44).…”

Section: Discussionmentioning

confidence: 99%

Multidimensional traction force microscopy reveals out-of-plane rotational moments about focal adhesions

Legant

Choi

Miller

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

251

239

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Recent methods have revealed that cells on planar substrates exert both shear (in-plane) and normal (out-of-plane) tractions against the extracellular matrix (ECM). However, the location and origin of the normal tractions with respect to the adhesive and cytoskeletal elements of cells have not been elucidated. We developed a highspatiotemporal-resolution, multidimensional (2.5D) traction force microscopy to measure and model the full 3D nature of cellular forces on planar 2D surfaces. We show that shear tractions are centered under elongated focal adhesions whereas upward and downward normal tractions are detected on distal (toward the cell edge) and proximal (toward the cell body) ends of adhesions, respectively. Together, these forces produce significant rotational moments about focal adhesions in both protruding and retracting peripheral regions. Temporal 2.5D traction force microscopy analysis of migrating and spreading cells shows that these rotational moments are highly dynamic, propagating outward with the leading edge of the cell. Finally, we developed a finite element model to examine how rotational moments could be generated about focal adhesions in a thin lamella. Our model suggests that rotational moments can be generated largely via shear lag transfer to the underlying ECM from actomyosin contractility applied at the intracellular surface of a rigid adhesion of finite thickness. Together, these data demonstrate and probe the origin of a previously unappreciated multidimensional stress profile associated with adhesions and highlight the importance of new approaches to characterize cellular forces.cell mechanics | mechanotransduction | migration | actin U nderstanding how cells generate and respond to mechanical forces is critical in cell biology. In anchorage-dependent cells, myosin-II cross-links and contracts actin filaments to generate tension, which is transmitted to the extracellular matrix (ECM) via integrin-mediated adhesions (1-4). The traction stresses (force per area) exerted between adhesions and the ECM drive cell spreading and migration in morphogenesis (5, 6), wound healing (7), and tumor metastasis (8, 9). In addition, these stresses induce changes in adhesion signaling, cytoskeletal reorganization, and gene expression (4, 10-13), thereby regulating functions such as proliferation (14, 15) and differentiation (16,17).Measurements of cellular traction stresses have advanced our understanding of mechanotransduction and enabled quantitative modeling of cellular interactions with the ECM (18-20). These measurements reveal that cells exert inwardly oriented tractions at their periphery, where focal adhesions grow centripetally (3,4,21). However, the vast majority of methods (collectively termed traction force microscopy, TFM) have assumed that cells exert only shear forces (parallel to the plane of the substrate). Interestingly, recent studies have demonstrated that cells on planar substrata exert significant vertical (normal) tractions, indicating that patterns of cellular force generation...

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“…Cell migration requires dramatic changes in cell shape and in adhesion to the extracellular matrix, and both these processes must be spatiotemporally coordinated (Wolfenson et al, 2009;Gardel et al, 2010).To analyze focal adhesions and actin cytoskeleton, MTLn3-EGFR Ctr and p140Cap cells were plated on collagen, serum deprived for 4 h and stimulated with EGF for 5-15 min. As shown in Figure 4a, p140 cells show increased paxillin staining of focal adhesions compared with control cells.…”

Section: P140cap Inhibits Directed Cell Migrationmentioning

confidence: 99%

p140Cap suppresses the invasive properties of highly metastatic MTLn3-EGFR cells via impaired cortactin phosphorylation

et al. 2011

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We have recently shown that the adaptor protein p140Cap regulates tumor properties in terms of cell motility and growth. Here, by using the highly metastatic rat adenocarcinoma cell line MTLn3-epidermal growth factor receptor (EGFR), we assess the role of p140Cap in metastasis formation. Orthotopic transplantation of MTLn3-EGFR cells over-expressing p140Cap in Rag2À/À mice resulted in normal primary tumor growth compared with the controls. Strikingly, p140Cap over-expression causes an 80% inhibition in the number of lung metastases. p140Cap over-expressing cells display a 50% reduction in directional cell migration, an increased number and size of focal adhesions, and a strong impairment in the ability to invade in a 3D matrix. p140Cap overexpression affects EGFR signaling and tyrosine phosphorylation of cortactin in response to EGF stimulation. Intriguingly, p140Cap associates with cortactin via interaction with its second proline-rich domain to the cortactin SH3 domain. The phosphomimetic cortactin tyrosine 421 mutant rescues migration and invasive properties in p140Cap over-expressing cells. Taken together, these data demonstrate that p140Cap suppresses the invasive properties of highly metastatic breast carcinoma cells by inhibiting cortactin-dependent cell motility.

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The heel and toe of the cell's foot: A multifaceted approach for understanding the structure and dynamics of focal adhesions

Cited by 115 publications

References 132 publications

Evidence for a role of MRCK in mediating HeLa cell elongation induced by the C1 domain ligand HMI-1a3

Evidence for a role of MRCK in mediating HeLa cell elongation induced by the C1 domain ligand HMI-1a3

Multidimensional traction force microscopy reveals out-of-plane rotational moments about focal adhesions

p140Cap suppresses the invasive properties of highly metastatic MTLn3-EGFR cells via impaired cortactin phosphorylation

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