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Staff member

Pere Roca-Cusachs Soulere

Group Leader
+34 934 020 863
procaibecbarcelona.eu
CV Summary
Pere Roca-Cusachs obtained his PhD in cellular biophysics in 2007 from the Medical School at the University of Barcelona. He then worked in the lab of Prof. Michael Sheetz (Columbia University) as a post-doctoral researcher until 2011. He established his group at IBEC in 2012. is currently Full Professor at the University of Barcelona, and Group Leader at the Institute for bioengineering of Catalonia (IBEC). His group studies the physical and molecular mechanisms by which cells detect and respond to mechanical signals. He is an EMBO member, and recipient of the EMBO Young Investigator award, the 2017 City of Barcelona award to the life sciences, the 2019 Young Investigator Award of the European Biophysical Societies Association (EBSA), and the ICREA acadèmia award.
Staff member publications

Rossetti, Leone, Grosser, Steffen, Abenza, Juan Francisco, Valon, Leo, Roca-Cusachs, Pere, Alert, Ricard, Trepat, Xavier, (2024). Optogenetic generation of leader cells reveals a force-velocity relation for collective cell migration Nature Physics

During development, wound healing and cancer invasion, migrating cell clusters feature highly protrusive leader cells at their front. Leader cells are thought to pull and direct their cohort of followers, but whether their local action is enough to guide the entire cluster, or if a global mechanical organization is needed, remains controversial. Here we show that the effectiveness of the leader-follower organization is proportional to the asymmetry of traction and tension within cell clusters. By combining hydrogel micropatterning and optogenetic activation, we generate highly protrusive leaders at the edge of minimal cell clusters. We find that the induced leader can robustly drag one follower but not larger groups. By measuring traction forces and tension propagation in clusters of increasing size, we establish a quantitative relationship between group velocity and the asymmetry of the traction and tension profiles. Modelling motile clusters as active polar fluids, we explain this force-velocity relationship in terms of asymmetries in the active traction profile. Our results challenge the notion of autonomous leader cells, showing that collective cell migration requires global mechanical organization within the cluster. Leader cells play an important role in guiding migratory clusters in various biological processes. Now, the mechanical organization of leader and followers within a cell cluster is shown to enable collective migration.

JTD Keywords: Driven, Dynamics, Guidance, Require


Granero-Moya I, Venturini V, Belthier G, Groenen B, Molina-Jordán M, González-Martín M, Trepat X, van Rheenen J, Andreu I, Roca-Cusachs P, (2024). Nucleocytoplasmic transport senses mechanics independently of cell density in cell monolayers. Journal Of Cell Science 137, jcs.262363

Cells sense and respond to mechanical forces through mechanotransduction, which regulates processes in health and disease. In single adhesive cells, mechanotransduction involves the transmission of force from the extracellular matrix to the cell nucleus, where it affects nucleocytoplasmic transport (NCT) and the subsequent nuclear localization of transcriptional regulators such as YAP. However, if and how NCT is mechanosensitive in multicellular systems is unclear. Here, we characterize and use a fluorescent sensor of nucleocytoplasmic transport (Sencyt) and demonstrate that nucleocytoplasmic transport responds to mechanics but not cell density in cell monolayers. Using monolayers of both epithelial and mesenchymal phenotype, we show that NCT is altered in response both to osmotic shocks, and to the inhibition of cell contractility. Further, NCT correlates with the degree of nuclear deformation measured through nuclear solidity, a shape parameter related to nuclear envelope tension. In contrast, YAP but NCT is sensitive to cell density, showing that YAP response to cell-cell contacts is not via a mere mechanical effect of NCT. Our results demonstrate the generality of the mechanical regulation of NCT.

JTD Keywords: Cell nucleus, Mechanobiology, Mechanotransduction, Sensor


Conti, Sefora, Venturini, Valeria, Canellas-Socias, Adria, Cortina, Carme, Abenza, Juan F, Attolini, Camille Stephan-Otto, Guerra, Emily Middendorp, Xu, Catherine K, Li, Jia Hui, Rossetti, Leone, Stassi, Giorgio, Roca-Cusachs, Pere, Diz-Munoz, Alba, Ruprecht, Verena, Guck, Jochen, Batlle, Eduard, Labernadie, Anna, Trepat, Xavier, (2024). Membrane to cortex attachment determines different mechanical phenotypes in LGR5+and LGR5-colorectal cancer cells Nature Communications 15, 3363

Colorectal cancer (CRC) tumors are composed of heterogeneous and plastic cell populations, including a pool of cancer stem cells that express LGR5. Whether these distinct cell populations display different mechanical properties, and how these properties might contribute to metastasis is poorly understood. Using CRC patient derived organoids (PDOs), we find that compared to LGR5- cells, LGR5+ cancer stem cells are stiffer, adhere better to the extracellular matrix (ECM), move slower both as single cells and clusters, display higher nuclear YAP, show a higher survival rate in response to mechanical confinement, and form larger transendothelial gaps. These differences are largely explained by the downregulation of the membrane to cortex attachment proteins Ezrin/Radixin/Moesin (ERMs) in the LGR5+ cells. By analyzing single cell RNA-sequencing (scRNA-seq) expression patterns from a patient cohort, we show that this downregulation is a robust signature of colorectal tumors. Our results show that LGR5- cells display a mechanically dynamic phenotype suitable for dissemination from the primary tumor whereas LGR5+ cells display a mechanically stable and resilient phenotype suitable for extravasation and metastatic growth. The mechanical properties of heterogeneous cell populations in colorectal tumors and the relevance to cancer metastasis remain not fully understood. Here, the authors suggest that the variations in malignant phenotypes between LGR5-positive cancer stem cells and LGR5-negative cells could be due to their distinct mechanical phenotypes observed in vitro, determined by the membrane to cortex attachment proteins Ezrin/Radixin/Moesin.

JTD Keywords: , , Adhesion, Deformability, E-cadherin, Erm proteins, Expression, Metastasis, Organization, Plasticit, Stem-cells, Tumor-cells


Kechagia, Z, Roca-Cusachs, P, (2023). Cytoskeletal safeguards: Protecting the nucleus from mechanical perturbations Curr Opin Biomed Eng 28, 100494

The cell nucleus plays a key role in cellular mechanoresponses. 3D genome organisation, gene expression, and cell behaviour, in general, are affected by mechanical force application to the nucleus, which is transmitted from the cellular environment via a network of interconnected cytoskeletal components. To effectively regulate cell responses, these cytoskeletal components must not only exert forces but also withstand external forces when necessary. This review delves into the latest research concerning how the cytoskeleton safeguards the nucleus from mechanical perturbations. Spe-cifically, we focus on the three primary cytoskeletal polymers: actin, intermediate filaments, and microtubules, as well as their interactions with the cell nucleus. We discuss how the cyto-skeleton acts as a protective shield for the nucleus, ensuring structural integrity and conveying context-specific mechanoresponses.

JTD Keywords: Actin, Architecture, Cytoskeleton, Envelope, F-actin, Filaments, Force, Genome, Intermediate filaments, Lamin, Mechanotransduction, Membrane protein, Microtubules, Nesprin-1, Nucleus


Kechagia, Z, Sáez, P, Gómez-González, M, Canales, B, Viswanadha, S, Zamarbide, M, Andreu, I, Koorman, T, Beedle, AEM, Elosegui-Artola, A, Derksen, PWB, Trepat, X, Arroyo, M, Roca-Cusachs, P, (2023). The laminin-keratin link shields the nucleus from mechanical deformation and signalling Nature Materials 22, 1409-1420

The mechanical properties of the extracellular matrix dictate tissue behaviour. In epithelial tissues, laminin is a very abundant extracellular matrix component and a key supporting element. Here we show that laminin hinders the mechanoresponses of breast epithelial cells by shielding the nucleus from mechanical deformation. Coating substrates with laminin-111-unlike fibronectin or collagen I-impairs cell response to substrate rigidity and YAP nuclear localization. Blocking the laminin-specific integrin β4 increases nuclear YAP ratios in a rigidity-dependent manner without affecting the cell forces or focal adhesions. By combining mechanical perturbations and mathematical modelling, we show that β4 integrins establish a mechanical linkage between the substrate and keratin cytoskeleton, which stiffens the network and shields the nucleus from actomyosin-mediated mechanical deformation. In turn, this affects the nuclear YAP mechanoresponses, chromatin methylation and cell invasion in three dimensions. Our results demonstrate a mechanism by which tissues can regulate their sensitivity to mechanical signals.© 2023. The Author(s).

JTD Keywords: actin, cell migration, filaments, force transmission, localization, membrane, motility, proteins, yap, Cell adhesion, Cytoskeleton, Extracellular matrix, Fibronectins, Integrin alpha-6-beta-4, Integrins, Keratins, Laminin


Beedle, AE, Roca-Cusachs, P, (2023). The reversibility of cellular mechano-activation Current Opinion In Cell Biology 84, 102229

The cellular microenvironment is highly heterogeneous and dynamic. Therefore, cells must be equipped with molecular tools to adapt and respond to constantly fluctuating inputs. One such input is mechanical force, which activates signalling and regulates cell behaviour in the process of mechanotransduction. Whereas the mechanisms activating mechanotransduction are well studied, the reversibility of this process, whereby cells disassemble and reverse force-activated signalling pathways upon cessation of mechanical stimulation is far less understood. In this review we will outline some of the key experimental techniques to investigate the reversibility of mechanical signalling, and key discoveries arising from them.Copyright © 2023 The Authors. Published by Elsevier Ltd.. All rights reserved.

JTD Keywords: mechanical memory, memory, reversibility, Mechanical memory, Mechanotransduction, Nuclear, Reversibility


Noordstra, I, Hermoso, MD, Schimmel, L, Bonfim-Melo, A, Currin-Ross, D, Duong, CN, Kalappurakkal, JM, Morris, RG, Vestweber, D, Mayor, S, Gordon, E, Roca-Cusachs, P, Yap, AS, (2023). An E-cadherin-actin clutch translates the mechanical force of cortical flow for cell-cell contact to inhibit epithelial cell locomotion Developmental Cell 58, 1748-+

Adherens junctions (AJs) allow cell contact to inhibit epithelial migration yet also permit epithelia to move as coherent sheets. How, then, do cells identify which contacts will inhibit locomotion? Here, we show that in human epithelial cells this arises from the orientation of cortical flows at AJs. When the leader cells from different migrating sheets make head-on contact with one another, they assemble AJs that couple together oppositely directed cortical flows. This applies a tensile signal to the actin-binding domain (ABD) of a-cate-nin, which provides a clutch to promote lateral adhesion growth and inhibit the lamellipodial activity neces-sary for migration. In contrast, AJs found between leader cells in the same migrating sheet have cortical flows aligned in the same direction, and no such mechanical inhibition takes place. Therefore, a-catenin mechano-sensitivity in the clutch between E-cadherin and cortical F-actin allows cells to interpret the direction of motion via cortical flows and signal for contact to inhibit locomotion.

JTD Keywords: Clutch, Contact inhibition of locomotion, Cortical flow, E-cadherin adhesion, Mechanical tension, Α-catenin


Quiroga, X, Walani, N, Disanza, A, Chavero, A, Mittens, A, Tebar, F, Trepat, X, Parton, RG, Geli, MI, Scita, G, Arroyo, M, Le Roux, AL, Roca-Cusachs, P, (2023). A mechanosensing mechanism controls plasma membrane shape homeostasis at the nanoscale Elife 12, e72316

As cells migrate and experience forces from their surroundings, they constantly undergo mechanical deformations which reshape their plasma membrane (PM). To maintain homeostasis, cells need to detect and restore such changes, not only in terms of overall PM area and tension as previously described, but also in terms of local, nanoscale topography. Here, we describe a novel phenomenon, by which cells sense and restore mechanically induced PM nanoscale deformations. We show that cell stretch and subsequent compression reshape the PM in a way that generates local membrane evaginations in the 100 nm scale. These evaginations are recognized by I-BAR proteins, which triggers a burst of actin polymerization mediated by Rac1 and Arp2/3. The actin polymerization burst subsequently re-flattens the evagination, completing the mechanochemical feedback loop. Our results demonstrate a new mechanosensing mechanism for PM shape homeostasis, with potential applicability in different physiological scenarios.© 2023, Quiroga et al.

JTD Keywords: arp2/3 complex, bar, bar proteins, cdc42, cells, domain, human, irsp53, membrane biophysics, mouse, proteins, rac, tension, Actin polymerization, Actins, Bar proteins, Cell biology, Cell membrane, Homeostasis, Human, Mechanobiology, Membrane biophysics, Mouse, Physics of living systems


Marín-Llauradó, A, Kale, S, Ouzeri, A, Golde, T, Sunyer, R, Torres-Sánchez, A, Latorre, E, Gómez-González, M, Roca-Cusachs, P, Arroyo, M, Trepat, X, (2023). Mapping mechanical stress in curved epithelia of designed size and shape Nature Communications 14, 4014

The function of organs such as lungs, kidneys and mammary glands relies on the three-dimensional geometry of their epithelium. To adopt shapes such as spheres, tubes and ellipsoids, epithelia generate mechanical stresses that are generally unknown. Here we engineer curved epithelial monolayers of controlled size and shape and map their state of stress. We design pressurized epithelia with circular, rectangular and ellipsoidal footprints. We develop a computational method, called curved monolayer stress microscopy, to map the stress tensor in these epithelia. This method establishes a correspondence between epithelial shape and mechanical stress without assumptions of material properties. In epithelia with spherical geometry we show that stress weakly increases with areal strain in a size-independent manner. In epithelia with rectangular and ellipsoidal cross-section we find pronounced stress anisotropies that impact cell alignment. Our approach enables a systematic study of how geometry and stress influence epithelial fate and function in three-dimensions.© 2023. The Author(s).

JTD Keywords: cell, forces, morphogenesis, tension, E-cadherin, Epithelial cells, Epithelium, Microscopy, Stress, mechanical


Abenza, JF, Rossetti, L, Mouelhi, M, Burgués, J, Andreu, I, Kennedy, K, Roca-Cusachs, P, Marco, S, García-Ojalvo, J, Trepat, X, (2023). Mechanical control of the mammalian circadian clock via YAP/TAZ and TEAD Journal Of Cell Biology 222, e202209120

Autonomous circadian clocks exist in nearly every mammalian cell type. These cellular clocks are subjected to a multilayered regulation sensitive to the mechanochemical cell microenvironment. Whereas the biochemical signaling that controls the cellular circadian clock is increasingly well understood, mechanisms underlying regulation by mechanical cues are largely unknown. Here we show that the fibroblast circadian clock is mechanically regulated through YAP/TAZ nuclear levels. We use high-throughput analysis of single-cell circadian rhythms and apply controlled mechanical, biochemical, and genetic perturbations to study the expression of the clock gene Rev-erbα. We observe that Rev-erbα circadian oscillations are disrupted with YAP/TAZ nuclear translocation. By targeted mutations and overexpression of YAP/TAZ, we show that this mechanobiological regulation, which also impacts core components of the clock such as Bmal1 and Cry1, depends on the binding of YAP/TAZ to the transcriptional effector TEAD. This mechanism could explain the impairment of circadian rhythms observed when YAP/TAZ activity is upregulated, as in cancer and aging.© 2023 Abenza et al.

JTD Keywords: activation, dynamics, forces, growth, hippo pathway, liver, platform, time, transcription, Animals, Circadian clocks, Circadian rhythm, Gene-expression, Mammals, Signal transduction, Tea domain transcription factors, Transcriptional coactivator with pdz-binding motif proteins, Yap-signaling proteins


Pesce, M, Duda, GN, Forte, G, Girao, H, Raya, A, Roca-Cusachs, P, Sluijter, JPG, Tschöpe, C, Van Linthout, S, (2023). Cardiac fibroblasts and mechanosensation in heart development, health and disease Nature Reviews Cardiology 20, 309-324

The term 'mechanosensation' describes the capacity of cells to translate mechanical stimuli into the coordinated regulation of intracellular signals, cellular function, gene expression and epigenetic programming. This capacity is related not only to the sensitivity of the cells to tissue motion, but also to the decryption of tissue geometric arrangement and mechanical properties. The cardiac stroma, composed of fibroblasts, has been historically considered a mechanically passive component of the heart. However, the latest research suggests that the mechanical functions of these cells are an active and necessary component of the developmental biology programme of the heart that is involved in myocardial growth and homeostasis, and a crucial determinant of cardiac repair and disease. In this Review, we discuss the general concept of cell mechanosensation and force generation as potent regulators in heart development and pathology, and describe the integration of mechanical and biohumoral pathways predisposing the heart to fibrosis and failure. Next, we address the use of 3D culture systems to integrate tissue mechanics to mimic cardiac remodelling. Finally, we highlight the potential of mechanotherapeutic strategies, including pharmacological treatment and device-mediated left ventricular unloading, to reverse remodelling in the failing heart.© 2022. Springer Nature Limited.

JTD Keywords: cardiomyocyte proliferation, cross-linking, extracellular-matrix, focal adhesions, gene-expression, mechanical regulation, myocardial-infarction, substrate stiffness affects, t-cells, Ventricular assist device


Alegre-Cebollada, J, Huerta-Lopez, C, Clemente-Manteca, A, Velazquez-Carreras, D, Espinosa, FM, Saez, P, Martinez-del-Pozo, A, Garcia-Garcia, M, Martin-Colomo, S, Garcia, R, Roca-Cusachs, P, Elosegui-Artola, A, del Pozo, MA, Herrero-Galán, E, Plaza, GR, (2023). Cell response to substrate energy dissipation outweighs rigidity sensing Biophysical Journal 122, 292A-292A

Pallares, ME, Pi-Jauma, I, Fortunato, IC, Grazu, V, Gomez-Gonzalez, M, Roca-Cusachs, P, de la Fuente, JM, Alert, R, Sunyer, R, Casademunt, J, Trepat, X, (2023). Stiffness-dependent active wetting enables optimal collective cell durotaxis Nature Physics 19, 279-289

The directed migration of cellular clusters enables morphogenesis, wound healing and collective cancer invasion. Gradients of substrate stiffness direct the migration of cellular clusters in a process called collective durotaxis, but the underlying mechanisms remain unclear. Here we unveil a connection between collective durotaxis and the wetting properties of cellular clusters. We show that clusters of cancer cells dewet soft substrates and wet stiff ones. At intermediate stiffness-at the crossover from low to high wettability-clusters on uniform-stiffness substrates become maximally motile, and clusters on stiffness gradients exhibit optimal durotaxis. Durotactic velocity increases with cluster size, stiffness gradient and actomyosin activity. We demonstrate this behaviour on substrates coated with the cell-cell adhesion protein E-cadherin and then establish its generality on substrates coated with extracellular matrix. We develop an active wetting model that explains collective durotaxis in terms of a balance between in-plane active traction and tissue contractility and out-of-plane surface tension. Finally, we show that the distribution of cluster displacements has a heavy tail, with infrequent but large cellular hops that contribute to durotactic migration. Our study demonstrates a physical mechanism of collective durotaxis, through both cell-cell and cell-substrate adhesion ligands, based on the wetting properties of active droplets.

JTD Keywords: Adhesion, Dynamics, E-cadherin, Gradient, Migration, Model, Motility, Movements, Rigidity, Substrate stiffness


Lerche, M, Mathieu, M, Malerod, L, Pedersen, N, Hamidi, H, Chastney, M, Bruininks, B, Kaptan, S, Jacquemet, G, Vattulainen, I, Roca-Cusachs, P, Brech, A, Perez, F, Boncompain, G, Miserey, S, Ivaska, J, (2023). Regulation of Cell Dynamics by Rapid Transport of Integrins Through the Biosynthetic Pathway (MS21) Molecular Biology Of The Cell 34, 327-327

Roca-Cusachs, P, (2023). Transducing - and shielding - mechanical signals from integrins to the nucleus (SG123) Molecular Biology Of The Cell 34, 98-98

Sunyer, R, Pallares, ME, Pi-Jaumà, I, Fortunato, IC, Grazú, V, Gómez-González, M, Roca-Cusachs, P, de la Furente, JM, Alert, R, Casademunt, J, Trepat, X, (2023). Optimal collective durotaxis through E-cadherin adhesions Molecular Biology Of The Cell 34, 303-304

Lerche, M, Mathieu, M, Malerod, L, Pedersen, N, Hamidi, H, Chastney, M, Bruininks, B, Kaptan, S, Jacquemet, G, Vattulainen, I, Roca-Cusachs, P, Brech, A, Perez, F, Boncompain, G, Miserey, S, Ivaska, J, (2023). Regulation of Cell Dynamics by Rapid Transport of Integrins Through the Biosynthetic Pathway (MS21) Molecular Biology Of The Cell 34, 309-309

Lolo, FN, Walani, N, Seemann, E, Zalvidea, D, Pavón, DM, Cojoc, G, Zamai, M, de Lesegno, CV, de Benito, FM, Sánchez-Alvarez, M, Uriarte, JJ, Echarri, A, Jiménez-Carretero, D, Escolano, JC, Sánchez, SA, Caiolfa, VR, Navajas, D, Trepat, X, Guck, J, Lamaze, C, Roca-Cusachs, P, Kessels, MM, Qualmann, B, Arroyo, M, Del Pozo, MA, (2023). Caveolin-1 dolines form a distinct and rapid caveolae-independent mechanoadaptation system Nature Cell Biology 25, 120-133

In response to different types and intensities of mechanical force, cells modulate their physical properties and adapt their plasma membrane (PM). Caveolae are PM nano-invaginations that contribute to mechanoadaptation, buffering tension changes. However, whether core caveolar proteins contribute to PM tension accommodation independently from the caveolar assembly is unknown. Here we provide experimental and computational evidence supporting that caveolin-1 confers deformability and mechanoprotection independently from caveolae, through modulation of PM curvature. Freeze-fracture electron microscopy reveals that caveolin-1 stabilizes non-caveolar invaginations-dolines-capable of responding to low-medium mechanical forces, impacting downstream mechanotransduction and conferring mechanoprotection to cells devoid of caveolae. Upon cavin-1/PTRF binding, doline size is restricted and membrane buffering is limited to relatively high forces, capable of flattening caveolae. Thus, caveolae and dolines constitute two distinct albeit complementary components of a buffering system that allows cells to adapt efficiently to a broad range of mechanical stimuli.© 2022. The Author(s).

JTD Keywords: cavin, cell-migration, cholesterol, extracellular-matrix, nanoscale organization, particle-size, polarization, size distribution, tension, Plasma-membrane


Zambarda, C, Gonzalez, CP, Schoenit, A, Veits, N, Schimmer, C, Jung, RM, Ollech, D, Christian, J, Roca-Cusachs, P, Trepat, X, Cavalcanti-Adam, EA, (2022). Epithelial cell cluster size affects force distribution in response to EGF-induced collective contractility European Journal Of Cell Biology 101, 151274

Several factors present in the extracellular environment regulate epithelial cell adhesion and dynamics. Among them, growth factors such as EGF, upon binding to their receptors at the cell surface, get internalized and directly activate the acto-myosin machinery. In this study we present the effects of EGF on the contractility of epithelial cancer cell colonies in confined geometry of different sizes. We show that the extent to which EGF triggers contractility scales with the cluster size and thus the number of cells. Moreover, the collective contractility results in a radial distribution of traction forces, which are dependent on integrin β1 peripheral adhesions and transmitted to neighboring cells through adherens junctions. Taken together, EGF-induced contractility acts on the mechanical crosstalk and linkage between the cell-cell and cell-matrix compartments, regulating collective responses.Copyright © 2022 The Authors. Published by Elsevier GmbH.. All rights reserved.

JTD Keywords: actin, activation, actomyosin, adherens junctions, adhesion, e-cadherin, egf, maturation, mechanical regulation, micropatterning, migration, traction forces, transduction, transmission, Actomyosin, Adherens junctions, Cell adhesion, Cell membrane, Collective contractility, Egf, Epidermal growth factor, Epidermal-growth-factor, Epithelial cells, Micropatterning, Myosins, Traction forces


Lolo, FN, Pavón, DM, Grande, A, Artola, AE, Segatori, VI, Sánchez, S, Trepat, X, Roca-Cusachs, P, del Pozo, MA, (2022). Caveolae couple mechanical stress to integrin recycling and activation Elife 11, e82348

Cells are subjected to multiple mechanical inputs throughout their lives. Their ability to detect these environmental cues is called mechanosensing, a process in which integrins play an important role. During cellular mechanosensing, plasma membrane (PM) tension is adjusted to mechanical stress through the buffering action of caveolae; however, little is known about the role of caveolae in early integrin mechanosensing regulation. Here, we show that Cav1KO fibroblasts increase adhesion to FN-coated beads when pulled with magnetic tweezers, as compared to wild type fibroblasts. This phenotype is Rho-independent and mainly derived from increased active b1-integrin content on the surface of Cav1KO fibroblasts. FRAP analysis and endocytosis/recycling assays revealed that active b1-integrin is mostly endocytosed through the CLIC/GEEC pathway and is more rapidly recycled to the PM in Cav1KO fibroblasts, in a Rab4 and PM tension-dependent manner. Moreover, the threshold for PM tension-driven b1-integrin activation is lower in Cav1KO MEFs than in wild type MEFs, through a mechanism dependent on talin activity. Our findings suggest that caveolae couple mechanical stress to integrin cycling and activation, thereby regulating the early steps of the cellular mechanosensing response.© 2022, Lolo et al.

JTD Keywords: adhesion, alpha-v-beta-3, cell, integrin activation, internalization, kinase, mechanosensing, mediated endocytosis, mouse, stiffness, talin, trafficking, Animals, Caveolae, Cell adhesion, Cell biology, Fibroblasts, Integrin activation, Integrin beta1, Integrin recycling, Integrins, Mechanosensing, Membrane tension, Mice, Mouse, Stress, mechanical


Barbacena, P, Dominguez-Cejudo, M, Fonseca, CG, Gómez-González, M, Faure, LM, Zarkada, G, Pena, A, Pezzarossa, A, Ramalho, D, Giarratano, Y, Ouarné, M, Barata, D, Fortunato, IC, Misikova, LH, Mauldin, I, Carvalho, Y, Trepat, X, Roca-Cusachs, P, Eichmann, A, Bernabeu, MO, Franco, CA, (2022). Competition for endothelial cell polarity drives vascular morphogenesis in the mouse retina Developmental Cell 57, 2321-2333

Blood-vessel formation generates unique vascular patterns in each individual. The principles governing the apparent stochasticity of this process remain to be elucidated. Using mathematical methods, we find that the transition between two fundamental vascular morphogenetic programs-sprouting angiogenesis and vascular remodeling-is established by a shift of collective front-to-rear polarity of endothelial cells in the mouse retina. We demonstrate that the competition between biochemical (VEGFA) and mechanical (blood-flow-induced shear stress) cues controls this collective polarity shift. Shear stress increases tension at focal adhesions overriding VEGFA-driven collective polarization, which relies on tension at adherens junctions. We propose that vascular morphogenetic cues compete to regulate individual cell polarity and migration through tension shifts that translates into tissue-level emergent behaviors, ultimately leading to uniquely organized vascular patterns.Copyright © 2022 The Author(s). Published by Elsevier Inc. All rights reserved.

JTD Keywords: activation, angiogenesis, dynamics, flow, forces, image, mechanisms, vinculin, Angiogenesis, Cell polarity, Fluid shear, Mechanobiology, Morphogenesis, Shear stress


Phuyal, S, Djaerff, E, Le Roux, AL, Baker, MJ, Fankhauser, D, Mahdizadeh, SJ, Reiterer, V, Parizadeh, A, Felder, E, Kahlhofer, JC, Teis, D, Kazanietz, MG, Geley, S, Eriksson, L, Roca-Cusachs, P, Farhan, H, (2022). Mechanical strain stimulates COPII-dependent secretory trafficking via Rac1 Embo Journal 41, e110596

Cells are constantly exposed to various chemical and physical stimuli. While much has been learned about the biochemical factors that regulate secretory trafficking from the endoplasmic reticulum (ER), much less is known about whether and how this trafficking is subject to regulation by mechanical signals. Here, we show that subjecting cells to mechanical strain both induces the formation of ER exit sites (ERES) and accelerates ER-to-Golgi trafficking. We found that cells with impaired ERES function were less capable of expanding their surface area when placed under mechanical stress and were more prone to develop plasma membrane defects when subjected to stretching. Thus, coupling of ERES function to mechanotransduction appears to confer resistance of cells to mechanical stress. Furthermore, we show that the coupling of mechanotransduction to ERES formation was mediated via a previously unappreciated ER-localized pool of the small GTPase Rac1. Mechanistically, we show that Rac1 interacts with the small GTPase Sar1 to drive budding of COPII carriers and stimulates ER-to-Golgi transport. This interaction therefore represents an unprecedented link between mechanical strain and export from the ER.© 2022 The Authors. Published under the terms of the CC BY 4.0 license.

JTD Keywords: cells, copii, docking, endoplasmic reticulum, endoplasmic-reticulum, er, gtpase, mechanobiology, proliferation, protein, reticulum exit sites, web server, Copii, Fast interaction refinement, Mechanobiology


Faure, LM, Roca-Cusachs, P, (2022). Ruffled in water, smooth in honey Nature Physics 18, 966-967

Cells can sense the mechanical properties of their environment. By adjusting the ruffling of their membranes, cells respond to different viscosities of their surrounding liquid medium.

JTD


Beedle, AEM, Roca-Cusachs, P, (2022). In search of a softer environment Nature Materials 21, 995-996

By maximizing cell-substrate force transmission, cancer cells can migrate towards either stiffer or softer substrate regions.

JTD Keywords: Durotaxis, Stiffness


Andreu, I, Granero-Moya, I, Chahare, NR, Clein, K, Molina-Jordan, M, Beedle, AEM, Elosegui-Artola, A, Abenza, JF, Rossetti, L, Trepat, X, Raveh, B, Roca-Cusachs, P, (2022). Mechanical force application to the nucleus regulates nucleocytoplasmic transport Nature Cell Biology 24, 896-905

Mechanical force controls fundamental cellular processes in health and disease, and increasing evidence shows that the nucleus both experiences and senses applied forces. Such forces can lead to the nuclear translocation of proteins, but whether force controls nucleocytoplasmic transport, and how, remains unknown. Here we show that nuclear forces differentially control passive and facilitated nucleocytoplasmic transport, setting the rules for the mechanosensitivity of shuttling proteins. We demonstrate that nuclear force increases permeability across nuclear pore complexes, with a dependence on molecular weight that is stronger for passive than for facilitated diffusion. Owing to this differential effect, force leads to the translocation of cargoes into or out of the nucleus within a given range of molecular weight and affinity for nuclear transport receptors. Further, we show that the mechanosensitivity of several transcriptional regulators can be both explained by this mechanism and engineered exogenously by introducing appropriate nuclear localization signals. Our work unveils a mechanism of mechanically induced signalling, probably operating in parallel with others, with potential applicability across signalling pathways.; Andreu et al. show that force regulates nucleocytoplasmic transport by weakening the permeability barrier of nuclear pore complexes, affecting passive and facilitated diffusion in different ways.

JTD Keywords: Activation, Inhibitor, Matrix, Mechanotransduction, Nesprins, Nucleoporins, Permeability barrier, Pore complex, Proteins, Transmission


Andreu, I, Granero-Moya, I, Garcia-Manyes, S, Roca-Cusachs, P, (2022). Understanding the role of mechanics in nucleocytoplasmic transport Apl Bioengineering 6, 20901

Cell nuclei are submitted to mechanical forces, which in turn affect nuclear and cell functions. Recent evidence shows that a crucial mechanically regulated nuclear function is nucleocytoplasmic transport, mediated by nuclear pore complexes (NPCs). Mechanical regulation occurs at two levels: first, by force application to the nucleus, which increases NPC permeability likely through NPC stretch. Second, by the mechanical properties of the transported proteins themselves, as mechanically labile proteins translocate through NPCs faster than mechanically stiff ones. In this perspective, we discuss this evidence and the associated mechanisms by which mechanics can regulate the nucleo-cytoplasmic partitioning of proteins. Finally, we analyze how mechanical regulation of nucleocytoplasmic transport can provide a systematic approach to the study of mechanobiology and open new avenues both in fundamental and applied research. (C) 2022 Author(s).

JTD Keywords: Architecture, Association, Force, Nuclear-pore complex, Pathways, Protein import, Sun1


Rätze, MAK, Koorman, T, Sijnesael, T, Bassey-Archibong, B, van de Ven, R, Enserink, L, Visser, D, Jaksani, S, Viciano, I, Bakker, ERM, Richard, F, Tutt, A, O'Leary, L, Fitzpatrick, A, Roca-Cusachs, P, van Diest, PJ, Desmedt, C, Daniel, JM, Isacke, CM, Derksen, PWB, (2022). Loss of E-cadherin leads to Id2-dependent inhibition of cell cycle progression in metastatic lobular breast cancer Oncogene 41, 2932-2944

Invasive lobular breast carcinoma (ILC) is characterized by proliferative indolence and long-term latency relapses. This study aimed to identify how disseminating ILC cells control the balance between quiescence and cell cycle re-entry. In the absence of anchorage, ILC cells undergo a sustained cell cycle arrest in G0/G1 while maintaining viability. From the genes that are upregulated in anchorage independent ILC cells, we selected Inhibitor of DNA binding 2 (Id2), a mediator of cell cycle progression. Using loss-of-function experiments, we demonstrate that Id2 is essential for anchorage independent survival (anoikis resistance) in vitro and lung colonization in mice. Importantly, we find that under anchorage independent conditions, E-cadherin loss promotes expression of Id2 in multiple mouse and (organotypic) human models of ILC, an event that is caused by a direct p120-catenin/Kaiso-dependent transcriptional de-repression of the canonical Kaiso binding sequence TCCTGCNA. Conversely, stable inducible restoration of E-cadherin expression in the ILC cell line SUM44PE inhibits Id2 expression and anoikis resistance. We show evidence that Id2 accumulates in the cytosol, where it induces a sustained and CDK4/6-dependent G0/G1 cell cycle arrest through interaction with hypo-phosphorylated Rb. Finally, we find that Id2 is indeed enriched in ILC when compared to other breast cancers, and confirm cytosolic Id2 protein expression in primary ILC samples. In sum, we have linked mutational inactivation of E-cadherin to direct inhibition of cell cycle progression. Our work indicates that loss of E-cadherin and subsequent expression of Id2 drive indolence and dissemination of ILC. As such, E-cadherin and Id2 are promising candidates to stratify low and intermediate grade invasive breast cancers for the use of clinical cell cycle intervention drugs.

JTD Keywords: anoikis resistance, carcinoma, d1, differentiation, gene-expression, growth, id2, proliferation, repression, Mammary epithelial-cells


Estaje, EB, Dalby, M, Roca-Cusachs, P, Salmeron-Sanchez, M, Cantini, M, (2022). SURFACE VISCOSITY CONTROLS STEM CELL FATE (Abstract 1319) Tissue Engineering Part a 28, S369-S369

The microenvironment where cells reside provides them with arange of cues that collectively affect their fate. These include bio-chemical cues, mediated by interactions with the extracellular matrix(via integrins) and between cells (via cadherins), and physicalproperties, such as stiffness and viscosity. While the effect of stiff-ness on cell fate has been widely studied, the role of viscosity hasbeen only recently addressed (1). In our group, we demonstrated thatcells sense viscosity using the same mechanotransductive mecha-nisms that they use for stiffness (2). Here, we investigate the role ofviscous interactions in controlling stem cell fate. We designed mo-bile surfaces based on supported lipid bilayers of controlled viscos-ity, which target the adhesive crosstalk between integrins (RGDreceptors) and cadherins (HAVDI-containing proteins) (3).Using human mesenchymal stem cells, we observed an increase incell adhesion and a decrease in actin flow when the viscosity of thesurface and the concentration of RGD increased. When HAVDI wasadded to the surface, cell adhesion was reduced, while the actin flowincreased. These changes provoked alterations in the activation ofmechanotransductive pathways: the nuclear translocation of me-chanosensitive YAP increased with viscosity and decreased withHAVDI ligation. Finally, early differentiation markers were differ-entially expressed, revealing an effect of viscosity on cell fate: whilemore mobile surfaces favored adipogenesis and HAVDI ligation onthem triggered chondrogenesis, more viscous substrates promotedosteogenesis. Further investigations on the signaling pathways in-volved will allow to establish a paradigm to collectively understandand exploit cell response to viscous interactions

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Le Roux, AL, Tozzi, C, Walani, N, Quiroga, X, Zalvidea, D, Trepat, X, Staykova, M, Arroyo, M, Roca-Cusachs, P, (2021). Dynamic mechanochemical feedback between curved membranes and BAR protein self-organization Nature Communications 12, 6550

In many physiological situations, BAR proteins reshape membranes with pre-existing curvature (templates), contributing to essential cellular processes. However, the mechanism and the biological implications of this reshaping process remain unclear. Here we show, both experimentally and through modelling, that BAR proteins reshape low curvature membrane templates through a mechanochemical phase transition. This phenomenon depends on initial template shape and involves the co-existence and progressive transition between distinct local states in terms of molecular organization (protein arrangement and density) and membrane shape (template size and spherical versus cylindrical curvature). Further, we demonstrate in cells that this phenomenon enables a mechanotransduction mode, in which cellular stretch leads to the mechanical formation of membrane templates, which are then reshaped into tubules by BAR proteins. Our results demonstrate the interplay between membrane mechanics and BAR protein molecular organization, integrating curvature sensing and generation in a comprehensive framework with implications for cell mechanical responses.

JTD Keywords: aggregation, amphiphysin, domains, vesicles, Article, Cell, Cell component, Curvature, Detection method, Geomembrane, Mechanotransduction, Membrane, Molecular analysis, Phase transition, Physiology, Protein, Self organization


Andreu, I, Falcones, B, Hurst, S, Chahare, N, Quiroga, X, Le Roux, AL, Kechagia, Z, Beedle, AEM, Elosegui-Artola, A, Trepat, X, Farre, R, Betz, T, Almendros, I, Roca-Cusachs, P, (2021). The force loading rate drives cell mechanosensing through both reinforcement and cytoskeletal softening Nature Communications 12, 4229

Cell response to force regulates essential processes in health and disease. However, the fundamental mechanical variables that cells sense and respond to remain unclear. Here we show that the rate of force application (loading rate) drives mechanosensing, as predicted by a molecular clutch model. By applying dynamic force regimes to cells through substrate stretching, optical tweezers, and atomic force microscopy, we find that increasing loading rates trigger talin-dependent mechanosensing, leading to adhesion growth and reinforcement, and YAP nuclear localization. However, above a given threshold the actin cytoskeleton softens, decreasing loading rates and preventing reinforcement. By stretching rat lungs in vivo, we show that a similar phenomenon may occur. Our results show that cell sensing of external forces and of passive mechanical parameters (like tissue stiffness) can be understood through the same mechanisms, driven by the properties under force of the mechanosensing molecules involved. Cells sense mechanical forces from their environment, but the precise mechanical variable sensed by cells is unclear. Here, the authors show that cells can sense the rate of force application, known as the loading rate, with effects on YAP nuclear localization and cytoskeletal stiffness remodelling.

JTD Keywords: Actin cytoskeleton, Actin filament, Actin-filament, Adhesion, Animal, Animals, Atomic force microscopy, Breathing, Cell, Cell adhesion, Cell culture, Cell nucleus, Cells, cultured, Cytoplasm, Extracellular-matrix, Fibroblast, Fibroblasts, Fibronectin, Frequency, Gene knockdown, Gene knockdown techniques, Genetics, Germfree animal, Integrin, Intracellular signaling peptides and proteins, Knockout mouse, Lung, Male, Mechanotransduction, Mechanotransduction, cellular, Metabolism, Mice, Mice, knockout, Microscopy, atomic force, Mouse, Optical tweezers, Paxillin, Physiology, Primary cell culture, Pxn protein, mouse, Rat, Rats, Rats, sprague-dawley, Respiration, Signal peptide, Softening, Specific pathogen-free organisms, Sprague dawley rat, Stress, Substrate, Substrate rigidity, Talin, Talin protein, mouse, Tln2 protein, mouse, Traction, Transmission, Ultrastructure, Yap1 protein, rat


Pérez-González, C, Ceada, G, Greco, F, Matejcic, M, Gómez-González, M, Castro, N, Menendez, A, Kale, S, Krndija, D, Clark, AG, Gannavarapu, VR, Alvarez-Varela, A, Roca-Cusachs, P, Batlle, E, Vignjevic, DM, Arroyo, M, Trepat, X, (2021). Mechanical compartmentalization of the intestinal organoid enables crypt folding and collective cell migration Nature Cell Biology 23, 745-757

Intestinal organoids capture essential features of the intestinal epithelium such as crypt folding, cellular compartmentalization and collective movements. Each of these processes and their coordination require patterned forces that are at present unknown. Here we map three-dimensional cellular forces in mouse intestinal organoids grown on soft hydrogels. We show that these organoids exhibit a non-monotonic stress distribution that defines mechanical and functional compartments. The stem cell compartment pushes the extracellular matrix and folds through apical constriction, whereas the transit amplifying zone pulls the extracellular matrix and elongates through basal constriction. The size of the stem cell compartment depends on the extracellular-matrix stiffness and endogenous cellular forces. Computational modelling reveals that crypt shape and force distribution rely on cell surface tensions following cortical actomyosin density. Finally, cells are pulled out of the crypt along a gradient of increasing tension. Our study unveils how patterned forces enable compartmentalization, folding and collective migration in the intestinal epithelium. Perez-Gonzalez et al. explore the mechanical properties of intestinal organoids, and report the existence of distinct mechanical domains and that cells are pulled out of the central crypt along a gradient of increasing tension.

JTD Keywords: Forces, Growth, Gut, Monolayers, Morphogenesis, Reveal, Stem-cells, Tension


Tozzi, C, Walani, N, Le Roux, AL, Roca-Cusachs, P, Arroyo, M, (2021). A theory of ordering of elongated and curved proteins on membranes driven by density and curvature Soft Matter 17, 3367-3379

Cell membranes interact with a myriad of curvature-active proteins that control membrane morphology and are responsible for mechanosensation and mechanotransduction. Some of these proteins, such as those containing BAR domains, are curved and elongated, and hence may adopt different states of orientational order, from isotropic to maximize entropy to nematic as a result of crowding or to adapt to the curvature of the underlying membrane. Here, extending the classical work of Onsager for ordering in hard particle systems and that of [E. S. Nascimento et al., Phys. Rev. E, 2017, 96, 022704], we develop a mean-field density functional theory to predict the orientational order and evaluate the free energy of ensembles of elongated and curved objects on curved membranes. This theory depends on the microscopic properties of the particles and explains how a density-dependent isotropic-to-nematic transition is modified by anisotropic curvature. We also examine the coexistence of isotropic and nematic phases. This theory predicts how ordering depends on geometry but we assume here that the geometry is fixed. It also lays the ground to understand the interplay between membrane reshaping by BAR proteins and molecular order, examined by [Le Roux et al., submitted, 2020]. This journal is

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Lerche, Martina, Elosegui-Artola, Alberto, Kechagia, Jenny Z., Guzmán, Camilo, Georgiadou, Maria, Andreu, Ion, Gullberg, Donald, Roca-Cusachs, Pere, Peuhu, Emilia, Ivaska, Johanna, (2020). Integrin binding dynamics modulate ligand-specific mechanosensing in mammary gland fibroblasts iScience 23, (3), 100907

The link between integrin activity regulation and cellular mechanosensing of tissue rigidity, especially on different extracellular matrix ligands, remains poorly understood. Here, we find that primary mouse mammary gland stromal fibroblasts (MSFs) are able to spread efficiently, generate high forces, and display nuclear YAP on soft collagen-coated substrates, resembling the soft mammary gland tissue. We describe that loss of the integrin inhibitor, SHARPIN, impedes MSF spreading specifically on soft type I collagen but not on fibronectin. Through quantitative experiments and computational modeling, we find that SHARPIN-deficient MSFs display faster force-induced unbinding of adhesions from collagen-coated beads. Faster unbinding, in turn, impairs force transmission in these cells, particularly, at the stiffness optimum observed for wild-type cells. Mechanistically, we link the impaired mechanotransduction of SHARPIN-deficient cells on collagen to reduced levels of collagen-binding integrin α11β1. Thus integrin activity regulation and α11β1 play a role in collagen-specific mechanosensing in MSFs.

JTD Keywords: Biological Sciences, Cell Biology, Functional Aspects of Cell Biology


Guixé-Muntet, Sergi, Ortega-Ribera, Martí, Wang, Cong, Selicean, Sonia, Andreu, Ion, Kechagia, Jenny Z., Fondevila, Constantino, Roca-Cusachs, Pere, Dufour, Jean-François, Bosch, Jaime, Berzigotti, Annalisa, Gracia-Sancho, Jordi, (2020). Nuclear deformation mediates liver cell mechanosensing in cirrhosis JHEP Reports 2, (5), 100145

Background & AimsLiver stiffness is increased in advanced chronic liver disease (ACLD) and accurately predicts prognosis in this population. Recent data suggest that extracellular matrix stiffness per se may modulate the phenotype of liver cells. We aimed at investigating the effect of matrix stiffness on the phenotype of liver cells of rats with cirrhosis, assessing its influence on their response to antifibrotic strategies and evaluating associated molecular mechanisms.

JTD Keywords: Chronic liver disease, Hepatocyte, HSC, LSEC, Stiffness


Kechagia, Jenny Z., Ivaska, Johanna, Roca-Cusachs, Pere, (2019). Integrins as biomechanical sensors of the microenvironment Nature Reviews Molecular Cell Biology 20, (8), 457-473

Integrins, and integrin-mediated adhesions, have long been recognized to provide the main molecular link attaching cells to the extracellular matrix (ECM) and to serve as bidirectional hubs transmitting signals between cells and their environment. Recent evidence has shown that their combined biochemical and mechanical properties also allow integrins to sense, respond to and interact with ECM of differing properties with exquisite specificity. Here, we review this work first by providing an overview of how integrin function is regulated from both a biochemical and a mechanical perspective, affecting integrin cell-surface availability, binding properties, activation or clustering. Then, we address how this biomechanical regulation allows integrins to respond to different ECM physicochemical properties and signals, such as rigidity, composition and spatial distribution. Finally, we discuss the importance of this sensing for major cell functions by taking cell migration and cancer as examples.

JTD Keywords: Cell migration, Extracellular matrix, Integrins, Mechanotransduction, Single-molecule biophysics


Garreta, Elena, Prado, Patricia, Tarantino, Carolina, Oria, Roger, Fanlo, Lucia, Martí, Elisa, Zalvidea, Dobryna, Trepat, Xavier, Roca-Cusachs, Pere, Gavaldà -Navarro, Aleix, Cozzuto, Luca, Campistol, Josep M., Izpisúa Belmonte, Juan Carlos, Hurtado del Pozo, Carmen, Montserrat, Nuria, (2019). Fine tuning the extracellular environment accelerates the derivation of kidney organoids from human pluripotent stem cells Nature Materials 18, 397-405

The generation of organoids is one of the biggest scientific advances in regenerative medicine. Here, by lengthening the time that human pluripotent stem cells (hPSCs) were exposed to a three-dimensional microenvironment, and by applying defined renal inductive signals, we generated kidney organoids that transcriptomically matched second-trimester human fetal kidneys. We validated these results using ex vivo and in vitro assays that model renal development. Furthermore, we developed a transplantation method that utilizes the chick chorioallantoic membrane. This approach created a soft in vivo microenvironment that promoted the growth and differentiation of implanted kidney organoids, as well as providing a vascular component. The stiffness of the in ovo chorioallantoic membrane microenvironment was recapitulated in vitro by fabricating compliant hydrogels. These biomaterials promoted the efficient generation of renal vesicles and nephron structures, demonstrating that a soft environment accelerates the differentiation of hPSC-derived kidney organoids.

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Uroz, Marina, Garcia-Puig, Anna, Tekeli, Isil, Elosegui-Artola, Alberto, Abenza, Juan F., Marín-Llauradó, Ariadna, Pujals, Silvia, Conte, Vito, Albertazzi, Lorenzo, Roca-Cusachs, Pere, Raya, Ángel, Trepat, Xavier, (2019). Traction forces at the cytokinetic ring regulate cell division and polyploidy in the migrating zebrafish epicardium Nature Materials 18, 1015-1023

Epithelial repair and regeneration are driven by collective cell migration and division. Both cellular functions involve tightly controlled mechanical events, but how physical forces regulate cell division in migrating epithelia is largely unknown. Here we show that cells dividing in the migrating zebrafish epicardium exert large cell–extracellular matrix (ECM) forces during cytokinesis. These forces point towards the division axis and are exerted through focal adhesions that connect the cytokinetic ring to the underlying ECM. When subjected to high loading rates, these cytokinetic focal adhesions prevent closure of the contractile ring, leading to multi-nucleation through cytokinetic failure. By combining a clutch model with experiments on substrates of different rigidity, ECM composition and ligand density, we show that failed cytokinesis is triggered by adhesion reinforcement downstream of increased myosin density. The mechanical interaction between the cytokinetic ring and the ECM thus provides a mechanism for the regulation of cell division and polyploidy that may have implications in regeneration and cancer.

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Infante, Elvira, Stannard, Andrew, Board, Stephanie J., Rico-Lastres, Palma, Rostkova, Elena, Beedle, Amy E. M., Lezamiz, Ainhoa, Wang, Yong Jian, Gulaidi Breen, Samuel, Panagaki, Fani, Sundar Rajan, Vinoth, Shanahan, Catherine, Roca-Cusachs, Pere, Garcia-Manyes, Sergi, (2019). The mechanical stability of proteins regulates their translocation rate into the cell nucleus Nature Physics 15, 973-981

A cell’s ability to react to mechanical stimuli is known to be affected by the transport of transcription factors, the proteins responsible for regulating transcription of DNA into RNA, across the membrane enveloping its nucleus. Yet the molecular mechanisms by which mechanical cues control this process remain unclear. Here we show that one such protein, myocardin-related transcription factor A (MRTFA), is imported into the nucleus at a rate that is inversely correlated with its nanomechanical stability, but independent of its thermodynamic stability. Attaching mechanically stable proteins to MRTFA results in reduced gene expression and the subsequent slowing down of cell migration. We conclude that the mechanical unfolding of proteins regulates their nuclear translocation rate, and highlight the role of the nuclear pore complex as a selective mechanosensor that is capable of detecting forces as low as 10 pN. The modulation of the mechanical stability of transcription factors may represent a general strategy for the control of gene expression.

JTD Keywords: Biological physics, Biophysics, Chemistry, Nanoscience and technology


Roux, Anabel-Lise Lee, Quiroga, Xarxa, Walani, Nikhil, Arroyo, Marino, Roca-Cusachs, Pere, (2019). The plasma membrane as a mechanochemical transducer Philosophical Transactions of the Royal Society B: Biological Sciences 374, (1779), 20180221

Cells are constantly submitted to external mechanical stresses, which they must withstand and respond to. By forming a physical boundary between cells and their environment that is also a biochemical platform, the plasma membrane (PM) is a key interface mediating both cellular response to mechanical stimuli, and subsequent biochemical responses. Here, we review the role of the PM as a mechanosensing structure. We first analyse how the PM responds to mechanical stresses, and then discuss how this mechanical response triggers downstream biochemical responses. The molecular players involved in PM mechanochemical transduction include sensors of membrane unfolding, membrane tension, membrane curvature or membrane domain rearrangement. These sensors trigger signalling cascades fundamental both in healthy scenarios and in diseases such as cancer, which cells harness to maintain integrity, keep or restore homeostasis and adapt to their external environment.

JTD Keywords: Plasma membrane, Mechanotransduction, Membrane tension, Mechanosensor


Wickström, S. A., Roca-Cusachs, P., (2019). Special issue on “mechanotransduction in cell fate determination” – From molecular switches to organ-level regulation Experimental Cell Research 382, (1), 111452

Stereotypic formation of tissues requires coordination of cell fate with adhesive and cytoskeletal cues that control cell shape, positioning and motility. The importance of physical ques in shaping development has been demonstrated already a decade ago by observations that their removal arrests embryogenesis. Recent advances in technology and methods to quantify and experimentally manipulate adhesive and mechanical properties of cells and tissues has revolutionized the field. While many aspects of mechanical signaling have been covered by excellent reviews in the past, we invited experts in their respective fields of cell biology and biophysics to review the most recent, exciting breakthroughs on the role of mechanotransduction in cell fate regulation. Amongst these breakthroughs are the advancement of imaging techniques that have allowed single molecule resolution analyses of adhesive structures and quantitative analyses of the mechanical properties of the extracellular matrix and cells, the identification of nucleocytoplasmic transport and ion channels as key players in mechanosensitive cell fate decisions, and the interdisciplinary studies combining physics and biology to understand the role of tension in coupling single cell behaviors to changes in tissue state.

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Uroz, Marina, Wistorf, Sabrina, Serra-Picamal, Xavier, Conte, Vito, Sales-Pardo, Marta, Roca-Cusachs, Pere, Guimerà, Roger, Trepat, Xavier, (2018). Regulation of cell cycle progression by cell–cell and cell–matrix forces Nature Cell Biology 20, (6), 646-654

It has long been proposed that the cell cycle is regulated by physical forces at the cell–cell and cell–extracellular matrix (ECM) interfaces. However, the evolution of these forces during the cycle has never been measured in a tissue, and whether this evolution affects cell cycle progression is unknown. Here, we quantified cell–cell tension and cell–ECM traction throughout the complete cycle of a large cell population in a growing epithelium. These measurements unveil temporal mechanical patterns that span the entire cell cycle and regulate its duration, the G1–S transition and mitotic rounding. Cells subjected to higher intercellular tension exhibit a higher probability to transition from G1 to S, as well as shorter G1 and S–G2–M phases. Moreover, we show that tension and mechanical energy are better predictors of the duration of G1 than measured geometric properties. Tension increases during the cell cycle but decreases 3 hours before mitosis. Using optogenetic control of contractility, we show that this tension drop favours mitotic rounding. Our results establish that cell cycle progression is regulated cooperatively by forces between the dividing cell and its neighbours.

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Elosegui-Artola, Alberto, Trepat, Xavier, Roca-Cusachs, Pere, (2018). Control of mechanotransduction by molecular clutch dynamics Trends in Cell Biology 28, (5), 356-367

The linkage of cells to their microenvironment is mediated by a series of bonds that dynamically engage and disengage, in what has been conceptualized as the molecular clutch model. Whereas this model has long been employed to describe actin cytoskeleton and cell migration dynamics, it has recently been proposed to also explain mechanotransduction (i.e., the process by which cells convert mechanical signals from their environment into biochemical signals). Here we review the current understanding on how cell dynamics and mechanotransduction are driven by molecular clutch dynamics and its master regulator, the force loading rate. Throughout this Review, we place a specific emphasis on the quantitative prediction of cell response enabled by combined experimental and theoretical approaches.

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Thottacherry, Joseph Jose, Kosmalska, Anita Joanna, Elosegui-Artola, Alberto, Pradhan, Susav, Sharma, Sumit, Singh, Parvinder P., Guadamillas, Marta C., Chaudhary, Natasha, Vishwakarma, Ram, Trepat, Xavier, del Pozo, Miguel A., Parton, Robert G., Pullarkat, Pramod, Roca-Cusachs, Pere, Mayor, Satyajit, (2018). Mechanochemical feedback and control of endocytosis and membrane tension Nature Communications 9, 4217

Plasma membrane tension is an important factor that regulates many key cellular processes. Membrane trafficking is tightly coupled to membrane tension and can modulate the latter by addition or removal of the membrane. However, the cellular pathway(s) involved in these processes are poorly understood. Although a number of endocytic processes function simultaneously at the cell surface, we find that a dynamin and clathrin-independent pathway, the CLIC/GEEC (CG) pathway, is rapidly and specifically upregulated upon reduction of tension. On the other hand, inhibition of the CG pathway results in lower membrane tension, while up regulation significantly enhances membrane tension. We find that vinculin, a well-studied mechanotransducer, mediates the tension-dependent regulation of the CG pathway. Vinculin negatively regulates a key CG pathway regulator, GBF1, at the plasma membrane in a tension dependent manner. Thus, the CG pathway operates in a negative feedback loop with membrane tension which leads to a homeostatic regulation of membrane tension.

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Bennett, Mark, Cantini, Marco, Reboud, Julien, Cooper, Jonathan M., Roca-Cusachs, Pere, Salmeron-Sanchez, Manuel, (2018). Molecular clutch drives cell response to surface viscosity Proceedings of the National Academy of Sciences of the United States of America 115, (6), 1192-1197

Cell response to matrix rigidity has been explained by the mechanical properties of the actin-talin-integrin-fibronectin clutch. Here the molecular clutch model is extended to account for cell interactions with purely viscous surfaces (i.e., without an elastic component). Supported lipid bilayers present an idealized and controllable system through which to study this concept. Using lipids of different diffusion coefficients, the mobility (i.e., surface viscosity) of the presented ligands (in this case RGD) was altered by an order of magnitude. Cell size and cytoskeletal organization were proportional to viscosity. Furthermore, there was a higher number of focal adhesions and a higher phosphorylation of FAK on less-mobile (more-viscous) surfaces. Actin retrograde flow, an indicator of the force exerted on surfaces, was also seen to be faster on more mobile surfaces. This has consequential effects on downstream molecules; the mechanosensitive YAP protein localized to the nucleus more on less-mobile (more-viscous) surfaces and differentiation of myoblast cells was enhanced on higher viscosity. This behavior was explained within the framework of the molecular clutch model, with lower viscosity leading to a low force loading rate, preventing the exposure of mechanosensitive proteins, and with a higher viscosity causing a higher force loading rate exposing these sites, activating downstream pathways. Consequently, the understanding of how viscosity (regardless of matrix stiffness) influences cell response adds a further tool to engineer materials that control cell behavior.

JTD Keywords: Matrix rigidity, Molecular clutch, Surface viscosity, Mechanotransduction, Cell differentiation


Gauthier, Nils C., Roca-Cusachs, Pere, (2018). Mechanosensing at integrin-mediated cell–matrix adhesions: from molecular to integrated mechanisms Current Opinion in Cell Biology 50, 20-26

Integrin-mediated adhesions between cells and the extracellular matrix are fundamental for cell function, and one of their main roles is to sense and respond to mechanical force. Here we discuss the different mechanisms that can confer mechanosensitivity to adhesions. We first address molecular mechanisms mediated by force-induced changes in molecular properties, such as binding dynamics or protein conformation. Then, we discuss recent evidence on how these mechanisms are integrated with cellular and extracellular parameters such as myosin and actin activity, membrane tension, and ECM properties, endowing cells with an exquisite ability to both detect and respond to physical and mechanical cues from their environment.

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Roca-Cusachs, Pere, (2018). Cell scientist to watch - Pere Roca-Cusachs Journal of Cell Science 131, (16), jcs222596

Pere Roca-Cusachs received his doctorate in cellular biophysics from the University of Barcelona, Spain, in 2007, having worked with Daniel Navajas on mechanical properties of cells using atomic force microscopy. He then joined the laboratory of Michael Sheetz at Columbia University, USA, to study integrin-mediated mechanotransduction. In 2012, Pere established his own research group with a joint position between the Institute for Bioengineering of Catalonia (IBEC) and the University of Barcelona. His research focuses on the mechanisms cells employ to detect and respond to mechanical signals. Pere is the recipient of an EMBO young investigator award and has received numerous national awards.

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Escribano, Jorge, Sunyer, Raimon, Sánchez, María Teresa, Trepat, Xavier, Roca-Cusachs, Pere, García-Aznar, José Manuel, (2018). A hybrid computational model for collective cell durotaxis Biomechanics and Modeling in Mechanobiology 17, (4), 1037-1052

Collective cell migration is regulated by a complex set of mechanical interactions and cellular mechanisms. Collective migration emerges from mechanisms occurring at single cell level, involving processes like contraction, polymerization and depolymerization, of cell–cell interactions and of cell–substrate adhesion. Here, we present a computational framework which simulates the dynamics of this emergent behavior conditioned by substrates with stiffness gradients. The computational model reproduces the cell’s ability to move toward the stiffer part of the substrate, process known as durotaxis. It combines the continuous formulation of truss elements and a particle-based approach to simulate the dynamics of cell–matrix adhesions and cell–cell interactions. Using this hybrid approach, researchers can quickly create a quantitative model to understand the regulatory role of different mechanical conditions on the dynamics of collective cell migration. Our model shows that durotaxis occurs due to the ability of cells to deform the substrate more in the part of lower stiffness than in the stiffer part. This effect explains why cell collective movement is more effective than single cell movement in stiffness gradient conditions. In addition, we numerically evaluate how gradient stiffness properties, cell monolayer size and force transmission between cells and extracellular matrix are crucial in regulating durotaxis.

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Neri, L., Lasa, M., Elosegui-Artola, A., D'Avola, D., Carte, B., Gazquez, C., Alve, S., Roca-Cusachs, P., Iñarrairaegui, M., Herrero, J., Prieto, J., Sangro, B., Aldabe, R., (2017). NatB-mediated protein N-α-terminal acetylation is a potential therapeutic target in hepatocellular carcinoma Oncotarget 8, (25), 40967-40981

The identification of new targets for systemic therapy of hepatocellular carcinoma (HCC) is an urgent medical need. Recently, we showed that hNatB catalyzes the N-α-terminal acetylation of 15% of the human proteome and that this action is necessary for proper actin cytoskeleton structure and function. In tumors, cytoskeletal changes influence motility, invasion, survival, cell growth and tumor progression, making the cytoskeleton a very attractive antitumor target. Here, we show that hNatB subunits are upregulated in in over 59% HCC tumors compared to non-tumor tissue and that this upregulation is associated with microscopic vascular invasion. We found that hNatB silencing blocks proliferation and tumor formation in HCC cell lines in association with hampered DNA synthesis and impaired progression through the S and the G2/M phases. Growth inhibition is mediated by the degradation of two hNatB substrates, tropomyosin and CDK2, which occurs when these proteins lack N-α-terminal acetylation. In addition, hNatB inhibition disrupts the actin cytoskeleton, focal adhesions and tight/adherens junctions, abrogating two proliferative signaling pathways, Hippo/YAP and ERK1/2. Therefore, inhibition of NatB activity represents an interesting new approach to treating HCC by blocking cell proliferation and disrupting actin cytoskeleton function.

JTD Keywords: CDK2, Cell cycle arrest, Cell-cell junctions, Focal adhesions, Tropomyosin


Oria, Roger, Wiegand, Tina, Escribano, Jorge, Elosegui-Artola, Alberto, Uriarte, Juan Jose, Moreno-Pulido, Cristian, Platzman, Ilia, Delcanale, Pietro, Albertazzi, Lorenzo, Navajas, Daniel, Trepat, Xavier, García-Aznar, José Manuel, Cavalcanti-Adam, Elisabetta Ada, Roca-Cusachs, Pere, (2017). Force loading explains spatial sensing of ligands by cells Nature 552, 219-224

Cells can sense the density and distribution of extracellular matrix (ECM) molecules by means of individual integrin proteins and larger, integrin-containing adhesion complexes within the cell membrane. This spatial sensing drives cellular activity in a variety of normal and pathological contexts1,2. Previous studies of cells on rigid glass surfaces have shown that spatial sensing of ECM ligands takes place at the nanometre scale, with integrin clustering and subsequent formation of focal adhesions impaired when single integrin–ligand bonds are separated by more than a few tens of nanometres3,4,5,6. It has thus been suggested that a crosslinking ‘adaptor’ protein of this size might connect integrins to the actin cytoskeleton, acting as a molecular ruler that senses ligand spacing directly3,7,8,9. Here, we develop gels whose rigidity and nanometre-scale distribution of ECM ligands can be controlled and altered. We find that increasing the spacing between ligands promotes the growth of focal adhesions on low-rigidity substrates, but leads to adhesion collapse on more-rigid substrates. Furthermore, disordering the ligand distribution drastically increases adhesion growth, but reduces the rigidity threshold for adhesion collapse. The growth and collapse of focal adhesions are mirrored by, respectively, the nuclear or cytosolic localization of the transcriptional regulator protein YAP. We explain these findings not through direct sensing of ligand spacing, but by using an expanded computational molecular-clutch model10,11, in which individual integrin–ECM bonds—the molecular clutches—respond to force loading by recruiting extra integrins, up to a maximum value. This generates more clutches, redistributing the overall force among them, and reducing the force loading per clutch. At high rigidity and high ligand spacing, maximum recruitment is reached, preventing further force redistribution and leading to adhesion collapse. Measurements of cellular traction forces and actin flow speeds support our model. Our results provide a general framework for how cells sense spatial and physical information at the nanoscale, precisely tuning the range of conditions at which they form adhesions and activate transcriptional regulation.

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Rodriguez-Franco, P., Brugués, A., Marin-Llaurado, A., Conte, V., Solanas, G., Batlle, E., Fredberg, J. J., Roca-Cusachs, P., Sunyer, R., Trepat, X., (2017). Long-lived force patterns and deformation waves at repulsive epithelial boundaries Nature Materials 16, (10), 1029-1036

For an organism to develop and maintain homeostasis, cell types with distinct functions must often be separated by physical boundaries. The formation and maintenance of such boundaries are commonly attributed to mechanisms restricted to the cells lining the boundary. Here we show that, besides these local subcellular mechanisms, the formation and maintenance of tissue boundaries involves long-lived, long-ranged mechanical events. Following contact between two epithelial monolayers expressing, respectively, EphB2 and its ligand ephrinB1, both monolayers exhibit oscillatory patterns of traction forces and intercellular stresses that tend to pull cell-matrix adhesions away from the boundary. With time, monolayers jam, accompanied by the emergence of deformation waves that propagate away from the boundary. This phenomenon is not specific to EphB2/ephrinB1 repulsion but is also present during the formation of boundaries with an inert interface and during fusion of homotypic epithelial layers. Our findings thus unveil a global physical mechanism that sustains tissue separation independently of the biochemical and mechanical features of the local tissue boundary.

JTD Keywords: Biological physics, Cellular motility


Elosegui-Artola, A., Andreu, I., Beedle, A. E. M., Lezamiz, A., Uroz, M., Kosmalska, A. J., Oria, R., Kechagia, J. Z., Rico-Lastres, P., Le Roux, A. L., Shanahan, C. M., Trepat, X., Navajas, D., Garcia-Manyes, S., Roca-Cusachs, P., (2017). Force triggers YAP nuclear entry by regulating transport across nuclear pores Cell 171, (6), 1397-1410

YAP is a mechanosensitive transcriptional activator with a critical role in cancer, regeneration, and organ size control. Here, we show that force applied to the nucleus directly drives YAP nuclear translocation by decreasing the mechanical restriction of nuclear pores to molecular transport. Exposure to a stiff environment leads cells to establish a mechanical connection between the nucleus and the cytoskeleton, allowing forces exerted through focal adhesions to reach the nucleus. Force transmission then leads to nuclear flattening, which stretches nuclear pores, reduces their mechanical resistance to molecular transport, and increases YAP nuclear import. The restriction to transport is further regulated by the mechanical stability of the transported protein, which determines both active nuclear transport of YAP and passive transport of small proteins. Our results unveil a mechanosensing mechanism mediated directly by nuclear pores, demonstrated for YAP but with potential general applicability in transcriptional regulation. Force-dependent changes in nuclear pores control protein access to the nucleus.

JTD Keywords: Atomic force microscopy, Hippo pathway, Mechanosensing, Mechanotransduction, Molecular mechanical stability, Nuclear mechanics, Nuclear pores, Nuclear transport, Rigidity sensing, Transcription regulation


Labernadie, A., Kato, T., Brugués, A., Serra-Picamal, X., Derzsi, S., Arwert, E., Weston, A., González-Tarragó, V., Elosegui-Artola, A., Albertazzi, L., Alcaraz, J., Roca-Cusachs, P., Sahai, E., Trepat, X., (2017). A mechanically active heterotypic E-cadherin/N-cadherin adhesion enables fibroblasts to drive cancer cell invasion Nature Cell Biology 19, (3), 224-237

Cancer-associated fibroblasts (CAFs) promote tumour invasion and metastasis. We show that CAFs exert a physical force on cancer cells that enables their collective invasion. Force transmission is mediated by a heterophilic adhesion involving N-cadherin at the CAF membrane and E-cadherin at the cancer cell membrane. This adhesion is mechanically active; when subjected to force it triggers β-catenin recruitment and adhesion reinforcement dependent on α-catenin/vinculin interaction. Impairment of E-cadherin/N-cadherin adhesion abrogates the ability of CAFs to guide collective cell migration and blocks cancer cell invasion. N-cadherin also mediates repolarization of the CAFs away from the cancer cells. In parallel, nectins and afadin are recruited to the cancer cell/CAF interface and CAF repolarization is afadin dependent. Heterotypic junctions between CAFs and cancer cells are observed in patient-derived material. Together, our findings show that a mechanically active heterophilic adhesion between CAFs and cancer cells enables cooperative tumour invasion.

JTD


Roca-Cusachs, Pere, Conte, Vito, Trepat, Xavier, (2017). Quantifying forces in cell biology Nature Cell Biology 19, (7), 742-751

Cells exert, sense, and respond to physical forces through an astounding diversity of mechanisms. Here we review recently developed tools to quantify the forces generated by cells. We first review technologies based on sensors of known or assumed mechanical properties, and discuss their applicability and limitations. We then proceed to draw an analogy between these human-made sensors and force sensing in the cell. As mechanics is increasingly revealed to play a fundamental role in cell function we envisage that tools to quantify physical forces may soon become widely applied in life-sciences laboratories.

JTD


Pontes, B., Monzo, P., Gole, L., Le Roux, A. L., Kosmalska, A. J., Tam, Z. Y., Luo, W., Kan, S., Viasnoff, V., Roca-Cusachs, P., Tucker-Kellogg, L., Gauthier, N. C., (2017). Membrane tension controls adhesion positioning at the leading edge of cells Journal of Cell Biology , 216, (9), 2959-2977

Cell migration is dependent on adhesion dynamics and actin cytoskeleton remodeling at the leading edge. These events may be physically constrained by the plasma membrane. Here, we show that the mechanical signal produced by an increase in plasma membrane tension triggers the positioning of new rows of adhesions at the leading edge. During protrusion, as membrane tension increases, velocity slows, and the lamellipodium buckles upward in a myosin II-independent manner. The buckling occurs between the front of the lamellipodium, where nascent adhesions are positioned in rows, and the base of the lamellipodium, where a vinculin-dependent clutch couples actin to previously positioned adhesions. As membrane tension decreases, protrusion resumes and buckling disappears, until the next cycle. We propose that the mechanical signal of membrane tension exerts upstream control in mechanotransduction by periodically compressing and relaxing the lamellipodium, leading to the positioning of adhesions at the leading edge of cells.

JTD


González-Tarragó, V., Elosegui-Artola, A., Bazellières, E., Oria, R., Pérez-González, C., Roca-Cusachs, P., (2017). Binding of ZO-1 to α5β1 integrins regulates the mechanical properties of α5β1–fibronectin links Molecular Biology of the Cell , 28, (14), 1847-1852

Fundamental processes in cell adhesion, motility, and rigidity adaptation are regulated by integrin-mediated adhesion to the extracellular matrix (ECM). The link between the ECM component fibronectin (fn) and integrin α5β1 forms a complex with ZO-1 in cells at the edge of migrating monolayers, regulating cell migration. However, how this complex affects the α5β1-fn link is unknown. Here we show that the α5β1/ZO-1 complex decreases the resistance to force of α5β1-fn adhesions located at the edge of migrating cell monolayers while also increasing α5β1 recruitment. Consistently with a molecular clutch model of adhesion, this effect of ZO-1 leads to a decrease in the density and intensity of adhesions in cells at the edge of migrating monolayers. Taken together, our results unveil a new mode of integrin regulation through modification of the mechanical properties of integrin-ECM links, which may be harnessed by cells to control adhesion and migration.

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Elosegui-Artola, A., Roca-Cusachs, P., (2017). Amoebae as mechanosensitive tanks Biophysical Journal , 112, (12), 2457-2458

Whether employed to search for nutrients or to rearrange tissues, cell migration is essential to the function of uni- and multicellular systems, both in healthy conditions and in disease. Among the several described modes of migration, amoeboid migration is particularly intriguing due to its apparent simplicity. Indeed, it requires only low levels of cell-substrate adhesion, which does not even have to be mediated by specific molecular bonds. This type of migration can be observed across a broad range of cell types including neutrophils, lymphocytes, and tumor cells, although the best-studied case is that of the amoeba of the lower eukaryote Dictyostelium discoideum.

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Sunyer, R., Conte, V., Escribano, J., Elosegui-Artola, A., Labernadie, A., Valon, L., Navajas, D., García-Aznar, J. M., Muñoz, J. J., Roca-Cusachs, P., Trepat, X., (2016). Collective cell durotaxis emerges from long-range intercellular force transmission Science 353, (6304), 1157-1161

The ability of cells to follow gradients of extracellular matrix stiffness-durotaxis-has been implicated in development, fibrosis, and cancer. Here, we found multicellular clusters that exhibited durotaxis even if isolated constituent cells did not. This emergent mode of directed collective cell migration applied to a variety of epithelial cell types, required the action of myosin motors, and originated from supracellular transmission of contractile physical forces. To explain the observed phenomenology, we developed a generalized clutch model in which local stick-slip dynamics of cell-matrix adhesions was integrated to the tissue level through cell-cell junctions. Collective durotaxis is far more efficient than single-cell durotaxis; it thus emerges as a robust mechanism to direct cell migration during development, wound healing, and collective cancer cell invasion.

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Elosegui, Alberto, Oria, Roger, Chen, Yunfeng, Kosmalska, Anita, Perez-Gonzalez, Carlos, Castro, Natalia, Zhu, Cheng, Trepat, Xavier, Roca-Cusachs, Pere, (2016). Mechanical regulation of a molecular clutch defines force transmission and transduction in response to matrix rigidity Nature Cell Biology 18, (5), 540-548

Cell function depends on tissue rigidity, which cells probe by applying and transmitting forces to their extracellular matrix, and then transducing them into biochemical signals. Here we show that in response to matrix rigidity and density, force transmission and transduction are explained by the mechanical properties of the actin-talin-integrin-fibronectin clutch. We demonstrate that force transmission is regulated by a dynamic clutch mechanism, which unveils its fundamental biphasic force/rigidity relationship on talin depletion. Force transduction is triggered by talin unfolding above a stiffness threshold. Below this threshold, integrins unbind and release force before talin can unfold. Above the threshold, talin unfolds and binds to vinculin, leading to adhesion growth and YAP nuclear translocation. Matrix density, myosin contractility, integrin ligation and talin mechanical stability differently and nonlinearly regulate both force transmission and the transduction threshold. In all cases, coupling of talin unfolding dynamics to a theoretical clutch model quantitatively predicts cell response.

JTD


Wolfenson, Haguy, Meacci, Giovanni, Liu, Shuaimin, Stachowiak, Matthew R., Iskratsch, Thomas, Ghassemi, Saba, Roca-Cusachs, Pere, Oshaughnessy, Ben, Hone, James, Sheetz, Michael P., (2016). Tropomyosin controls sarcomere-like contractions for rigidity sensing and suppressing growth on soft matrices Nature Cell Biology 18, 33-42

Cells test the rigidity of the extracellular matrix by applying forces to it through integrin adhesions. Recent measurements show that these forces are applied by local micrometre-scale contractions, but how contraction force is regulated by rigidity is unknown. Here we performed high temporal- and spatial-resolution tracking of contractile forces by plating cells on sub-micrometre elastomeric pillars. We found that actomyosin-based sarcomere-like contractile units (CUs) simultaneously moved opposing pillars in net steps of [sim]2.5[thinsp]nm, independent of rigidity. What correlated with rigidity was the number of steps taken to reach a force level that activated recruitment of [alpha]-actinin to the CUs. When we removed actomyosin restriction by depleting tropomyosin 2.1, we observed larger steps and higher forces that resulted in aberrant rigidity sensing and growth of non-transformed cells on soft matrices. Thus, we conclude that tropomyosin 2.1 acts as a suppressor of growth on soft matrices by supporting proper rigidity sensing.

JTD Keywords: Cell adhesion, Mechanotransduction


Meacci, G., Wolfenson, H., Liu, S., Stachowiak, M. R., Iskratsch, T., Mathur, A., Ghassemi, S., Gauthier, N., Tabdanov, E., Lohner, J., Gondarenko, A., Chander, A. C., Roca-Cusachs, P., O'Shaughnessy, B., Hone, J., Sheetz, M. P., (2016). α-Actinin links extracellular matrix rigidity-sensing contractile units with periodic cell-edge retractions Molecular Biology of the Cell , 27, (22), 3471-3479

During spreading and migration, the leading edges of cells undergo periodic protrusion--retraction cycles. The functional purpose of these cycles is unclear. Here, using submicrometer polydimethylsiloxane pillars as substrates for cell spreading, we show that periodic edge retractions coincide with peak forces produced by local contractile units (CUs) that assemble and disassemble along the cell edge to test matrix rigidity. We find that, whereas actin rearward flow produces a relatively constant force inward, the peak of local contractile forces by CUs scales with rigidity. The cytoskeletal protein α-Actinin is shared between these two force-producing systems. It initially localizes to the CUs and subsequently moves inward with the actin flow. Knockdown of α-Actinin causes aberrant rigidity sensing, loss of CUs, loss of protrusion-retraction cycles, and, surprisingly, enables the cells to proliferate on soft matrices. We present a model based on these results in which local CUs drive rigidity sensing and adhesion formation.

JTD


Bazellières, Elsa, Conte, Vito, Elosegui, Alberto, Serra-Picamal, Xavier, Bintanel-Morcillo, María, Roca-Cusachs, Pere, Muñoz, José J., Sales-Pardo, Marta, Guimerà, Roger, Trepat, Xavier, (2015). Control of cell-cell forces and collective cell dynamics by the intercellular adhesome Nature Cell Biology 17, (4), 409-420

Dynamics of epithelial tissues determine key processes in development, tissue healing and cancer invasion. These processes are critically influenced by cell–cell adhesion forces. However, the identity of the proteins that resist and transmit forces at cell–cell junctions remains unclear, and how these proteins control tissue dynamics is largely unknown. Here we provide a systematic study of the interplay between cell–cell adhesion proteins, intercellular forces and epithelial tissue dynamics. We show that collective cellular responses to selective perturbations of the intercellular adhesome conform to three mechanical phenotypes. These phenotypes are controlled by different molecular modules and characterized by distinct relationships between cellular kinematics and intercellular forces. We show that these forces and their rates can be predicted by the concentrations of cadherins and catenins. Unexpectedly, we identified different mechanical roles for P-cadherin and E-cadherin; whereas P-cadherin predicts levels of intercellular force, E-cadherin predicts the rate at which intercellular force builds up.

JTD


Kosmalska, A. J., Casares, L., Elosegui, A., Thottacherry, J. J., Moreno-Vicente, R., González-Tarragó, V., Del Pozo, M. Á, Mayor, S., Arroyo, M., Navajas, D., Trepat, X., Gauthier, N. C., Roca-Cusachs, P., (2015). Physical principles of membrane remodelling during cell mechanoadaptation Nature Communications 6, 7292

Biological processes in any physiological environment involve changes in cell shape, which must be accommodated by their physical envelope - the bilayer membrane. However, the fundamental biophysical principles by which the cell membrane allows for and responds to shape changes remain unclear. Here we show that the 3D remodelling of the membrane in response to a broad diversity of physiological perturbations can be explained by a purely mechanical process. This process is passive, local, almost instantaneous, before any active remodelling and generates different types of membrane invaginations that can repeatedly store and release large fractions of the cell membrane. We further demonstrate that the shape of those invaginations is determined by the minimum elastic and adhesive energy required to store both membrane area and liquid volume at the cell-substrate interface. Once formed, cells reabsorb the invaginations through an active process with duration of the order of minutes.

JTD


Elosegui, A., Bazellières, E., Allen, M. D., Andreu, I., Oria, R., Sunyer, R., Gomm, J. J., Marshall, J. F., Jones, J. L., Trepat, X., Roca-Cusachs, P., (2014). Rigidity sensing and adaptation through regulation of integrin types Nature Materials 13, (6), 631-637

Tissue rigidity regulates processes in development, cancer and wound healing. However, how cells detect rigidity, and thereby modulate their behaviour, remains unknown. Here, we show that sensing and adaptation to matrix rigidity in breast myoepithelial cells is determined by the bond dynamics of different integrin types. Cell binding to fibronectin through either α5β1 integrins (constitutively expressed) or αvβ6 integrins (selectively expressed in cancer and development) adapts force generation, actin flow and integrin recruitment to rigidities associated with healthy or malignant tissue, respectively. In vitro experiments and theoretical modelling further demonstrate that this behaviour is explained by the different binding and unbinding rates of both integrin types to fibronectin. Moreover, rigidity sensing through differences in integrin bond dynamics applies both when integrins bind separately and when they compete for binding to fibronectin.

JTD


Roca-Cusachs, P., Del Rio, A., Puklin-Faucher, E., Gauthier, N. C., Biais, N., Sheetz, M. P., (2013). Integrin-dependent force transmission to the extracellular matrix by α-actinin triggers adhesion maturation Proceedings of the National Academy of Sciences of the United States of America 110, (15), E1361-E1370

Focal adhesions are mechanosensitive elements that enable mechanical communication between cells and the extracellular matrix. Here, we demonstrate a major mechanosensitive pathway in which α-actinin triggers adhesion maturation by linking integrins to actin in nascent adhesions. We show that depletion of the focal adhesion protein α-actinin enhances force generation in initial adhesions on fibronectin, but impairs mechanotransduction in a subsequent step, preventing adhesion maturation. Expression of an α-actinin fragment containing the integrin binding domain, however, dramatically reduces force generation in depleted cells. This behavior can be explained by a competition between talin (which mediates initial adhesion and force generation) and α-actinin for integrin binding. Indeed, we show in an in vitro assay that talin and α-actinin compete for binding to β3 integrins, but cooperate in binding to β1 integrins. Consistently, we find opposite effects of α-actinin depletion and expression of mutants on substrates that bind β3 integrins (fibronectin and vitronectin) versus substrates that only bind β1 integrins (collagen). We thus suggest that nascent adhesions composed of β3 integrins are initially linked to the actin cytoskeleton by talin, and then α-actinin competes with talin to bind β3 integrins. Force transmitted through α-actinin then triggers adhesion maturation. Once adhesions have matured, α-actinin recruitment correlates with force generation, suggesting that α-actinin is the main link transmitting force between integrins and the cytoskeleton in mature adhesions. Such a multistep process enables cells to adjust forces on matrices, unveiling a role of α-actinin that is different from its well-studied function as an actin cross-linker.

JTD


Roca-Cusachs, P., Sunyer, R., Trepat, X., (2013). Mechanical guidance of cell migration: lessons from chemotaxis Current Opinion in Cell Biology 25, (5), 543-549

For an organism to develop, for a wound to heal, or for a tumor to invade, cells must be able to migrate following directional cues. It is widely accepted that directed cell migration is enabled by cellular sensing of local gradients in the concentration of chemical factors. The main molecular players involved in this mode of cellular guidance - chemotaxis - have been identified and the combination of modeling and experimental approaches is progressively unveiling a clear picture of the underlying mechanisms. Evidence obtained over the past decade has shown that cells can also be guided by mechanical stimuli such as physical forces or gradients in extracellular matrix stiffness. Mechanical guidance, which we refer here globally as mechanotaxis, is also thought to drive processes in development, cancer, and wound healing, but experimental evidence is scattered and mechanisms remain largely unknown. Here we use the better understood process of chemotaxis as a reference to define the building blocks that are required for cell guidance, and then discuss how these building blocks might be organized in mechanotaxis. We show that both chemotaxis and mechanotaxis involve an exquisite interplay between physical and chemical mechanisms to sense gradients, establish polarization, and drive directed migration.

JTD


Ghassemi, S., Meacci, G., Liu, S., Gondarenko, A. A., Mathur, A., Roca-Cusachs, P., Sheetz, M. P., Hone, J., (2012). Cells test substrate rigidity by local contractions on submicrometer pillars Proceedings of the National Academy of Sciences of the United States of America 109, (14), 5328-5333

Cell growth and differentiation are critically dependent upon matrix rigidity, yet many aspects of the cellular rigidity-sensing mechanism are not understood. Here, we analyze matrix forces after initial cell-matrix contact, when early rigidity-sensing events occur, using a series of elastomeric pillar arrays with dimensions extending to the submicron scale (2, 1, and 0.5 μm in diameter covering a range of stiffnesses). We observe that the cellular response is fundamentally different on micron-scale and submicron pillars. On 2-μm diameter pillars, adhesions form at the pillar periphery, forces are directed toward the center of the cell, and a constant maximum force is applied independent of stiffness. On 0.5-μm diameter pillars, adhesions form on the pillar tops, and local contractions between neighboring pillars are observed with a maximum displacement of ∼60 nm, independent of stiffness. Because mutants in rigidity sensing show no detectable displacement on 0.5-μm diameter pillars, there is a correlation between local contractions to 60 nm and rigidity sensing. Localization of myosin between submicron pillars demonstrates that submicron scale myosin filaments can cause these local contractions. Finally, submicron pillars can capture many details of cellular force generation that are missed on larger pillars and more closely mimic continuous surfaces.

JTD Keywords: Cell mechanics, Mechanotransduction, Nanofabrication


Roca-Cusachs, P., Iskratsch, T., Sheetz, M. P., (2012). Finding the weakest link: exploring integrin-mediated mechanical molecular pathways Journal of Cell Science 125, (13), 3025-3038

From the extracellular matrix to the cytoskeleton, a network of molecular links connects cells to their environment. Molecules in this network transmit and detect mechanical forces, which subsequently determine cell behavior and fate. Here, we reconstruct the mechanical pathway followed by these forces. From matrix proteins to actin through integrins and adaptor proteins, we review how forces affect the lifetime of bonds and stretch or alter the conformation of proteins, and how these mechanical changes are converted into biochemical signals in mechanotransduction events. We evaluate which of the proteins in the network can participate in mechanotransduction and which are simply responsible for transmitting forces in a dynamic network. Besides their individual properties, we also analyze how the mechanical responses of a protein are determined by their serial connections from the matrix to actin, their parallel connections in integrin clusters and by the rate at which force is applied to them. All these define mechanical molecular pathways in cells, which are emerging as key regulators of cell function alongside better studied biochemical pathways.

JTD Keywords: Cell adhesion, Cytoskeleton, Mechanotransduction


Gauthier, Nils C., Fardin, Marc Antoine, Roca-Cusachs, Pere, Sheetz, Michael P., (2011). Temporary increase in plasma membrane tension coordinates the activation of exocytosis and contraction during cell spreading Proceedings of the National Academy of Sciences of the United States of America 108, (35), 14467-14472

Cell migration and spreading involve the coordination of membrane trafficking, actomyosin contraction, and modifications to plasma membrane tension and area. The biochemical or biophysical basis for this coordination is however unknown. In this study, we show that during cell spreading, lamellipodia protrusion flattens plasma membrane folds and blebs and, once the plasma membrane area is depleted, there is a temporary increase in membrane tension by over twofold that is followed by activation of exocytosis and myosin contraction. Further, an artificial increase in plasma membrane tension stopped lamellipodia protrusion and activated an exocytotic burst. Subsequent decrease in tension restored spreading with activation of contraction. Conversely, blebbistatin inhibition of actomyosin contraction resulted in an even greater increase in plasma membrane tension and exocytosis activation. This spatio-temporal synchronization indicates that membrane tension is the signal that coordinates membrane trafficking, actomyosin contraction, and plasma membrane area change. We suggest that cells use plasma membrane tension as a global physical parameter to control cell motility.

JTD Keywords: Surface-area regulation, Cytoskeleton adhesion, Erythrocyte-membrane, Extensional flow, Elastic tether, Force


Moore, S. W., Roca-Cusachs, P., Sheetz, M. P., (2010). Stretchy proteins on stretchy substrates: The important elements of integrin-mediated rigidity sensing Developmental Cell 19, (2), 194-206

Matrix and tissue rigidity guides many cellular processes, including the differentiation of stem cells and the migration of cells in health and disease. Cells actively and transiently test rigidity using mechanisms limited by inherent physical parameters that include the strength of extracellular attachments, the pulling capacity on these attachments, and the sensitivity of the mechanotransduction system. Here, we focus on rigidity sensing mediated through the integrin family of extracellular matrix receptors and linked proteins and discuss the evidence supporting these proteins as mechanosensors.

JTD Keywords: Focal adhesion kinase, Atomic Force Microscopy, Smooth-muscle cells, Traction forces, Living cells, Mechanical force, Locomoting cells


Roca-Cusachs, P., Gauthier, N. C., del Rio, A., Sheetz, M. P., (2009). Clustering of alpha(5)beta(1) integrins determines adhesion strength whereas alpha(v)beta(3) and talin enable mechanotransduction Proceedings of the National Academy of Sciences of the United States of America 106, (38), 16245-16250

A key molecular link between cells and the extracellular matrix is the binding between fibronectin and integrins alpha(5)beta(1) and alpha(v)beta(3). However, the roles of these different integrins in establishing adhesion remain unclear. We tested the adhesion strength of fibronectin-integrin-cytoskeleton linkages by applying physiological nanonewton forces to fibronectin-coated magnetic beads bound to cells. We report that the clustering of fibronectin domains within 40 nm led to integrin alpha(5)beta(1) recruitment, and increased the ability to sustain force by over six-fold. This force was supported by alpha(5)beta(1) integrin clusters. Importantly, we did not detect a role of either integrin alpha(v)beta(3) or talin 1 or 2 in maintaining adhesion strength. Instead, these molecules enabled the connection to the cytoskeleton and reinforcement in response to an applied force. Thus, high matrix forces are primarily supported by clustered alpha(5)beta(1) integrins, while less stable links to alpha(v)beta(3) integrins initiate mechanotransduction, resulting in reinforcement of integrin-cytoskeleton linkages through talin-dependent bonds.

JTD Keywords: Cell-adhesion, Mechanical force, Vinculin-binding, Fibronectin, Activation, Dynamics, Domain, Alpha-v-beta-3, Translocation, Bonds


Gavara, N., Roca-Cusachs, P., Sunyer, R., Farre, R., Navajas, D., (2008). Mapping cell-matrix stresses during stretch reveals inelastic reorganization of the cytoskeleton Biophysical Journal , 95, (1), 464-471

The mechanical properties of the living cell are intimately related to cell signaling biology through cytoskeletal tension. The tension borne by the cytoskeleton (CSK) is in part generated internally by the actomyosin machinery and externally by stretch. Here we studied how cytoskeletal tension is modified during stretch and the tensional changes undergone by the sites of cell-matrix interaction. To this end we developed a novel technique to map cell-matrix stresses during application of stretch. We found that cell-matrix stresses increased with imposition of stretch but dropped below baseline levels on stretch release. Inhibition of the actomyosin machinery resulted in a larger relative increase in CSK tension with stretch and in a smaller drop in tension after stretch release. Cell-matrix stress maps showed that the loci of cell adhesion initially bearing greater stress also exhibited larger drops in traction forces after stretch removal. Our results suggest that stretch partially disrupts the actin-myosin apparatus and the cytoskeletal structures that support the largest CSK tension. These findings indicate that cells use the mechanical energy injected by stretch to rapidly reorganize their structure and redistribute tension.

JTD Keywords: Cell Line, Computer Simulation, Cytoskeleton/ physiology, Elasticity, Epithelial Cells/ physiology, Extracellular Matrix/ physiology, Humans, Mechanotransduction, Cellular/ physiology, Models, Biological, Stress, Mechanical


Roca-Cusachs, P., Alcaraz, J., Sunyer, R., Samitier, J., Farre, R., Navajas, D., (2008). Micropatterning of single endothelial cell shape reveals a tight coupling between nuclear volume in G1 and proliferation Biophysical Journal , 94, (12), 4984-4995

Shape-dependent local differentials in cell proliferation are considered to be a major driving mechanism of structuring processes in vivo, such as embryogenesis, wound healing, and angiogenesis. However, the specific biophysical signaling by which changes in cell shape contribute to cell cycle regulation remains poorly understood. Here, we describe our study of the roles of nuclear volume and cytoskeletal mechanics in mediating shape control of proliferation in single endothelial cells. Micropatterned adhesive islands were used to independently control cell spreading and elongation. We show that, irrespective of elongation, nuclear volume and apparent chromatin decondensation of cells in G1 systematically increased with cell spreading and highly correlated with DNA synthesis (percent of cells in the S phase). In contrast, cell elongation dramatically affected the organization of the actin cytoskeleton, markedly reduced both cytoskeletal stiffness (measured dorsally with atomic force microscopy) and contractility (measured ventrally with traction microscopy), and increased mechanical anisotropy, without affecting either DNA synthesis or nuclear volume. Our results reveal that the nuclear volume in G1 is predictive of the proliferative status of single endothelial cells within a population, whereas cell stiffness and contractility are not. These findings show that the effects of cell mechanics in shape control of proliferation are far more complex than a linear or straightforward relationship. Our data are consistent with a mechanism by which spreading of cells in G1 partially enhances proliferation by inducing nuclear swelling and decreasing chromatin condensation, thereby rendering DNA more accessible to the replication machinery.

JTD Keywords: Cell Line, Cell Nucleus/ physiology, Cell Proliferation, Cell Size, Computer Simulation, Endothelial Cells/ cytology/ physiology, G1 Phase/ physiology, Humans, Mechanotransduction, Cellular/ physiology, Models, Biological, Statistics as Topic


Rico, Félix, Roca-Cusachs, Pere, Sunyer, Raimon, Farré, Ramon, Navajas, Daniel, (2007). Cell dynamic adhesion and elastic properties probed with cylindrical atomic force microscopy cantilever tips Journal of Molecular Recognition John Wiley & Sons, Ltd. 20, (6), 459-466

Cell adhesion is required for essential biological functions such as migration, tissue formation and wound healing, and it is mediated by individual molecules that bind specifically to ligands on other cells or on the extracellular matrix. Atomic force microscopy (AFM) has been successfully used to measure cell adhesion at both single molecule and whole cell levels. However, the measurement of inherent cell adhesion properties requires a constant cell-probe contact area during indentation, a requirement which is not fulfilled in common pyramidal or spherical AFM tips. We developed a procedure using focused ion beam (FIB) technology by which we modified silicon pyramidal AFM cantilever tips to obtain flat-ended cylindrical tips with a constant and known area of contact. The tips were validated on elastic gels and living cells. Cylindrical tips showed a fairly linear force-indentation behaviour on both gels and cells for indentations > 200nm. Cylindrical tips coated with ligands were used to quantify inherent dynamic cell adhesion and elastic properties. Force, work of adhesion and elasticity showed a marked dynamic response. In contrast, the deformation applied to the cells before rupture was fairly constant within the probed dynamic range. Taken together, these results suggest that the dynamic adhesion strength is counterbalanced by the dynamic elastic response to keep a constant cell deformation regardless of the applied pulling rate.

JTD Keywords: AFM, Cell adhesion, Cell mechanics, Cell stiffness