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1.
Elife ; 122023 08 07.
Artigo em Inglês | MEDLINE | ID: mdl-37548995

RESUMO

Cell-generated forces play a major role in coordinating the large-scale behavior of cell assemblies, in particular during development, wound healing, and cancer. Mechanical signals propagate faster than biochemical signals, but can have similar effects, especially in epithelial tissues with strong cell-cell adhesion. However, a quantitative description of the transmission chain from force generation in a sender cell, force propagation across cell-cell boundaries, and the concomitant response of receiver cells is missing. For a quantitative analysis of this important situation, here we propose a minimal model system of two epithelial cells on an H-pattern ('cell doublet'). After optogenetically activating RhoA, a major regulator of cell contractility, in the sender cell, we measure the mechanical response of the receiver cell by traction force and monolayer stress microscopies. In general, we find that the receiver cells show an active response so that the cell doublet forms a coherent unit. However, force propagation and response of the receiver cell also strongly depend on the mechano-structural polarization in the cell assembly, which is controlled by cell-matrix adhesion to the adhesive micropattern. We find that the response of the receiver cell is stronger when the mechano-structural polarization axis is oriented perpendicular to the direction of force propagation, reminiscent of the Poisson effect in passive materials. We finally show that the same effects are at work in small tissues. Our work demonstrates that cellular organization and active mechanical response of a tissue are key to maintain signal strength and lead to the emergence of elasticity, which means that signals are not dissipated like in a viscous system, but can propagate over large distances.


Assuntos
Células Epiteliais , Fenômenos Mecânicos , Células Epiteliais/fisiologia , Epitélio , Adesão Celular/fisiologia , Elasticidade , Estresse Mecânico
2.
Nat Commun ; 14(1): 2740, 2023 05 22.
Artigo em Inglês | MEDLINE | ID: mdl-37217519

RESUMO

Cell migration is crucial for cancer dissemination. We find that AMP-activated protein kinase (AMPK) controls cell migration by acting as an adhesion sensing molecular hub. In 3-dimensional matrices, fast-migrating amoeboid cancer cells exert low adhesion/low traction linked to low ATP/AMP, leading to AMPK activation. In turn, AMPK plays a dual role controlling mitochondrial dynamics and cytoskeletal remodelling. High AMPK activity in low adhering migratory cells, induces mitochondrial fission, resulting in lower oxidative phosphorylation and lower mitochondrial ATP. Concurrently, AMPK inactivates Myosin Phosphatase, increasing Myosin II-dependent amoeboid migration. Reducing adhesion or mitochondrial fusion or activating AMPK induces efficient rounded-amoeboid migration. AMPK inhibition suppresses metastatic potential of amoeboid cancer cells in vivo, while a mitochondrial/AMPK-driven switch is observed in regions of human tumours where amoeboid cells are disseminating. We unveil how mitochondrial dynamics control cell migration and suggest that AMPK is a mechano-metabolic sensor linking energetics and the cytoskeleton.


Assuntos
Proteínas Quinases Ativadas por AMP , Dinâmica Mitocondrial , Neoplasias , Humanos , Trifosfato de Adenosina/metabolismo , Proteínas Quinases Ativadas por AMP/metabolismo , Adesão Celular , Movimento Celular/fisiologia , Miosina Tipo II/metabolismo , Fosforilação Oxidativa , Fosforilação
3.
Proc Natl Acad Sci U S A ; 119(49): e2201600119, 2022 12 06.
Artigo em Inglês | MEDLINE | ID: mdl-36454762

RESUMO

The direction in which a cell divides is set by the orientation of its mitotic spindle and is important for determining cell fate, controlling tissue shape, and maintaining tissue architecture. Divisions parallel to the epithelial plane sustain tissue expansion. By contrast, divisions perpendicular to the plane promote tissue stratification and lead to the loss of epithelial cells from the tissue-an event that has been suggested to promote metastasis. Much is known about the molecular machinery involved in orienting the spindle, but less is known about the contribution of mechanical factors, such as tissue tension, in ensuring spindle orientation in the plane of the epithelium. This is important as epithelia are continuously subjected to mechanical stresses. To explore this further, we subjected suspended epithelial monolayers devoid of extracellular matrix to varying levels of tissue tension to study the orientation of cell divisions relative to the tissue plane. This analysis revealed that lowering tissue tension by compressing epithelial monolayers or by inhibiting myosin contractility increased the frequency of out-of-plane divisions. Reciprocally, increasing tissue tension by elevating cell contractility or by tissue stretching restored accurate in-plane cell divisions. Moreover, a characterization of the geometry of cells within these epithelia suggested that spindles can sense tissue tension through its impact on tension at subcellular surfaces, independently of their shape. Overall, these data suggest that accurate spindle orientation in the plane of the epithelium relies on a threshold level of tension at intercellular junctions.


Assuntos
Células Epiteliais , Junções Intercelulares , Epitélio , Divisão Celular , Matriz Extracelular
5.
Nature ; 609(7927): 469-470, 2022 09.
Artigo em Inglês | MEDLINE | ID: mdl-35978164
6.
Proc Natl Acad Sci U S A ; 119(26): e2121868119, 2022 06 28.
Artigo em Inglês | MEDLINE | ID: mdl-35727980

RESUMO

Proper orientation of the mitotic spindle plays a crucial role in embryos, during tissue development, and in adults, where it functions to dissipate mechanical stress to maintain tissue integrity and homeostasis. While mitotic spindles have been shown to reorient in response to external mechanical stresses, the subcellular cues that mediate spindle reorientation remain unclear. Here, we used a combination of optogenetics and computational modeling to investigate how mitotic spindles respond to inhomogeneous tension within the actomyosin cortex. Strikingly, we found that the optogenetic activation of RhoA only influences spindle orientation when it is induced at both poles of the cell. Under these conditions, the sudden local increase in cortical tension induced by RhoA activation reduces pulling forces exerted by cortical regulators on astral microtubules. This leads to a perturbation of the balance of torques exerted on the spindle, which causes it to rotate. Thus, spindle rotation in response to mechanical stress is an emergent phenomenon arising from the interaction between the spindle positioning machinery and the cell cortex.


Assuntos
Microtúbulos , Fuso Acromático , Estresse Mecânico , Actomiosina/metabolismo , Simulação por Computador , Citoplasma , Microtúbulos/metabolismo , Optogenética , Fuso Acromático/fisiologia , Proteína rhoA de Ligação ao GTP/metabolismo
7.
Development ; 149(12)2022 06 15.
Artigo em Inglês | MEDLINE | ID: mdl-35593440

RESUMO

Planar cell polarity (PCP) is the aligned cell polarity within a tissue plane. Mechanical signals are known to act as a global cue for PCP, yet their exact role is still unclear. In this study, we focused on PCP in the posterior neuroectoderm of Xenopus laevis and investigated how mechanical signals regulate polarity. We reveal that the neuroectoderm is under a greater tension in the anterior-posterior direction and that perturbation of this tension causes PCP disappearance. We show that application of uniaxial stretch to explant tissues can control the orientation of PCP and that cells sense the tissue stretch indirectly through a change in their shape, rather than directly through detection of anisotropic tension. Furthermore, we reveal that PCP is most strongly established when the orientation of tissue stretch coincides with that of diffusion of locally expressed Wnt ligands, suggesting a cooperative relationship between these two PCP regulators.


Assuntos
Polaridade Celular , Via de Sinalização Wnt , Animais , Polaridade Celular/fisiologia , Xenopus laevis
8.
Trends Cell Biol ; 32(6): 537-551, 2022 06.
Artigo em Inglês | MEDLINE | ID: mdl-35190218

RESUMO

During development and in adult physiology, living tissues are continuously subjected to mechanical stresses originating either from cellular processes intrinsic to the tissue or from external forces. As a consequence, rupture is a constant risk and can arise as a result of excessive stresses or because of tissue weakening through genetic abnormalities or pathologies. Tissue fracture is a multiscale process involving the unzipping of intercellular adhesions at the molecular scale in response to stresses arising at the tissue or cellular scale that are transmitted to adhesion complexes via the cytoskeleton. In this review we detail experimental characterization and theoretical approaches for understanding the fracture of living tissues at the tissue, cellular, and molecular scales.


Assuntos
Caderinas , Citoesqueleto , Caderinas/genética , Adesão Celular/fisiologia , Humanos , Mecanotransdução Celular/fisiologia , Estresse Mecânico
9.
iScience ; 24(12): 103482, 2021 Dec 17.
Artigo em Inglês | MEDLINE | ID: mdl-34927026

RESUMO

Cells maintain their volume through fine intracellular osmolarity regulation. Osmotic challenges drive fluid into or out of cells causing swelling or shrinkage, respectively. The dynamics of cell volume changes depending on the rheology of the cellular constituents and on how fast the fluid permeates through the membrane and cytoplasm. We investigated whether and how poroelasticity can describe volume dynamics in response to osmotic shocks. We exposed cells to osmotic perturbations and used defocusing epifluorescence microscopy on membrane-attached fluorescent nanospheres to track volume dynamics with high spatiotemporal resolution. We found that a poroelastic model that considers both geometrical and pressurization rates captures fluid-cytoskeleton interactions, which are rate-limiting factors in controlling volume changes at short timescales. Linking cellular responses to osmotic shocks and cell mechanics through poroelasticity can predict the cell state in health, disease, or in response to novel therapeutics.

10.
Nat Commun ; 12(1): 6511, 2021 11 11.
Artigo em Inglês | MEDLINE | ID: mdl-34764258

RESUMO

In animal cells, shape is mostly determined by the actomyosin cortex, a thin cytoskeletal network underlying the plasma membrane. Myosin motors generate tension in the cortex, and tension gradients result in cellular deformations. As such, many cell morphogenesis studies have focused on the mechanisms controlling myosin activity and recruitment to the cortex. Here, we demonstrate using super-resolution microscopy that myosin does not always overlap with actin at the cortex, but remains restricted towards the cytoplasm in cells with low cortex tension. We propose that this restricted penetration results from steric hindrance, as myosin minifilaments are considerably larger than the cortical actin meshsize. We identify myosin activity and actin network architecture as key regulators of myosin penetration into the cortex, and show that increasing myosin penetration increases cortical tension. Our study reveals that the spatial coordination of myosin and actin at the cortex regulates cell surface mechanics, and unveils an important mechanism whereby myosin size controls its action by limiting minifilament penetration into the cortical actin network. More generally, our findings suggest that protein size could regulate function in dense cytoskeletal structures.


Assuntos
Miosinas/metabolismo , Citoesqueleto de Actina/metabolismo , Actinas/metabolismo , Animais , Membrana Celular/metabolismo
11.
Int J Mol Sci ; 22(16)2021 Aug 20.
Artigo em Inglês | MEDLINE | ID: mdl-34445693

RESUMO

Mechanical forces acting on cell-cell adhesion modulate the barrier function of endothelial cells. The actively remodeled actin cytoskeleton impinges on cell-cell adhesion to counteract external forces. We applied stress on endothelial monolayers by mechanical stretch to uncover the role of BRAF in the stress-induced response. Control cells responded to external forces by organizing and stabilizing actin cables in the stretched cell junctions. This was accompanied by an increase in intercellular gap formation, which was prevented in BRAF knockdown monolayers. In the absence of BRAF, there was excess stress fiber formation due to the enhanced reorganization of actin fibers. Our findings suggest that stretch-induced intercellular gap formation, leading to a decrease in barrier function of blood vessels, can be reverted by BRAF RNAi. This is important when the endothelium experiences changes in external stresses caused by high blood pressure, leading to edema, or by immune or cancer cells in inflammation or metastasis.


Assuntos
Células Endoteliais/metabolismo , Junções Comunicantes/fisiologia , Proteínas Proto-Oncogênicas B-raf/metabolismo , Actinas/fisiologia , Adesão Celular/fisiologia , Células Cultivadas , Citoesqueleto/fisiologia , Células Endoteliais/fisiologia , Endotélio Vascular/citologia , Humanos , Junções Intercelulares/fisiologia , Fenômenos Mecânicos , Proteínas Proto-Oncogênicas B-raf/fisiologia
12.
Elife ; 102021 05 20.
Artigo em Inglês | MEDLINE | ID: mdl-34014166

RESUMO

How cells with different genetic makeups compete in tissues is an outstanding question in developmental biology and cancer research. Studies in recent years have revealed that cell competition can either be driven by short-range biochemical signalling or by long-range mechanical stresses in the tissue. To date, cell competition has generally been characterised at the population scale, leaving the single-cell-level mechanisms of competition elusive. Here, we use high time-resolution experimental data to construct a multi-scale agent-based model for epithelial cell competition and use it to gain a conceptual understanding of the cellular factors that governs competition in cell populations within tissues. We find that a key determinant of mechanical competition is the difference in homeostatic density between winners and losers, while differences in growth rates and tissue organisation do not affect competition end result. In contrast, the outcome and kinetics of biochemical competition is strongly influenced by local tissue organisation. Indeed, when loser cells are homogenously mixed with winners at the onset of competition, they are eradicated; however, when they are spatially separated, winner and loser cells coexist for long times. These findings suggest distinct biophysical origins for mechanical and biochemical modes of cell competition.


Assuntos
Competição entre as Células , Células Epiteliais/fisiologia , Mecanotransdução Celular , Modelos Biológicos , Animais , Apoptose , Fenômenos Biomecânicos , Comunicação Celular , Proliferação de Células , Simulação por Computador , Cães , Genótipo , Cinética , Células Madin Darby de Rim Canino , Fenótipo , Análise de Célula Única , Estresse Mecânico
13.
Nat Mater ; 20(3): 281-283, 2021 03.
Artigo em Inglês | MEDLINE | ID: mdl-33633346
14.
J Cell Sci ; 134(3)2021 02 08.
Artigo em Inglês | MEDLINE | ID: mdl-33408247

RESUMO

The migration of circulating neutrophils towards damaged or infected tissue is absolutely critical to the inflammatory response. L-selectin is a cell adhesion molecule abundantly expressed on circulating neutrophils. For over two decades, neutrophil L-selectin has been assigned the exclusive role of supporting tethering and rolling - the initial stages of the multi-step adhesion cascade. Here, we provide direct evidence for L-selectin contributing to neutrophil transendothelial migration (TEM). We show that L-selectin co-clusters with PECAM-1 - a well-characterised cell adhesion molecule involved in regulating neutrophil TEM. This co-clustering behaviour occurs specifically during TEM, which serves to augment ectodomain shedding of L-selectin and expedite the time taken for TEM (TTT) to complete. Blocking PECAM-1 signalling (through mutation of its cytoplasmic tail), PECAM-1-dependent adhesion or L-selectin shedding, leads to a significant delay in the TTT. Finally, we show that co-clustering of L-selectin with PECAM-1 occurs specifically across TNF- but not IL-1ß-activated endothelial monolayers - implying unique adhesion interactomes forming in a cytokine-specific manner. To our knowledge, this is the first report to implicate a non-canonical role for L-selectin in regulating neutrophil TEM.


Assuntos
Movimento Celular , Selectina L , Neutrófilos , Migração Transendotelial e Transepitelial , Adesão Celular , Endotélio Vascular , Humanos , Selectina L/genética
15.
Nat Cell Biol ; 22(7): 803-814, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32572169

RESUMO

Cell shape is controlled by the submembranous cortex, an actomyosin network mainly generated by two actin nucleators: the Arp2/3 complex and the formin mDia1. Changes in relative nucleator activity may alter cortical organization, mechanics and cell shape. Here we investigate how nucleation-promoting factors mediate interactions between nucleators. In vitro, the nucleation-promoting factor SPIN90 promotes formation of unbranched filaments by Arp2/3, a process thought to provide the initial filament for generation of dendritic networks. Paradoxically, in cells, SPIN90 appears to favour a formin-dominated cortex. Our in vitro experiments reveal that this feature stems mainly from two mechanisms: efficient recruitment of mDia1 to SPIN90-Arp2/3 nucleated filaments and formation of a ternary SPIN90-Arp2/3-mDia1 complex that greatly enhances filament nucleation. Both mechanisms yield rapidly elongating filaments with mDia1 at their barbed ends and SPIN90-Arp2/3 at their pointed ends. Thus, in networks, SPIN90 lowers branching densities and increases the proportion of long filaments elongated by mDia1.


Assuntos
Citoesqueleto de Actina/fisiologia , Complexo 2-3 de Proteínas Relacionadas à Actina/metabolismo , Actinas/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Forminas/metabolismo , Melanoma/patologia , Proteínas Musculares/metabolismo , Complexo 2-3 de Proteínas Relacionadas à Actina/genética , Proteínas Adaptadoras de Transdução de Sinal/genética , Animais , Blástula/citologia , Blástula/metabolismo , Forma Celular , Embrião não Mamífero/citologia , Embrião não Mamífero/metabolismo , Forminas/genética , Humanos , Melanoma/genética , Melanoma/metabolismo , Proteínas Musculares/genética , Xenopus laevis/crescimento & desenvolvimento , Xenopus laevis/metabolismo
16.
Curr Opin Cell Biol ; 66: 69-78, 2020 10.
Artigo em Inglês | MEDLINE | ID: mdl-32580115

RESUMO

The actin cortex is a thin layer of actin, myosin and actin-binding proteins that underlies the membrane of most animal cells. It is highly dynamic and can undergo remodelling on timescales of tens of seconds, thanks to protein turnover and myosin-mediated contractions. The cortex enables cells to resist external mechanical stresses, controls cell shape and allows cells to exert forces on their neighbours. Thus, its mechanical properties are the key to its physiological function. Here, we give an overview of how cortex composition, structure and dynamics control cortex mechanics and cell shape. We use mitosis as an example to illustrate how global and local regulation of cortex mechanics gives rise to a complex series of cell shape changes.


Assuntos
Actinas/metabolismo , Transdução de Sinais , Citoesqueleto de Actina/metabolismo , Actomiosina/metabolismo , Animais , Fenômenos Biomecânicos , Humanos , Modelos Biológicos
17.
Nat Mater ; 19(9): 1019-1025, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32451510

RESUMO

Cortical stiffness is an important cellular property that changes during migration, adhesion and growth. Previous atomic force microscopy (AFM) indentation measurements of cells cultured on deformable substrates have suggested that cells adapt their stiffness to that of their surroundings. Here we show that the force applied by AFM to a cell results in a significant deformation of the underlying substrate if this substrate is softer than the cell. This 'soft substrate effect' leads to an underestimation of a cell's elastic modulus when analysing data using a standard Hertz model, as confirmed by finite element modelling and AFM measurements of calibrated polyacrylamide beads, microglial cells and fibroblasts. To account for this substrate deformation, we developed a 'composite cell-substrate model'. Correcting for the substrate indentation revealed that cortical cell stiffness is largely independent of substrate mechanics, which has major implications for our interpretation of many physiological and pathological processes.


Assuntos
Córtex Cerebral/citologia , Diferenciação Celular , Módulo de Elasticidade , Microscopia de Força Atômica/métodos , Especificidade por Substrato
18.
Proc Natl Acad Sci U S A ; 117(17): 9377-9383, 2020 04 28.
Artigo em Inglês | MEDLINE | ID: mdl-32284424

RESUMO

Epithelial monolayers are two-dimensional cell sheets which compartmentalize the body and organs of multicellular organisms. Their morphogenesis during development or pathology results from patterned endogenous and exogenous forces and their interplay with tissue mechanical properties. In particular, bending of epithelia is thought to result from active torques generated by the polarization of myosin motors along their apicobasal axis. However, the contribution of these out-of-plane forces to morphogenesis remains challenging to evaluate because of the lack of direct mechanical measurement. Here we use epithelial curling to characterize the out-of-plane mechanics of epithelial monolayers. We find that curls of high curvature form spontaneously at the free edge of epithelial monolayers devoid of substrate in vivo and in vitro. Curling originates from an enrichment of myosin in the basal domain that generates an active spontaneous curvature. By measuring the force necessary to flatten curls, we can then estimate the active torques and the bending modulus of the tissue. Finally, we show that the extent of curling is controlled by the interplay between in-plane and out-of-plane stresses in the monolayer. Such mechanical coupling emphasizes a possible role for in-plane stresses in shaping epithelia during morphogenesis.


Assuntos
Epitélio/fisiologia , Animais , Fenômenos Biomecânicos , Adesão Celular , Linhagem Celular , Cães , Elasticidade , Estresse Mecânico
19.
Mol Biol Cell ; 31(13): 1370-1379, 2020 06 15.
Artigo em Inglês | MEDLINE | ID: mdl-32320325

RESUMO

The ability of cells to divide along their longest axis has been proposed to play an important role in maintaining epithelial tissue homeostasis in many systems. Because the division plane is largely set by the position of the anaphase spindle, it is important to understand how spindles become oriented. While several molecules have been identified that play key roles in spindle orientation across systems, most notably Mud/NuMA and cortical dynein, the precise mechanism by which spindles detect and align with the long cell axis remain poorly understood. Here, in exploring the dynamics of spindle orientation in mechanically distinct regions of the fly notum, we find that the ability of cells to properly reorient their divisions depends on local tissue tension. Thus, spindles reorient to align with the long cell axis in regions where isotropic tension is elevated, but fail to do so in elongated cells within the crowded midline, where tension is low, or in regions that have been mechanically isolated from the rest of the tissue via laser ablation. Importantly, these differences in spindle behavior outside and inside the midline can be recapitulated by corresponding changes in tension induced by perturbations that alter nonmuscle myosin II activity. These data lead us to propose that isotropic tension within an epithelium provides cells with a mechanically stable substrate upon which localized cortical motor complexes can act on astral microtubules to orient the spindle.


Assuntos
Drosophila/metabolismo , Miosina Tipo II/metabolismo , Fuso Acromático/metabolismo , Animais , Drosophila/fisiologia , Fenômenos Mecânicos , Miosina Tipo II/química
20.
Dev Cell ; 52(2): 210-222.e7, 2020 01 27.
Artigo em Inglês | MEDLINE | ID: mdl-31928973

RESUMO

Most metazoan cells entering mitosis undergo characteristic rounding, which is important for accurate spindle positioning and chromosome separation. Rounding is driven by contractile tension generated by myosin motors in the sub-membranous actin cortex. Recent studies highlight that alongside myosin activity, cortical actin organization is a key regulator of cortex tension. Yet, how mitotic actin organization is controlled remains poorly understood. To address this, we characterized the F-actin interactome in spread interphase and round mitotic cells. Using super-resolution microscopy, we then screened for regulators of cortex architecture and identified the intermediate filament vimentin and the actin-vimentin linker plectin as unexpected candidates. We found that vimentin is recruited to the mitotic cortex in a plectin-dependent manner. We then showed that cortical vimentin controls actin network organization and mechanics in mitosis and is required for successful cell division in confinement. Together, our study highlights crucial interactions between cytoskeletal networks during cell division.


Assuntos
Citoesqueleto de Actina/fisiologia , Actinas/metabolismo , Fenômenos Fisiológicos Celulares , Filamentos Intermediários/fisiologia , Interfase/fisiologia , Mitose , Vimentina/metabolismo , Segregação de Cromossomos , Células HeLa , Humanos
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