scholarly journals Supervillin slows cell spreading by facilitating myosin II activation at the cell periphery

2007 ◽  
Vol 120 (21) ◽  
pp. 3792-3803 ◽  
Author(s):  
N. Takizawa ◽  
R. Ikebe ◽  
M. Ikebe ◽  
E. J. Luna
Keyword(s):  
Myosin Ii ◽  
Cell Spreading ◽  
Cell Periphery

scholarly journals The lamellipodium is a myosin independent mechanosensor

2017 ◽  
Author(s):  
Patrick W. Oakes ◽  
Tamara C. Bidone ◽  
Yvonne Beckham ◽  
Austin V. Skeeters ◽  
Guillermina R. Ramirez-San Juan ◽  
...  
Keyword(s):  
Cell Attachment ◽  
Myosin Ii ◽  
Adhesion Formation ◽  
Leading Edge ◽  
Cell Spreading ◽  
Substrate Stiffness ◽  
Cell Physiology ◽  
Cell Periphery ◽  
Initial Adhesion ◽  
Kinetics Of

AbstractThe ability of adherent cells to sense changes in the mechanical properties of their extracellular environments is critical to numerous aspects of their physiology. It has been well documented that cell attachment and spreading are sensitive to substrate stiffness. Here we demonstrate that this behavior is actually biphasic, with a transition that occurs around a Young’s modulus of ∼7 kPa. Furthermore, we demonstrate that, contrary to established assumptions, this property is independent of myosin II activity. Rather, we find that cell spreading on soft substrates is inhibited due to reduced nascent adhesion formation within the lamellipodium. Cells on soft substrates display normal leading edge protrusion activity, but these protrusions are not stabilized due to impaired adhesion assembly. Enhancing integrin-ECM affinity through addition of Mn2+ recovers nascent adhesion assembly and cell spreading on soft substrates. Using a computational model to simulate nascent adhesion assembly, we find that biophysical properties of the integrin-ECM bond are optimized to stabilize interactions above a threshold matrix stiffness that is consistent with the experimentally observations. Together these results suggest that myosin II-independent forces in the lamellipodium are responsible for mechanosensation by regulating new adhesion assembly, which in turn, directly controls cell spreading. This myosin II-independent mechanism of substrate stiffness sensing could potentially regulate a number of other stiffness sensitive processes.Significance StatementCell physiology can be regulated by the mechanics of the extracellular environment. Here, we demonstrate that cell spreading is a mechanosensitive process regulated by weak forces generated at the cell periphery and independent of motor activity. We show that stiffness sensing depends on the kinetics of the initial adhesion bonds that are subjected to forces driven by protein polymerization. This work demonstrates how the binding kinetics of adhesion molecules are sensitively tuned to a range of forces that enable mechanosensation.

HVEM Analysis of Microfilament Patterns in The Margins of Tissue Cells

1980 ◽  
Vol 38 ◽  
pp. 808-811
Author(s):  
Carol Allen
Keyword(s):  
Cell Movement ◽  
Cell Spreading ◽  
Living Cells ◽  
Tissue Cell ◽  
Large Sample Size ◽  
Cell Periphery ◽  
Whole Cell ◽  
Critical Point Drying ◽  
Lamellar Region ◽  
Tissue Cells

When provided with a suitable solid substrate, tissue cells undergo a rapid conversion from the spherical form expressed in suspension culture to a characteristic flattened morphology. As a result of this conversion, called cell spreading, the cell nucleus and organelles come to occupy a central region of “deep cytoplasm” which slopes steeply into a peripheral “lamellar” region less than 1 pm thick at its outer edge and generally free of cell organelles. Cell spreading is accomplished by a continuous outward repositioning of the lamellar margins. Cell translocation on the substrate results when the activity of the lamellae on one side of the cell become dominant. When this occurs, the cell is “polarized” and moves in the direction of the “leading lamellae”. Careful analysis of tissue cell locomotion by time-lapse microphotography (1) has shown that the deformational movements of the leading lamellae occur in a repeating cycle of advance and retreat in the direction of cell movement and that the rate of such deformations are positively correlated with the speed of cell movement. In the present study, the physical basis for these movements of the cell margin has been examined by comparative light microscopy of living cells with whole-mount electron microscopy of fixed cells. Ultrastructural observations were made on tissue cells grown on Formvar-coated grids, fixed with glutaraldehyde, further processed by critical-point drying, and then photographed in the High Voltage Electron Microscope. This processing and imaging system maintains the 3-dimensional organization of the whole cell, the relationship of the cell to the substrate, and affords a large sample size which facilitates quantitative analysis. Comparative analysis of film records of living cells with the whole-cell micrographs revealed that specific patterns of microfilament organization consistently accompany recognizable stages of lamellar formation and movement. The margins of spreading cells and the leading lamellae of locomoting cells showed a similar pattern of MF repositionings (Figs. 1-4). These results will be discussed in terms of a working model for the mechanics of lamellar motility which includes the following major features: (a) lamellar protrusion results when an intracellular force is exerted at a locally weak area of the cell periphery; (b) the association of cortical MFs with one another determines the local resistance to this force; (c) where MF-to-MF association is weak, the cell periphery expands and some cortical MFs are dragged passively forward; (d) contact of the expanded area with the substrate then triggers the lateral association and reorientation of these cortical MFs into MF bundles parallel to the direction of the expansion; and (e) an active interaction between these MF bundles associated with the cortex of the expanded lamellae and the cortical MFs which remained in the sub-lamellar region then pulls the latter MFs forward toward the expanded area. Thus, the advance of the cell periphery on the substrate occurs in two stages: a passive phase in which some cortical MFs are dragged outward by the force acting to expand the cell periphery, and an active phase in which additional cortical MFs are pulled forward by interaction with the first set. Subsequent interactions between peripheral microfilament bundles and filaments in the deeper cytoplasm could then transmit the advance gained by lamellar expansion to the bulk of the cytoplasm.

scholarly journals Contribution of myosin II activity to cell spreading dynamics

Soft Matter ◽  
2016 ◽  
Vol 12 (2) ◽  
pp. 500-507 ◽  
Author(s):  
Noam Nisenholz ◽  
Aishwarya Paknikar ◽  
Sarah Köster ◽  
Assaf Zemel
Keyword(s):  
Myosin Ii ◽  
Cell Spreading ◽  
Spreading Dynamics

Coronin localizes to leading edges and is involved in cell spreading and lamellipodium extension in vertebrate cells

1999 ◽  
Vol 112 (17) ◽  
pp. 2833-2842 ◽  
Author(s):  
M. Mishima ◽  
E. Nishida
Keyword(s):  
Amino Acids ◽  
Cell Spreading ◽  
Immunofluorescent Staining ◽  
Actin Binding ◽  
Cellular Slime Mold ◽  
Cell Periphery ◽  
Terminal Amino ◽  
Wd Repeat ◽  
Leading Edges

Coronin is a WD repeat-containing actin-binding protein, which was originally identified in the cellular slime mold Dictyostelium. Coronin-null Dictyostelium cells show defects in cytokinesis, cell motility and phagocytosis. Although the existence of coronin in higher eukaryotes has been reported, its function in vertebrate cells has not been elucidated. We cloned a Xenopus homolog of coronin (Xcoronin) and examined its actin-binding properties, subcellular localization and possible functions. Xcoronin consists of 480 amino acids and is 63% identical to human coronin (p57). Bacterially expressed recombinant Xcoronin co-sedimented with F-actin in vitro. The WD repeat domain (residues 64–299) alone did not have any affinity for F-actin. Anti-Xcoronin antibodies reacted specifically with a single 57 kDa protein present in an extract of the Xenopus A6 cell line. Indirect immunofluorescent staining of A6 cells revealed that Xcoronin is present in the cytoplasm and concentrated in the cell periphery in membrane ruffles. During spreading after replating or wound healing after scratching a confluent monolayer, Xcoronin became concentrated in the leading edges of lamellipodia. A GFP-fusion protein of Xcoronin showed a subcellular distribution essentially identical to endogenous Xcoronin. The localization of Xcoronin to the cell periphery was resistant to treatment with 0.1% Triton X-100. The deletion of 63 N-terminal amino acids or of 65 C-terminal amino acids abolished the localization of Xcoronin to the cell periphery. Xcoronin expressed in 3T3 fibroblasts was concentrated to the leading edges of lamellipodia induced by active Rac. Remarkably, expression of a truncated form of Xcoronin (64–299), but not of full-length Xcoronin, significantly decreased the rate of cell spreading after replating and markedly inhibited lamellipodium extension induced by active Rac. These results suggest that Xcoronin plays an important role in lamellipodium extension and cell spreading.

scholarly journals Lamellipodium is a myosin-independent mechanosensor

2018 ◽  
Vol 115 (11) ◽  
pp. 2646-2651 ◽  
Author(s):  
Patrick W. Oakes ◽  
Tamara C. Bidone ◽  
Yvonne Beckham ◽  
Austin V. Skeeters ◽  
Guillermina R. Ramirez-San Juan ◽  
...  
Keyword(s):  
Cell Attachment ◽  
Myosin Ii ◽  
Adhesion Formation ◽  
Leading Edge ◽  
Cell Spreading ◽  
Substrate Stiffness ◽  
Adherent Cells ◽  
Matrix Stiffness ◽  
Biophysical Properties ◽  
Independent Mechanism

The ability of adherent cells to sense changes in the mechanical properties of their extracellular environments is critical to numerous aspects of their physiology. It has been well documented that cell attachment and spreading are sensitive to substrate stiffness. Here, we demonstrate that this behavior is actually biphasic, with a transition that occurs around a Young’s modulus of ∼7 kPa. Furthermore, we demonstrate that, contrary to established assumptions, this property is independent of myosin II activity. Rather, we find that cell spreading on soft substrates is inhibited due to reduced myosin-II independent nascent adhesion formation within the lamellipodium. Cells on soft substrates display normal leading-edge protrusion activity, but these protrusions are not stabilized due to impaired adhesion assembly. Enhancing integrin–ECM affinity through addition of Mn2+ recovers nascent adhesion assembly and cell spreading on soft substrates. Using a computational model to simulate nascent adhesion assembly, we find that biophysical properties of the integrin–ECM bond are optimized to stabilize interactions above a threshold matrix stiffness that is consistent with the experimental observations. Together, these results suggest that myosin II-independent forces in the lamellipodium are responsible for mechanosensation by regulating new adhesion assembly, which, in turn, directly controls cell spreading. This myosin II-independent mechanism of substrate stiffness sensing could potentially regulate a number of other stiffness-sensitive processes.

scholarly journals Myosin rings and spreading in mouse blastomeres.

1984 ◽  
Vol 99 (3) ◽  
pp. 1145-1150 ◽  
Author(s):  
J S Sobel
Keyword(s):  
Contact Region ◽  
Cell Spreading ◽  
Cell Contact ◽  
Cell Periphery ◽  
Cell Substrate ◽  
Preimplantation Mouse Embryo ◽  
Preimplantation Mouse ◽  
Filament Bundles ◽  
Surface Polysaccharides ◽  
Induced Changes

The relationship between myosin organization and cell spreading in the preimplantation mouse embryo was studied by indirect immunofluorescence in embryos cultured on lectin-coated substrates. Binding of cell surface polysaccharides to substrate-bound concanavalin A and wheat germ agglutinin induced changes in myosin distribution that resembled those which occur during cell-cell contact interaction. This involved an initial loss of myosin from the contact region that was associated with the development of stable cell-substrate attachments. In addition, a ring of myosin was formed along the edge of the cells' contact to the substrate. The presence of such a ring may be related to the potential for subsequent cell spreading. A myosin ring was also identified in the apical junctional region of the outer morula cells where it similarly separated the cell periphery into contacted and free peripheral domains. Following these changes in myosin organization the embryos spread on the substrate by extension of lamellipodia. These movements were coupled to the dissolution of the myosin ring and the reorganization of myosin into filament bundles. The sequence of changes in the pattern of myosin distribution suggests that contact regulation of myosin organization plays an important role in controlling the spreading behavior of blastomeres and perhaps more generally in the organization of cells into epithelia.

Active FHOD1 promotes the formation of functional actin stress fibers

2019 ◽  
Vol 476 (20) ◽  
pp. 2953-2963 ◽  
Author(s):  
Xuemeng Shi ◽  
Shuangshuang Zhao ◽  
Jinping Cai ◽  
Gary Wong ◽  
Yaming Jiu
Keyword(s):  
Myosin Ii ◽  
Critical Role ◽  
Active Form ◽  
Focal Adhesions ◽  
Stress Fibers ◽  
Cell Periphery ◽  
Physiological Relevance ◽  
Human Osteosarcoma Cells ◽  
And Migration ◽  
Actin Stress Fibers

Abstract The formin FHOD1 acts as a nucleating, capping and bundling protein of actin filaments. In cells, release from the C-terminal diaphanous autoregulatory domain (DAD) of FHOD1 stimulates the protein into the active form. However, the cellular physiological relevance of active form FHOD1 and the phenotypic regulation by FHOD1 depletion are not completely understood. Here, we show that in contrast with the cytosolic diffused expression of auto-inhibited FHOD1, active FHOD1 by C-terminal truncation was recruited into all three types of actin stress fibers in human osteosarcoma cells. Notably, the recruited active FHOD1 was more incorporated with myosin II than α-actinin, and associated with both naïve and mature focal adhesions. Active FHOD1 displayed faster turnover than actin molecules on ventral stress fibers. Moreover, we witnessed the emergence of active FHOD1 from the cell periphery, which subsequently moved centripetally together with transverse arcs. Furthermore, FHOD1 knockdown resulted in defective maturation of actomyosin bundles and subsequently longer non-contractile dorsal stress fibers, whereas the turnover of both actin and myosin II were maintained normally. Importantly, the loss of FHOD1 led to slower actin centripetal flow, resulting in abnormal cell spreading and migration defects. Taken together, these results reveal a critical role of FHOD1 in temporal- and spatial- control of the morphology and dynamics of functional actin stress fibers during variable cell behavior.

Myosin II-dependent cylindrical protrusions induced by quinine inDictyostelium: antagonizing effects of actin polymerization at the leading edge

2001 ◽  
Vol 114 (11) ◽  
pp. 2155-2165
Author(s):  
Kunito Yoshida ◽  
Kei Inouye
Keyword(s):  
Cell Membrane ◽  
Actin Polymerization ◽  
Myosin Ii ◽  
Cell Movement ◽  
Leading Edge ◽  
Contractile Vacuole ◽  
Cell Periphery ◽  
Slowing Down ◽  
On Line ◽  
Cylindrical Protrusion

We found that amoeboid cells of Dictyostelium are induced by a millimolar concentration of quinine to form a rapidly elongating, cylindrical protrusion, which often led to sustained locomotion of the cells. Formation of the protrusion was initiated by fusion of a contractile vacuole with the cell membrane. During protrusion extension, a patch of the contractile vacuole membrane stayed undiffused on the leading edge of the protrusion for over 30 seconds. Protrusion formation was not inhibited by high osmolarity of the external medium (at least up to 400 mosM). By contrast, mutant cells lacking myosin II (mhc− cells) failed to extend protrusions upon exposure to quinine. When GFP-myosin-expressing cells were exposed to quinine, GFP-myosin was accumulated in the cell periphery forming a layer under the cell membrane, but a newly formed protrusion was initially devoid of a GFP-myosin layer, which gradually formed and extended from the base of the protrusion. F-actin was absent in the leading front of the protrusion during the period of its rapid elongation, and the formation of a layer of F-actin in the front was closely correlated with its slowing-down or retraction. Periodical or continuous detachment of the F-actin layer from the apical membrane of the protrusion, accompanied by a transient increase in the elongation speed at the site of detachment, was observed in some of the protrusions. The detached F-actin layers, which formed a spiral layer of F-actin in the case of continuous detachment, moved in the opposite direction of protrusion elongation. In the presence of both cytochalasin A and quinine, the protrusions formed were not cylindrical but spherical, which swallowed up the entire cellular contents. The estimated bulk flux into the expanding spherical protrusions of such cells was four-times higher than the flux into the elongating cylindrical protrusions of the cells treated with quinine alone. These results indicate that the force responsible for the quinine-induced protrusion is mainly due to contraction of the cell body, which requires normal myosin II functions, while actin polymerization is important in restricting the direction of its expansion. We will discuss the possible significance of tail contraction in cell movement in the multicellular phase of Dictyostelium development, where cell locomotion similar to that induced by quinine is often observed without quinine treatment, and in protrusion elongation in general.Movies available on-line

scholarly journals Integrin-mediated Signals Regulated by Members of the Rho Family of GTPases

1998 ◽  
Vol 142 (2) ◽  
pp. 573-586 ◽  
Author(s):  
Edwin A. Clark ◽  
Warren G. King ◽  
Joan S. Brugge ◽  
Marc Symons ◽  
Richard O. Hynes
Keyword(s):  
Signaling Pathways ◽  
Matrix Protein ◽  
Focal Adhesions ◽  
Cell Spreading ◽  
Extracellular Matrix Protein ◽  
Cell Periphery ◽  
Dependent Manner ◽  
Membrane Ruffling ◽  
Initial Adhesion ◽  
Rho Family

The organization of the actin cytoskeleton can be regulated by soluble factors that trigger signal transduction events involving the Rho family of GTPases. Since adhesive interactions are also capable of organizing the actin-based cytoskeleton, we examined the role of Cdc42-, Rac-, and Rho-dependent signaling pathways in regulating the cytoskeleton during integrin-mediated adhesion and cell spreading using dominant-inhibitory mutants of these GTPases. When Rat1 cells initially adhere to the extracellular matrix protein fibronectin, punctate focal complexes form at the cell periphery. Concomitant with focal complex formation, we observed some phosphorylation of the focal adhesion kinase (FAK) and Src, which occurred independently of Rho family GTPases. However, subsequent phosphorylation of FAK and paxillin occurs in a Rho-dependent manner. Moreover, we found Rho dependence of the assembly of large focal adhesions from which actin stress fibers radiate. Initial adhesion to fibronectin also stimulates membrane ruffling; we show that this ruffling is independent of Rho but is dependent on both Cdc42 and Rac. Furthermore, we observed that Cdc42 controls the integrin-dependent activation of extracellular signal–regulated kinase 2 and of Akt, a kinase whose activity has been demonstrated to be dependent on phosphatidylinositol (PI) 3-kinase. Since Rac-dependent membrane ruffling can be stimulated by PI 3-kinase, it appears that Cdc42, PI 3-kinase, and Rac lie on a distinct pathway that regulates adhesion-induced membrane ruffling. In contrast to the differential regulation of integrin-mediated signaling by Cdc42, Rac, and Rho, we observed that all three GTPases regulate cell spreading, an event that may indirectly control cellular architecture. Therefore, several separable signaling pathways regulated by different members of the Rho family of GTPases converge to control adhesion-dependent changes in the organization of the cytoskeleton, changes that regulate cell morphology and behavior.

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