FilGAP and its close relatives: a mediator of Rho–Rac antagonism that regulates cell morphology and migration

2013 ◽  
Vol 453 (1) ◽  
pp. 17-25 ◽  
Author(s):  
Fumihiko Nakamura
Keyword(s):  
Actin Cytoskeleton ◽  
Gene Product ◽  
Rho Kinase ◽  
Leading Edge ◽  
Filamin A ◽  
Filamentous Actin ◽  
Rhoa Activity ◽  
Cell Protrusion ◽  
And Migration ◽  
Close Relatives

Cell migration, phagocytosis and cytokinesis are mechanically intensive cellular processes that are mediated by the dynamic assembly and contractility of the actin cytoskeleton. GAPs (GTPase-activating proteins) control activities of the Rho family proteins including Cdc42, Rac1 and RhoA, which are prominent upstream regulators of the actin cytoskeleton. The present review concerns a class of Rho GAPs, FilGAP (ARHGAP24 gene product) and its close relatives (ARHGAP22 and AHRGAP25 gene products). FilGAP is a GAP for Rac1 and a binding partner of FLNa (filamin A), a widely expressed F-actin (filamentous actin)-cross-linking protein that binds many different proteins that are important in cell regulation. Phosphorylation of FilGAP serine/threonine residues and binding to FLNa modulate FilGAP's GAP activity and, as a result, its ability to regulate cell protrusion and spreading. FLNa binds to FilGAP at F-actin-enriched sites, such as at the leading edge of the cell where Rac1 activity is controlled to inhibit actin assembly. FilGAP then dissociates from FLNa in actin networks by myosin-dependent mechanical deformation of FLNa's FilGAP-binding site to relocate at the plasma membrane by binding to polyphosphoinositides. Since actomyosin contraction is activated downstream of RhoA–ROCK (Rho-kinase), RhoA activity regulates Rac1 through FilGAP by signalling to the force-generating system. FilGAP and the ARHGAP22 gene product also act as mediators between RhoA and Rac1 pathways, which lead to amoeboid and mesenchymal modes of cell movements respectively. Therefore FilGAP and its close relatives are key regulators that promote the reciprocal inhibitory relationship between RhoA and Rac1 in cell shape changes and the mesenchymal–amoeboid transition in tumour cells.

Lysophosphatidic acid-mediated augmentation of cardiomyocyte lipoprotein lipase involves actin cytoskeleton reorganization

2005 ◽  
Vol 288 (6) ◽  
pp. H2802-H2810 ◽  
Author(s):  
Thomas Pulinilkunnil ◽  
Ding An ◽  
Sanjoy Ghosh ◽  
Dake Qi ◽  
Girish Kewalramani ◽  
...  
Keyword(s):  
Actin Cytoskeleton ◽  
Lipoprotein Lipase ◽  
Lysophosphatidic Acid ◽  
Actin Polymerization ◽  
Contractile Function ◽  
Rho Kinase ◽  
Filamentous Actin ◽  
Membrane Translocation ◽  
Downstream Effector ◽  
Cytoskeleton Reorganization

The lipoprotein lipase (LPL)-augmenting property of lysophosphatidylcholine requires the formation of lysophosphatidic acid (LPA) ( J Mol Cell Cardiol 37: 931–938, 2004). Given that the actin cytoskeleton has been implicated in regulating cardiomyocyte LPL, we examined whether LPL secretion after LPA involves actin cytoskeleton reassembly. Incubation of myocytes with LPA (1–100 nM) increased basal and heparin-releasable LPL (HR-LPL), an effect that was independent of shifts in LPL mRNA. The influence of LPA on myocyte LPL was reflected at the coronary lumen, with substantial increases of the enzyme at this location. Incubation of myocytes with cytochalasin D not only blocked LPA-induced augmentation of HR-LPL but also abrogated filamentous actin formation. These effects of LPA were likely receptor mediated. Exposure of myocytes to LPA facilitated significant membrane translocation of RhoA and its downstream effector Rho kinase I (ROCK I), and blocking this effect with Y-27632 appreciably reduced basal and HR-LPL activity. Incubation of adipose tissue with LPA also significantly enhanced basal and HR-LPL activity, suggesting that sarcomeric actin likely has a limited role in influencing the LPL secretory function of LPA in the myocyte. Comparable to LPA, hyperglycemia also caused significant membrane translocation of RhoA and ROCK I in hearts isolated from diazoxide-treated animals, effects that were abrogated using insulin. Overall, our data suggest that comparable to hyperglycemia, LPA-induced increases in cardiac LPL occurred via posttranscriptional mechanisms and processes that likely required RhoA activation and actin polymerization. Whether this increase in LPL augments triglyceride deposition in the heart leading to eventual impairment in contractile function is currently unknown.

scholarly journals Pak1 regulates focal adhesion strength, myosin IIA distribution, and actin dynamics to optimize cell migration

2011 ◽  
Vol 193 (7) ◽  
pp. 1289-1303 ◽  
Author(s):  
Violaine D. Delorme-Walker ◽  
Jeffrey R. Peterson ◽  
Jonathan Chernoff ◽  
Clare M. Waterman ◽  
Gaudenz Danuser ◽  
...  
Keyword(s):  
Cell Migration ◽  
Cell Motility ◽  
Adhesion Strength ◽  
Focal Adhesion ◽  
Leading Edge ◽  
Actin Dynamics ◽  
Filamentous Actin ◽  
Downstream Effectors ◽  
Myosin Iia ◽  
And Migration

Cell motility requires the spatial and temporal coordination of forces in the actomyosin cytoskeleton with extracellular adhesion. The biochemical mechanism that coordinates filamentous actin (F-actin) assembly, myosin contractility, adhesion dynamics, and motility to maintain the balance between adhesion and contraction remains unknown. In this paper, we show that p21-activated kinases (Paks), downstream effectors of the small guanosine triphosphatases Rac and Cdc42, biochemically couple leading-edge actin dynamics to focal adhesion (FA) dynamics. Quantitative live cell microscopy assays revealed that the inhibition of Paks abolished F-actin flow in the lamella, displaced myosin IIA from the cell edge, and decreased FA turnover. We show that, by controlling the dynamics of these three systems, Paks regulate the protrusive activity and migration of epithelial cells. Furthermore, we found that expressing Pak1 was sufficient to overcome the inhibitory effects of excess adhesion strength on cell motility. These findings establish Paks as critical molecules coordinating cytoskeletal systems for efficient cell migration.

Rho GTPases and the control of cell behaviour

2005 ◽  
Vol 33 (5) ◽  
pp. 891-895 ◽  
Author(s):  
A. Hall
Keyword(s):  
Actin Cytoskeleton ◽  
Rho Gtpases ◽  
Signal Transduction Pathway ◽  
Small Gtpases ◽  
Membrane Receptors ◽  
Mitogen Activated Protein Kinase ◽  
Cell Periphery ◽  
Filamentous Actin ◽  
Cell Behaviour ◽  
Cell Protrusion

Rho, Rac and Cdc42, three members of the Rho family of small GTPases, each control a signal transduction pathway linking membrane receptors to the assembly and disassembly of the actin cytoskeleton and of associated integrin adhesion complexes. Rho regulates stress fibre and focal adhesion assembly, Rac regulates the formation of lamellipodia protrusions and membrane ruffles, and Cdc42 triggers filopodial extensions at the cell periphery. These observations have led to the suggestion that wherever filamentous actin is used to drive a cellular process, Rho GTPases are likely to play an important regulatory role. Rho GTPases have also been reported to control other cellular activities, such as the JNK (c-Jun N-terminal kinase) and p38 MAPK (mitogen-activated protein kinase) cascades, an NADPH oxidase enzyme complex, the transcription factors NF-κB (nuclear factor κB) and SRF (serum-response factor), and progression through G1 of the cell cycle. Thus Rho, Rac and Cdc42 can regulate the actin cytoskeleton and gene transcription to promote co-ordinated changes in cell behaviour. We have been analysing the biochemical contributions of Rho GTPases in cell movement and have found that Rac controls cell protrusion, while Cdc42 controls cell polarity.

scholarly journals Shear stress–induced endothelial cell polarization is mediated by Rho and Rac but not Cdc42 or PI 3-kinases

2003 ◽  
Vol 161 (2) ◽  
pp. 429-439 ◽  
Author(s):  
Beata Wojciak-Stothard ◽  
Anne J. Ridley
Keyword(s):  
Shear Stress ◽  
Cell Shape ◽  
Rho Kinase ◽  
Cell Movement ◽  
Cell Polarization ◽  
Migration Speed ◽  
Rhoa Activity ◽  
Endothelial Response ◽  
And Migration ◽  
Shape Changes

Shear stress induces endothelial polarization and migration in the direction of flow accompanied by extensive remodeling of the actin cytoskeleton. The GTPases RhoA, Rac1, and Cdc42 are known to regulate cell shape changes through effects on the cytoskeleton and cell adhesion. We show here that all three GTPases become rapidly activated by shear stress, and that each is important for different aspects of the endothelial response. RhoA was activated within 5 min after stimulation with shear stress and led to cell rounding via Rho-kinase. Subsequently, the cells respread and elongated within the direction of shear stress as RhoA activity returned to baseline and Rac1 and Cdc42 reached peak activation. Cell elongation required Rac1 and Cdc42 but not phosphatidylinositide 3-kinases. Cdc42 and PI3Ks were not required to establish shear stress–induced polarity although they contributed to optimal migration speed. Instead, Rho and Rac1 regulated directionality of cell movement. Inhibition of Rho or Rho-kinase did not affect the cell speed but significantly increased cell displacement. Our results show that endothelial cells reorient in response to shear stress by a two-step process involving Rho-induced depolarization, followed by Rho/Rac-mediated polarization and migration in the direction of flow.

Plasticity of the actin cytoskeleton in response to extracellular matrix nanostructure and dimensionality

2014 ◽  
Vol 42 (5) ◽  
pp. 1356-1366 ◽  
Author(s):  
Josefine Starke ◽  
Bernhard Wehrle-Haller ◽  
Peter Friedl
Keyword(s):  
Extracellular Matrix ◽  
Actin Cytoskeleton ◽  
Shape Change ◽  
Leading Edge ◽  
High Speeds ◽  
Efficient Induction ◽  
Important Input ◽  
Single Force ◽  
And Migration

Mobile cells discriminate and adapt to mechanosensory input from extracellular matrix (ECM) topographies to undergo actin-based polarization, shape change and migration. We tested ‘cell-intrinsic’ and adaptive components of actin-based cell migration in response to widely used in vitro collagen-based substrates, including a continuous 2D surface, discontinuous fibril-based surfaces (2.5D) and fibril-based 3D geometries. Migrating B16F1 mouse melanoma cells expressing GFP–actin developed striking diversity and adaptation of cytoskeletal organization and migration efficacy in response to collagen organization. 2D geometry enabled keratinocyte-like cell spreading and lamellipod-driven motility, with barrier-free movement averaging the directional vectors from one or several leading edges. 3D fibrillar collagen imposed spindle-shaped polarity with a single cylindrical actin-rich leading edge and terminal filopod-like protrusions generating a single force vector. As a mixed phenotype, 2.5D environments prompted a broad but fractalized leading lamella, with multiple terminal filopod-like protrusions engaged with collagen fibrils to generate an average directional vector from multiple, often divergent, interactions. The migratory population reached >90% of the cells with high speeds for 2D, but only 10–30% of the cells and a 3-fold lower speed range for 2.5D and 3D substrates, suggesting substrate continuity as a major determinant of efficient induction and maintenance of migration. These findings implicate substrate geometry as an important input for plasticity and adaptation of the actin cytoskeleton to cope with varying ECM topography and highlight striking preference of moving cells for 2D continuous-shaped over more complex-shaped discontinuous 2.5 and 3D substrate geometries.

P190-B RhoGAP Is Critical for Hematopoietic Stem/Progenitor Cell Homing and Short Term Engraftment.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 1341-1341
Author(s):  
Haiming Xu ◽  
Kathleen Szczur ◽  
Yi Zheng ◽  
Jeffrey Settleman ◽  
David A. Williams ◽  
...  
Keyword(s):  
Cell Cycle ◽  
B Cells ◽  
Progenitor Cell ◽  
Leading Edge ◽  
Short Term ◽  
Hematopoietic Stem ◽  
Rhoa Activity ◽  
Membrane Protrusions ◽  
And Migration

Abstract Hematopoietic stem/progenitor cell (HSC/P) engraftment is a complex process which requires HSC/P to migrate across endothelium barrier from the blood towards the bone marrow (BM) cavities. The cellular and molecular mechanisms that regulate HSC/P engraftment are still poorly understood. Rho GTPases, Rac1, Rac2, CDC42 and RhoA, are major regulators of cell adhesion and migration via cytoskeleton rearrangement. While the roles of Rac and CDC42 in HSC/P functions have begun to be understood (Gu, Science 2003; Wang, Blood 2006), the role of RhoA has yet to be examined in physiological settings. To examine the role of RhoA in HSC/P engraftment, we used mice genetically deficient in the Rho-inhibitory protein p190-B RhoGAP (p190-B), which represent gain of RhoA activity (Sordella, Dev Cell, 2000). P190-B-deficient c-Kit+ cells demonstrated increased short-term engraftment in non-obese diabetic/severe combined-immunodeficiency (NOD/SCID) mice compared to WT cells. In addition, p190-B−/− was associated with increased colony forming-unit (CFU) homing to BM compared with WT (7.5%±1.8% vs 4.2%±0.6%, p<0.05). We next examined HSC/P adhesion and migration. P190-B−/− FL cells showed significantly increased CFU adhesion to fibronectin (FN) compared to WT FL cells (11.2%±1.2% vs 8.5%±1.02%, p<0.05), despite normal beta-1 integrin expression. Interestingly, p190-B−/− cells exhibited significantly increased CFU migration towards SDF-1α in the presence of FN (44.9%±1.3% vs 25.9%±4.2%, p<0.01). Cytoskeleton reorganization is critical during cell migration. To further investigate the role of RhoA activity in HSC/P migration at a mechanistic level, F-actin reorganization was examined. Upon integrin ligation and SDF-1α stimulation, c-Kit+ WT cells polarized with a single F-actin lamellipodia at the leading edge and a short uropod. In contrast, p190-B−/− showed multiple membrane protrusions of F-actin at the leading edge associated with abnormally elongated uropod in a significantly number of cells (16% vs 1%, p<0.05). In addition, in some p190-B−/− cells F-actin was assembled in a ring structure at the leading edge. Thus, RhoA may regulate HSC/P migration and homing by stimulating membrane protrusions via integrin signaling. HSC/P migration into BM cavities is restricted to the cells in the G0/G1-phase of the cell cycle. This restriction may be dependent on adhesion properties of these cells (Giet, Blood 1997; Orchell-Traycoff, Blood 2000). RhoA has been implicated in the coordination of cell cycle transit and migration in non hematopoietic cells. To test whether the increased CFU migration of p190-B-deficient cells was due to the ability of cells in cycle to migrate, c-Kit+ cells from WT and p190-B+/− BM were fractionated using the DNA dye Hoechst which allows separation of cells in G0/G1 versus S/G2+M. Remarkably, FN-induced migration of S/G2+M c-Kit+ p190-B+/− cells was significantly higher than that of WT cells. Therefore, this study suggests that the regulation of RhoA activity via p190-B is important for HSC/P short-term engraftment and homing maybe by coordinating migration and cell cycle transit and that rearrangement of the cytoskeleton likely plays an essential role during this process.

scholarly journals Molecular Tuning of Filamin A Activities in the Context of Adhesion and Migration

2020 ◽  
Vol 8 ◽  
Author(s):  
Isabelle Lamsoul ◽  
Loïc Dupré ◽  
Pierre G. Lutz
Keyword(s):  
Actin Cytoskeleton ◽  
Adult Life ◽  
Scaffolding Protein ◽  
Actin Binding ◽  
Filamin A ◽  
Multiple Organs ◽  
Human Disorders ◽  
Cross Links ◽  
Orthogonal Orientation ◽  
And Migration

The dynamic organization of actin cytoskeleton meshworks relies on multiple actin-binding proteins endowed with distinct actin-remodeling activities. Filamin A is a large multi-domain scaffolding protein that cross-links actin filaments with orthogonal orientation in response to various stimuli. As such it plays key roles in the modulation of cell shape, cell motility, and differentiation throughout development and adult life. The essentiality and complexity of Filamin A is highlighted by mutations that lead to a variety of severe human disorders affecting multiple organs. One of the most conserved activity of Filamin A is to bridge the actin cytoskeleton to integrins, thereby maintaining the later in an inactive state. We here review the numerous mechanisms cells have developed to adjust Filamin A content and activity and focus on the function of Filamin A as a gatekeeper to integrin activation and associated adhesion and motility.

scholarly journals RhoA Inactivation by p190RhoGAP Regulates Cell Spreading and Migration by Promoting Membrane Protrusion and Polarity

2001 ◽  
Vol 12 (9) ◽  
pp. 2711-2720 ◽  
Author(s):  
William T. Arthur ◽  
Keith Burridge
Keyword(s):  
Extracellular Matrix ◽  
Biological Significance ◽  
Cell Spreading ◽  
Small Gtpases ◽  
Dominant Negative ◽  
Wild Type ◽  
Rho Proteins ◽  
Rhoa Activity ◽  
Cell Protrusion ◽  
And Migration

The binding of extracellular matrix proteins to integrins triggers rearrangements in the actin cytoskeleton by regulating the Rho family of small GTPases. The signaling events that mediate changes in the activity of Rho proteins in response to the extracellular matrix remain largely unknown. We have demonstrated in previous studies that integrin signaling transiently suppresses RhoA activity through stimulation of p190RhoGAP. Here, we investigated the biological significance of adhesion-dependent RhoA inactivation by manipulating p190RhoGAP signaling in Rat1 fibroblasts. The inhibition of RhoA activity that is induced transiently by adhesion was antagonized by expression of dominant negative p190RhoGAP. This resulted in impaired cell spreading on a fibronectin substrate, reduced cell protrusion, and premature assembly of stress fibers. Conversely, overexpression of p190RhoGAP augmented cell spreading. Dominant negative p190RhoGAP elevated RhoA activity in cells on fibronectin and inhibited migration, whereas overexpression of the wild-type GAP decreased RhoA activity, promoted the formation of membrane protrusions, and enhanced motility. Cells expressing dominant negative p190RhoGAP, but not control cells or cells overexpressing the wild-type GAP, were unable to establish polarity in the direction of migration. Taken together, these data demonstrate that integrin-triggered RhoA inhibition by p190RhoGAP enhances spreading and migration by regulating cell protrusion and polarity.

Abstract 732: The P2Y 2 Receptor Selectively Binds to Filamin A to Regulate Spreading and Migration of Vascular Smooth Muscle Cells

Circulation ◽  
2007 ◽  
Vol 116 (suppl_16) ◽  
Author(s):  
Ningpu Yu ◽  
Gerardo E Fernandez ◽  
Gary A Weisman ◽  
Cheikh I Seye
Keyword(s):  
Smooth Muscle ◽  
Cell Migration ◽  
Vascular Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Actin Cytoskeleton ◽  
Vascular Smooth Muscle Cells ◽  
Filamin A ◽  
Wild Type ◽  
Adenoviral Infection ◽  
And Migration

The functional expression of the G protein-coupled P2Y 2 nucleotide receptor has been associated with the development of intimal lesions. Activation of this receptor also stimulates actin cytoskeleton reorganization and migration of vascular smooth muscle cells (SMCs). Since cell migration has been linked to the dynamic reorganization of the actin cytoskeleton, which transmits biochemical signals and forces necessary for cell locomotion, the aim of the present study was to identify cytoskeletal proteins that bind to the P2Y 2 receptor and potentially regulate SMC migration. Using the yeast two-hybrid system, we isolated filamin A, a filamentous actin-cross linking protein that interacts with the C-terminal domain of the P2Y 2 receptor and we used deletion mapping to identify amino acid deletion in the P2Y 2 receptor that led to selective loss of filamin A binding. Ex-vivo treatment of aortic explants with the P2Y 2 receptor agonist UTP (10 μmol/L) significantly promoted migration of SMCs in wild type but not in P2Y 2 receptor −/− mice. Likewise, using a Transwell migration assay we showed that UTP increased migration of SMCs in wild type (3.5 folds, P<0.01 vs. untreated cells) but not P2Y 2 receptor −/− mice (P<0.4 vs. untreated cells). Adenoviral infection of the full length P2Y 2 receptor restored UTP-mediated cell migration in P2Y 2 receptor −/− mice whereas infection of a mutant P2Y 2 receptor that does not bind filamin A did not. UTP-induced migration was preceded by a rapid phosphorylation of filamin A that was not observed either in P2Y 2 receptor −/− SMCs or in P2Y 2 receptor −/− SMCs infected with the mutant P2Y 2 receptor that does not bind filamin A. Treatment of SMCs from wild type mice with UTP (10 μmol/L) caused a 4-fold increase in spreading to collagen I as compared to unstimulated cells. The UTP-mediated increase of SMC spreading was not found in P2Y 2 receptor −/− SMCs but was restored by adenoviral infection of the full length P2Y 2 receptor cDNA into these cells. However, adenoviral delivery of a mutant receptor −/− which does not bind to filamin A did not restore UTP-mediated spreading of P2Y 2 receptor SMCs to collagen. This study demonstrates that P2Y 2 -dependent modulation of the actin cytoskeleton selectively regulates spreading and migration of SMC.

scholarly journals Actin retrograde flow actively aligns and orients ligand-engaged integrins in focal adhesions

2017 ◽  
Vol 114 (40) ◽  
pp. 10648-10653 ◽  
Author(s):  
Vinay Swaminathan ◽  
Joseph Mathew Kalappurakkal ◽  
Shalin B. Mehta ◽  
Pontus Nordenfelt ◽  
Travis I. Moore ◽  
...  
Keyword(s):  
Actin Cytoskeleton ◽  
Focal Adhesions ◽  
Leading Edge ◽  
Retrograde Flow ◽  
Molecular Scaffold ◽  
Cell Adhesion And Migration ◽  
Αvβ3 Integrins ◽  
Mediate Cell ◽  
And Migration ◽  
Migrating Cells

Integrins are transmembrane receptors that, upon activation, bind extracellular ligands and link them to the actin filament (F-actin) cytoskeleton to mediate cell adhesion and migration. Cytoskeletal forces in migrating cells generated by polymerization- or contractility-driven “retrograde flow” of F-actin from the cell leading edge have been hypothesized to mediate integrin activation for ligand binding. This predicts that these forces should align and orient activated, ligand-bound integrins at the leading edge. Here, polarization-sensitive fluorescence microscopy of GFP-αVβ3 integrins in fibroblasts shows that integrins are coaligned in a specific orientation within focal adhesions (FAs) in a manner dependent on binding immobilized ligand and a talin-mediated linkage to the F-actin cytoskeleton. These findings, together with Rosetta modeling, suggest that integrins in FA are coaligned and may be highly tilted by cytoskeletal forces. Thus, the F-actin cytoskeleton sculpts an anisotropic molecular scaffold in FAs, and this feature may underlie the ability of migrating cells to sense directional extracellular cues.

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