scholarly journals CLIC4 is regulated by RhoA-mDia2 signaling through Profilin-1 binding to modulate filopodium length

2018 ◽  
Author(s):  
Elisabetta Argenzio ◽  
Katarzyna M. Kedziora ◽  
Leila Nahidiazar ◽  
Tadamoto Isogai ◽  
Anastassis Perrakis ◽  
...  
Keyword(s):  
Actin Polymerization ◽  
Binding Protein ◽  
Feedback Mechanism ◽  
Actin Binding ◽  
Actin Assembly ◽  
Wild Type ◽  
Cortical Actin ◽  
Conserved Residues ◽  
Rhoa Activation ◽  
Necessary And Sufficient

AbstractCLIC4 is a cytosolic protein implicated in diverse actin-based processes, including integrin trafficking, cell adhesion and tubulogenesis. CLIC4 is rapidly recruited to the plasma membrane by G12/13-coupled receptor agonists and then partly co-localizes with β1 integrins. Receptor-mediated CLIC4 translocation depends on actin polymerization, but the mechanism and functional significance of CLIC4 trafficking are unknown. Here we show that RhoA activation by either LPA or EGF is necessary and sufficient for CLIC4 translocation, with a regulatory role for the RhoA effector mDia2, an inducer of actin polymerization. We find that CLIC4 directly interacts with the G-actin-binding protein Profilin-1 via conserved residues that are required for CLIC4 trafficking and lie in a concave surface. Consistently, silencing of Profilin-1 impaired CLIC4 trafficking induced by either LPA or EGF. CLIC4 knockdown promoted the formation of long integrin-dependent filopodia, a phenotype rescued by wild-type CLIC4 but not by trafficking-incompetent CLIC4(C35A). Our results establish CLIC4 as a Profilin-1-binding protein and suggest that CLIC4 translocation provides a feedback mechanism to modulate mDia2/Profilin-1-driven cortical actin assembly and membrane protrusion.

scholarly journals Sla1p Is a Functionally Modular Component of the Yeast Cortical Actin Cytoskeleton Required for Correct Localization of Both Rho1p-GTPase and Sla2p, a Protein with Talin Homology

1999 ◽  
Vol 10 (4) ◽  
pp. 1061-1075 ◽  
Author(s):  
Kathryn R. Ayscough ◽  
Jennifer J. Eby ◽  
Thomas Lila ◽  
Hilary Dewar ◽  
Keith G. Kozminski ◽  
...  
Keyword(s):  
Cell Walls ◽  
Binding Protein ◽  
Actin Binding ◽  
Actin Assembly ◽  
Cortical Actin ◽  
Actin Binding Protein ◽  
Actin Patch ◽  
Mutant Forms ◽  
Modular Component ◽  
Terminal Repeat Region

SLA1 was identified previously in budding yeast in a genetic screen for mutations that caused a requirement for the actin-binding protein Abp1p and was shown to be required for normal cortical actin patch structure and organization. Here, we show that Sla1p, like Abp1p, localizes to cortical actin patches. Furthermore, Sla1p is required for the correct localization of Sla2p, an actin-binding protein with homology to talin implicated in endocytosis, and the Rho1p-GTPase, which is associated with the cell wall biosynthesis enzyme β-1,3-glucan synthase. Mislocalization of Rho1p in sla1 null cells is consistent with our observation that these cells possess aberrantly thick cell walls.  Expression of mutant forms of Sla1p in which specific domains were deleted showed that the phenotypes associated with the full deletion are functionally separable. In particular, a region of Sla1p encompassing the third SH3 domain is important for growth at high temperatures, for the organization of cortical actin patches, and for nucleated actin assembly in a permeabilized yeast cell assay. The apparent redundancy between Sla1p and Abp1p resides in the C-terminal repeat region of Sla1p. A homologue of SLA1 was identified inSchizosaccharomyces pombe. Despite relatively low overall sequence homology, this gene was able to rescue the temperature sensitivity associated with a deletion of SLA1 inSaccharomyces cerevisiae.

scholarly journals Cortactin Localization to Sites of Actin Assembly in Lamellipodia Requires Interactions with F-Actin and the Arp2/3 Complex

2000 ◽  
Vol 151 (1) ◽  
pp. 29-40 ◽  
Author(s):  
Scott A. Weed ◽  
Andrei V. Karginov ◽  
Dorothy A. Schafer ◽  
Alissa M. Weaver ◽  
Andrew W. Kinley ◽  
...  
Keyword(s):  
Tandem Repeat ◽  
Actin Polymerization ◽  
Spatial Organization ◽  
Pdz Domain ◽  
Actin Binding ◽  
Cell Periphery ◽  
Actin Assembly ◽  
Cortical Actin ◽  
Amino Terminal ◽  
Receptor Complexes

Cortactin is an actin-binding protein that is enriched within the lamellipodia of motile cells and in neuronal growth cones. Here, we report that cortactin is localized with the actin-related protein (Arp) 2/3 complex at sites of actin polymerization within the lamellipodia. Two distinct sequence motifs of cortactin contribute to its interaction with the cortical actin network: the fourth of six tandem repeats and the amino-terminal acidic region (NTA). Cortactin variants lacking either the fourth tandem repeat or the NTA failed to localize at the cell periphery. Tandem repeat four was necessary for cortactin to stably bind F-actin in vitro. The NTA region interacts directly with the Arp2/3 complex based on affinity chromatography, immunoprecipitation assays, and binding assays using purified components. Cortactin variants containing the NTA region were inefficient at promoting Arp2/3 actin nucleation activity. These data provide strong evidence that cortactin is specifically localized to sites of dynamic cortical actin assembly via simultaneous interaction with F-actin and the Arp2/3 complex. Cortactin interacts via its Src homology 3 (SH3) domain with ZO-1 and the SHANK family of postsynaptic density 95/dlg/ZO-1 homology (PDZ) domain–containing proteins, suggesting that cortactin contributes to the spatial organization of sites of actin polymerization coupled to selected cell surface transmembrane receptor complexes.

scholarly journals Haematopoietic lineage cell-specific protein 1 (HS1) promotes actin-related protein (Arp) 2/3 complex-mediated actin polymerization

2003 ◽  
Vol 371 (2) ◽  
pp. 485-493 ◽  
Author(s):  
Takehito URUNO ◽  
Peijun ZHANG ◽  
Jiali LIU ◽  
Jian-Jiang HAO ◽  
Xi ZHAN
Keyword(s):  
Actin Polymerization ◽  
De Novo ◽  
Primary Component ◽  
Immunofluorescent Staining ◽  
Actin Binding ◽  
Specific Gene ◽  
Actin Assembly ◽  
Related Protein ◽  
Filamentous Actin ◽  
Cortical Actin

HS1 (haematopoietic lineage cell-specific gene protein 1), a prominent substrate of intracellular protein tyrosine kinases in haematopoietic cells, is implicated in the immune response to extracellular stimuli and in cell differentiation induced by cytokines. Although HS1 contains a 37-amino acid tandem repeat motif and a C-terminal Src homology 3 domain and is closely related to the cortical-actin-associated protein cortactin, it lacks the fourth repeat that has been shown to be essential for cortactin binding to filamentous actin (F-actin). In this study, we examined the possible role of HS1 in the regulation of the actin cytoskeleton. Immunofluorescent staining demonstrated that HS1 co-localizes in the cytoplasm of cells with actin-related protein (Arp) 2/3 complex, the primary component of the cellular machinery responsible for de novo actin assembly. Furthermore, recombinant HS1 binds directly to Arp2/3 complex with an equilibrium dissociation constant (Kd) of 880nM. Although HS1 is a modest F-actin-binding protein with a Kd of 400nM, it increases the rate of the actin assembly mediated by Arp2/3 complex, and promotes the formation of branched actin filaments induced by Arp2/3 complex and a constitutively activated peptide of N-WASP (neural Wiskott–Aldrich syndrome protein). Our data suggest that HS1, like cortactin, plays an important role in the modulation of actin assembly.

scholarly journals Actin-binding protein promotes the bipolar and perpendicular branching of actin filaments.

1980 ◽  
Vol 87 (3) ◽  
pp. 841-848 ◽  
Author(s):  
J H Hartwig ◽  
J Tyler ◽  
T P Stossel
Keyword(s):  
Actin Filaments ◽  
Actin Polymerization ◽  
Average Length ◽  
Binding Protein ◽  
Actin Binding ◽  
Branch Points ◽  
Heavy Meromyosin ◽  
Actin Binding Protein ◽  
Protein Dimers ◽  
Protein Molecules

Branching filaments with striking perpendicularity form when actin polymerizes in the presence of macrophage actin-binding protein. Actin-binding protein molecules are visible at the branch points. Compared with actin polymerized in the absence of actin-binding proteins, not only do the filaments branch but the average length of the actin filaments decreases from 3.2 to 0.63 micrometer. Arrowhead complexes formed by addition of heavy meromyosin molecules to the branching actin filaments point toward the branch points. Actin-binding protein also accelerates the onset of actin polymerization. All of these findings show that actin filaments assemble from nucleating sites on actin-binding protein dimers. A branching polymerization of actin filaments from a preexisting lattice of actin filaments joined by actin-binding protein molecules could generate expansion of cortical cytoplasm in amoeboid cells.

scholarly journals Actin-binding Protein Drebrin Regulates HIV-1-triggered Actin Polymerization and Viral Infection

2013 ◽  
Vol 288 (39) ◽  
pp. 28382-28397 ◽  
Author(s):  
Mónica Gordón-Alonso ◽  
Vera Rocha-Perugini ◽  
Susana Álvarez ◽  
Ángeles Ursa ◽  
Nuria Izquierdo-Useros ◽  
...  
Keyword(s):  
Viral Infection ◽  
Actin Polymerization ◽  
Binding Protein ◽  
Actin Binding ◽  
Actin Binding Protein ◽  
Hiv 1

scholarly journals Identification of a Novel Cortactin SH3 Domain-Binding Protein and Its Localization to Growth Cones of Cultured Neurons

1998 ◽  
Vol 18 (10) ◽  
pp. 5838-5851 ◽  
Author(s):  
Yunrui Du ◽  
Scott A. Weed ◽  
Wen-Cheng Xiong ◽  
Trudy D. Marshall ◽  
J. Thomas Parsons
Keyword(s):  
Hippocampal Neurons ◽  
Binding Protein ◽  
Consensus Sequence ◽  
Growth Cones ◽  
Sh3 Domain ◽  
Actin Binding ◽  
Sequence Motifs ◽  
Cortical Actin ◽  
Cytoskeleton Reorganization

ABSTRACT Cortactin is an actin-binding protein that contains several potential signaling motifs including a Src homology 3 (SH3) domain at the distal C terminus. Translocation of cortactin to specific cortical actin structures and hyperphosphorylation of cortactin on tyrosine have been associated with the cortical cytoskeleton reorganization induced by a variety of cellular stimuli. The function of cortactin in these processes is largely unknown in part due to the lack of information about cellular binding partners for cortactin. Here we report the identification of a novel cortactin-binding protein of approximately 180 kDa by yeast two-hybrid interaction screening. The interaction of cortactin with this 180-kDa protein was confirmed by both in vitro and in vivo methods, and the SH3 domain of cortactin was found to direct this interaction. Since this protein represents the first reported natural ligand for the cortactin SH3 domain, we designated it CortBP1 for cortactin-binding protein 1. CortBP1 contains two recognizable sequence motifs within its C-terminal region, including a consensus sequence for cortactin SH3 domain-binding peptides and a sterile alpha motif. Northern and Western blot analysis indicated that CortBP1 is expressed predominately in brain tissue. Immunofluorescence studies revealed colocalization of CortBP1 with cortactin and cortical actin filaments in lamellipodia and membrane ruffles in fibroblasts expressing CortBP1. Colocalization of endogenous CortBP1 and cortactin was also observed in growth cones of developing hippocampal neurons, implicating CortBP1 and cortactin in cytoskeleton reorganization during neurite outgrowth.

Abstract 20484: Enah/Vasp-Like Protein is a Critical Component in S1P-Mediated Endothelial Lamellipodia Formation

Circulation ◽  
2014 ◽  
Vol 130 (suppl_2) ◽  
Author(s):  
Joseph B Mascarenhas ◽  
Ghassan Mouneimne ◽  
Carol C Gregorio ◽  
Mary E Brown ◽  
Ting Wang ◽  
...  
Keyword(s):  
Actin Polymerization ◽  
Human Lung ◽  
Actin Binding ◽  
Lipid Mediator ◽  
Dependent Manner ◽  
Cortical Actin ◽  
Pulmonary Endothelial Cells ◽  
Sphingosine 1 Phosphate ◽  
Barrier Regulation ◽  
Lamellipodia Formation

Ena/VASP like protein, or EVL, is an actin-binding protein that regulates cancer cell lamellipodia protrusive activity and cell motility via an actomyosin contractility-dependent mechanism. The function of EVL in human lung endothelial cell (EC) barrier regulation, especially by the endogenous bioactive lipid mediator sphingosine-1-phosphate (S1P), is largely unknown. In this current study, we demonstrated that EVL is an active component in S1P-mediated EC barrier enhancement and lamellipodia formation. Compared to other focal adhesion (FA) proteins such as paxillin, EVL protein expression is very low in human pulmonary endothelial cells (ECs). S1P (1 μM) challenge stimulates translocation of cytosolic EVL to FAs in ECs, which was attenuated by EVL knockdown (KD) by its selective siRNA. S1P also promoted significant EVL translocation to lamellipodia, further confirmed by tracking translocation of EVL-GFP fusion protein upon S1P stimulation in a time-dependent manner. In addition, S1P-mediated cortical actin filament formation is attenuated by EVL KD, further confirming the function of EVL in S1P-induced lamellipodia formation/cortical actin polymerization. S1P stimulates EVL phosphorylation by tyrosine kinase c-Abl which is attenuated by the c-Abl inhibitor, imatinib. Finally, EVL KD attenuated S1P-mediated EC barrier enhancement and paracellular gap resealing reflected by reduced transendothelial electrical resistance (TER) measurements. These findings confirm a novel role for EVL in human lung vascular barrier enhancement and cytoskeleton rearrangement by S1P.

scholarly journals The N-Terminus ofDictyosteliumScar Interacts with Abi and HSPC300 and Is Essential for Proper Regulation and Function

2007 ◽  
Vol 18 (5) ◽  
pp. 1609-1620 ◽  
Author(s):  
Diana Caracino ◽  
Cheryl Jones ◽  
Mark Compton ◽  
Charles L. Saxe
Keyword(s):  
Actin Polymerization ◽  
Terminal Region ◽  
Wild Type ◽  
Large Molecular Weight ◽  
Weight Protein ◽  
And Function ◽  
Necessary And Sufficient

Scar/WAVE proteins, members of the conserved Wiskott-Aldrich syndrome (WAS) family, promote actin polymerization by activating the Arp2/3 complex. A number of proteins, including a complex containing Nap1, PIR121, Abi1/2, and HSPC300, interact with Scar/WAVE, though the role of this complex in regulating Scar function remains unclear. Here we identify a short N-terminal region of Dictyostelium Scar that is necessary and sufficient for interaction with HSPC300 and Abi in vitro. Cells expressing Scar lacking this N-terminal region show abnormalities in F-actin distribution, cell morphology, movement, and cytokinesis. This is true even in the presence of wild-type Scar. The data suggest that the first 96 amino acids of Scar are necessary for participation in a large-molecular-weight protein complex, and that this Scar-containing complex is responsible for the proper localization and regulation of Scar. The presence of mis-regulated or unregulated Scar has significant deleterious effects on cells and may explain the need to keep Scar activity tightly controlled in vivo either by assembly in a complex or by rapid degradation.

scholarly journals Actin-binding protein G (AbpG) participates in modulating the actin cytoskeleton and cell migration in Dictyostelium discoideum

2015 ◽  
Vol 26 (6) ◽  
pp. 1084-1097 ◽  
Author(s):  
Wei-Chi Lin ◽  
Liang-Chen Wang ◽  
Te-Ling Pang ◽  
Mei-Yu Chen
Keyword(s):  
Cell Migration ◽  
Dictyostelium Discoideum ◽  
Molecular Mechanisms ◽  
Binding Protein ◽  
Protein Domain ◽  
Actin Dynamics ◽  
Actin Binding ◽  
Protein G ◽  
Wild Type ◽  
Actin Binding Protein

Cell migration is involved in various physiological and pathogenic events, and the complex underlying molecular mechanisms have not been fully elucidated. The simple eukaryote Dictyostelium discoideum displays chemotactic locomotion in stages of its life cycle. By characterizing a Dictyostelium mutant defective in chemotactic responses, we identified a novel actin-binding protein serving to modulate cell migration and named it actin-binding protein G (AbpG); this 971–amino acid (aa) protein contains an N-terminal type 2 calponin homology (CH2) domain followed by two large coiled-coil regions. In chemoattractant gradients, abpG− cells display normal directional persistence but migrate significantly more slowly than wild-type cells; expressing Flag-AbpG in mutant cells eliminates the motility defect. AbpG is enriched in cortical/lamellipodial regions and colocalizes well with F-actin; aa 401–600 and aa 501–550 fragments of AbpG show the same distribution as full-length AbpG. The aa 501–550 region of AbpG, which is essential for AbpG to localize to lamellipodia and to rescue the phenotype of abpG− cells, is sufficient for binding to F-actin and represents a novel actin-binding protein domain. Compared with wild-type cells, abpG− cells have significantly higher F-actin levels. Collectively our results suggest that AbpG may participate in modulating actin dynamics to optimize cell locomotion.

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