scholarly journals Dia-Interacting Protein Modulates Formin-Mediated Actin Assembly at the Cell Cortex

2007 ◽  
Vol 17 (7) ◽  
pp. 579-591 ◽  
Author(s):  
Kathryn M. Eisenmann ◽  
Elizabeth S. Harris ◽  
Susan M. Kitchen ◽  
Holly A. Holman ◽  
Henry N. Higgs ◽  
...  
Keyword(s):  
Cell Cortex ◽  
Actin Assembly ◽  
Interacting Protein

Osmotic stress-induced remodeling of the cortical cytoskeleton

2002 ◽  
Vol 283 (3) ◽  
pp. C850-C865 ◽  
Author(s):  
Caterina Di Ciano ◽  
Zilin Nie ◽  
Katalin Szászi ◽  
Alison Lewis ◽  
Takehito Uruno ◽  
...  
Keyword(s):  
Osmotic Stress ◽  
De Novo ◽  
Small Gtpases ◽  
Dominant Negative ◽  
Actin Binding ◽  
Cell Cortex ◽  
Actin Assembly ◽  
Signaling Mechanism ◽  
Actin Binding Domain ◽  
Cytoskeleton Remodeling

Osmotic stress is known to affect the cytoskeleton; however, this adaptive response has remained poorly characterized, and the underlying signaling pathways are unexplored. Here we show that hypertonicity induces submembranous de novo F-actin assembly concomitant with the peripheral translocation and colocalization of cortactin and the actin-related protein 2/3 (Arp2/3) complex, which are key components of the actin nucleation machinery. Additionally, hyperosmolarity promotes the association of cortactin with Arp2/3 as revealed by coimmunoprecipitation. Using various truncation or phosphorylation-incompetent mutants, we show that cortactin translocation requires the Arp2/3- or the F-actin binding domain, but the process is independent of the shrinkage-induced tyrosine phosphorylation of cortactin. Looking for an alternative signaling mechanism, we found that hypertonicity stimulates Rac and Cdc42. This appears to be a key event in the osmotically triggered cytoskeletal reorganization, because 1) constitutively active small GTPases translocate cortactin, 2) Rac and cortactin colocalize at the periphery of hypertonically challenged cells, and 3) dominant-negative Rac and Cdc42 inhibit the hypertonicity-provoked cortactin and Arp3 translocation. The Rho family-dependent cytoskeleton remodeling may be an important osmoprotective response that reinforces the cell cortex.

scholarly journals Actin dynamics coupled to clathrin-coated vesicle formation at the trans-Golgi network

2004 ◽  
Vol 165 (6) ◽  
pp. 781-788 ◽  
Author(s):  
Sebastien Carreno ◽  
Åsa E. Engqvist-Goldstein ◽  
Claire X. Zhang ◽  
Kent L. McDonald ◽  
David G. Drubin
Keyword(s):  
Time Lapse ◽  
Actin Dynamics ◽  
Coated Vesicle ◽  
Cell Cortex ◽  
Actin Assembly ◽  
Trans Golgi Network ◽  
Lysosome Biogenesis ◽  
Diverse Species ◽  
Golgi Network ◽  
Golgi Organization

In diverse species, actin assembly facilitates clathrin-coated vesicle (CCV) formation during endocytosis. This role might be an adaptation specific to the unique environment at the cell cortex, or it might be fundamental, facilitating CCV formation on different membranes. Proteins of the Sla2p/Hip1R family bind to actin and clathrin at endocytic sites in yeast and mammals. We hypothesized that Hip1R might also coordinate actin assembly with clathrin budding at the trans-Golgi network (TGN). Using deconvolution and time-lapse microscopy, we showed that Hip1R is present on CCVs emerging from the TGN. These vesicles contain the mannose 6-phosphate receptor involved in targeting proteins to the lysosome, and the actin nucleating Arp2/3 complex. Silencing of Hip1R expression by RNAi resulted in disruption of Golgi organization and accumulation of F-actin structures associated with CCVs on the TGN. Hip1R silencing and actin poisons slowed cathepsin D exit from the TGN. These studies establish roles for Hip1R and actin in CCV budding from the TGN for lysosome biogenesis.

scholarly journals Delay of HeLa cell cleavage into interphase using dihydrocytochalasin B: retention of a postmitotic spindle and telophase disc correlates with synchronous cleavage recovery.

1995 ◽  
Vol 131 (1) ◽  
pp. 191-205 ◽  
Author(s):  
S N Martineau ◽  
P R Andreassen ◽  
R L Margolis
Keyword(s):  
Mammalian Cell ◽  
Mitotic Spindle ◽  
Cyclin B ◽  
Cell Cortex ◽  
Cleavage Furrow ◽  
Actin Assembly ◽  
Cell Cleavage ◽  
Integral Structure ◽  
G1 Cells ◽  
Molecular Signals

The molecular signals that determine the position and timing of the cleavage furrow during mammalian cell cytokinesis are presently unknown. We have studied in detail the effect of dihydrocytochalasin B (DCB), a drug that interferes with actin assembly, on specific late mitotic events in synchronous HeLa cells. When cleavage furrow formation is blocked at 10 microM DCB, cells return to interphase by the criteria of reformation of nuclei with lamin borders, degradation of the cyclin B component of p34cdc2 kinase, and loss of mitosis specific MPM-2 antigens. However, the machinery for cell cleavage is retained for up to one hour into G1 when cleavage cannot proceed. The components retained consist prominently of a "postmitotic" spindle and a telophase disc, a structure templated by the mitotic spindle in anaphase that may determine the position and timing of the cleavage furrow. Upon release from DCB block, G1 cells proceed through a rapid and synchronous cleavage. We conclude that the mitotic spindle is not inevitably destroyed at the end of mitosis, but persists as an integral structure with the telophase disc in the absence of cleavage. We also conclude that cell cleavage can occur in G1, and is therefore an event metabolically independent of mitosis. The retained telophase disc may indeed signal the position of furrow formation, as G1 cleavage occurs only in the position where the retained disc underlies the cell cortex. The protocol we describe should now enable development of a model system for the study of mammalian cell cleavage as a synchronous event independent of mitosis.

scholarly journals Filopodia formation and endosome clustering induced by mutant plus-end–directed myosin VI

2017 ◽  
Vol 114 (7) ◽  
pp. 1595-1600 ◽  
Author(s):  
Thomas A. Masters ◽  
Folma Buss
Keyword(s):  
Actin Filaments ◽  
Leucine Zipper ◽  
Adaptor Protein ◽  
Cell Cortex ◽  
Myosin Vi ◽  
Actin Network ◽  
Ph Domain ◽  
Cortical Actin ◽  
Directed Movement ◽  
Interacting Protein

Myosin VI (MYO6) is the only myosin known to move toward the minus end of actin filaments. It has roles in numerous cellular processes, including maintenance of stereocilia structure, endocytosis, and autophagosome maturation. However, the functional necessity of minus-end–directed movement along actin is unclear as the underlying architecture of the local actin network is often unknown. To address this question, we engineered a mutant of MYO6, MYO6+, which undergoes plus-end–directed movement while retaining physiological cargo interactions in the tail. Expression of this mutant motor in HeLa cells led to a dramatic reorganization of cortical actin filaments and the formation of actin-rich filopodia. MYO6 is present on peripheral adaptor protein, phosphotyrosine interacting with PH domain and leucine zipper 1 (APPL1) signaling endosomes and MYO6+ expression causes a dramatic relocalization and clustering of this endocytic compartment in the cell cortex. MYO6+ and its adaptor GAIP interacting protein, C terminus (GIPC) accumulate at the tips of these filopodia, while APPL1 endosomes accumulate at the base. A combination of MYO6+ mutagenesis and siRNA-mediated depletion of MYO6 binding partners demonstrates that motor activity and binding to endosomal membranes mediated by GIPC and PI(4,5)P2 are crucial for filopodia formation. A similar reorganization of actin is induced by a constitutive dimer of MYO6+, indicating that multimerization of MYO6 on endosomes through binding to GIPC is required for this cellular activity and regulation of actin network structure. This unique engineered MYO6+ offers insights into both filopodia formation and MYO6 motor function at endosomes and at the plasma membrane.

scholarly journals Enteropathogenic Escherichia coli and Vaccinia Virus Do Not Require the Family of WASP-Interacting Proteins for Pathogen-Induced Actin Assembly

2012 ◽  
Vol 80 (12) ◽  
pp. 4071-4077 ◽  
Author(s):  
John J. Garber ◽  
Fuminao Takeshima ◽  
Inés M. Antón ◽  
Michiko K. Oyoshi ◽  
Anna Lyubimova ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Vaccinia Virus ◽  
Family Members ◽  
Ectopic Expression ◽  
Host Cells ◽  
Actin Assembly ◽  
Human Pathogens ◽  
Interacting Protein ◽  
Content Type ◽  
Syndrome Protein

ABSTRACTThe human pathogens enteropathogenicEscherichia coli(EPEC) and vaccinia virus trigger actin assembly in host cells by activating the host adaptor Nck and the actin nucleation promoter neural Wiskott-Aldrich syndrome protein (N-WASP). EPEC translocates effector molecules into host cells via type III secretion, and the interaction between the translocated intimin receptor (Tir) and the bacterial membrane protein intimin stimulates Nck and N-WASP recruitment, leading to the formation of actin pedestals beneath adherent bacteria. Vaccinia virus also recruits Nck and N-WASP to generate actin tails that promote cell-to-cell spread of the virus. In addition to Nck and N-WASP, WASP-interacting protein (WIP) localizes to vaccinia virus tails, and inhibition of actin tail formation upon ectopic expression of WIP mutants led to the suggestion that WIP is required for this process. Similar studies of WIP mutants, however, did not affect the ability of EPEC to form actin pedestals, arguing against an essential role for WIP in EPEC-induced actin assembly. In this study, we demonstrate that Nck and N-WASP are normally recruited by vaccinia virus and EPEC in the absence of WIP, and neither WIP nor the WIP family members CR16 and WIRE/WICH are essential for pathogen induced actin assembly. In addition, although Nck binds EPEC Tir directly, N-WASP is required for its localization during pedestal formation. Overall, these data highlight similar pathogenic strategies shared by EPEC and vaccinia virus by demonstrating a requirement for both Nck and N-WASP, but not WIP or WIP family members in pathogen-induced actin assembly.

scholarly journals Rho and F-actin self-organize within an artificial cell cortex

2021 ◽  
Author(s):  
Jennifer Landino ◽  
Marcin Leda ◽  
Ani Michaud ◽  
Zachary T. Swider ◽  
Mariah Prom ◽  
...  
Keyword(s):  
Cell Division ◽  
Rho Gtpase ◽  
Cortical Excitability ◽  
Cell Cortex ◽  
List Type ◽  
Actin Assembly ◽  
Cortical Dynamics ◽  
Artificial Cell ◽  
Egg Extract ◽  
Dynamic Patterns

SummaryThe cell cortex, comprised of the plasma membrane and underlying cytoskeleton, undergoes dynamic reorganizations during a variety of essential biological processes including cell adhesion, cell migration, and cell division1,2. During cell division and cell locomotion, for example, waves of filamentous-actin (F-actin) assembly and disassembly develop in the cell cortex in a process termed “cortical excitability”3–7. In developing frog and starfish embryos, cortical excitability is generated through coupled positive and negative feedback, with rapid activation of Rho-mediated F-actin assembly followed in space and time by F-actin-dependent inhibition of Rho8,9. These feedback loops are proposed to serve as a mechanism for amplification of active Rho signaling at the cell equator to support furrowing during cytokinesis, while also maintaining flexibility for rapid error correction in response to movement of the mitotic spindle during chromosome segregation10. In this paper, we develop an artificial cortex based on Xenopus egg extract and supported lipid bilayers (SLBs), to investigate cortical Rho and F-actin dynamics11. This reconstituted system spontaneously develops two distinct dynamic patterns: singular excitable Rho and F-actin waves and non-traveling oscillatory Rho and F-actin patches. Both types of dynamic patterns have properties and dependencies similar to the cortical excitability previously characterized in vivo9. These findings directly support the longstanding speculation that the cell cortex is a self-organizing structure and present a novel approach for investigating mechanisms of Rho-GTPase-mediated cortical dynamics.HighlightsAn artificial cell cortex comprising Xenopus egg extract on a supported lipid bilayer self-organizes into complex, dynamic patterns of active Rho and F-actinWe identified two types of reconstituted cortical dynamics – excitable waves and coherent oscillationsReconstituted waves and oscillations require Rho activity and F-actin polymerization

scholarly journals Daip1, a Dictyostelium Homologue of the Yeast Actin-Interacting Protein 1, Is Involved in Endocytosis, Cytokinesis, and Motility

1999 ◽  
Vol 146 (2) ◽  
pp. 453-464 ◽  
Author(s):  
Angelika Konzok ◽  
Igor Weber ◽  
Evelyn Simmeth ◽  
Ulrike Hacker ◽  
Markus Maniak ◽  
...  
Keyword(s):  
Fluorescent Protein ◽  
Fluid Phase ◽  
Gene Replacement ◽  
Cell Cortex ◽  
Filamentous Actin ◽  
Cortical Actin ◽  
Interacting Protein ◽  
Phagocytic Cups ◽  
Green Fluorescent

The 64-kD protein DAip1 from Dictyostelium contains nine WD40-repeats and is homologous to the actin-interacting protein 1, Aip1p, from Saccharomyces cerevisiae, and to related proteins from Caenorhabditis, Physarum, and higher eukaryotes. We show that DAip1 is localized to dynamic regions of the cell cortex that are enriched in filamentous actin: phagocytic cups, macropinosomes, lamellipodia, and other pseudopodia. In cells expressing green fluorescent protein (GFP)-tagged DAip1, the protein rapidly redistributes into newly formed cortical protrusions. Functions of DAip1 in vivo were assessed using null mutants generated by gene replacement, and by overexpressing DAip1. DAip1-null cells are impaired in growth and their rates of fluid-phase uptake, phagocytosis, and movement are reduced in comparison to wild-type rates. Cytokinesis is prolonged in DAip1-null cells and they tend to become multinucleate. On the basis of similar results obtained by DAip1 overexpression and effects of latrunculin-A treatment, we propose a function for DAip1 in the control of actin depolymerization in vivo, probably through interaction with cofilin. Our data suggest that DAip1 plays an important regulatory role in the rapid remodeling of the cortical actin meshwork.

scholarly journals Inhibition of polar actin assembly by astral microtubules is required for cytokinesis

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Anan Chen ◽  
Luisa Ulloa Severino ◽  
Thomas C. Panagiotou ◽  
Trevor F. Moraes ◽  
Darren A. Yuen ◽  
...  
Keyword(s):  
Cell Division ◽  
Actin Cytoskeleton ◽  
Actin Filament ◽  
Cell Cortex ◽  
Actin Assembly ◽  
Actin Network ◽  
Contractile Ring ◽  
Actin Networks ◽  
Membrane Blebbing ◽  
Actin Isoform

AbstractDuring cytokinesis, the actin cytoskeleton is partitioned into two spatially distinct actin isoform specific networks: a β-actin network that generates the equatorial contractile ring, and a γ-actin network that localizes to the cell cortex. Here we demonstrate that the opposing regulation of the β- and γ-actin networks is required for successful cytokinesis. While activation of the formin DIAPH3 at the cytokinetic furrow underlies β-actin filament production, we show that the γ-actin network is specifically depleted at the cell poles through the localized deactivation of the formin DIAPH1. During anaphase, CLIP170 is delivered by astral microtubules and displaces IQGAP1 from DIAPH1, leading to formin autoinhibition, a decrease in cortical stiffness and localized membrane blebbing. The contemporaneous production of a β-actin contractile ring at the cell equator and loss of γ-actin from the poles is required to generate a stable cytokinetic furrow and for the completion of cell division.

scholarly journals Platelet-associated IgAs and impaired GPVI responses in platelets lacking WIP

Blood ◽  
2009 ◽  
Vol 114 (21) ◽  
pp. 4729-4737 ◽  
Author(s):  
Hervé Falet ◽  
Michael P. Marchetti ◽  
Karin M. Hoffmeister ◽  
Michel J. Massaad ◽  
Raif S. Geha ◽  
...  
Keyword(s):  
Platelet Function ◽  
Actin Assembly ◽  
Irreversible Loss ◽  
Interacting Protein ◽  
Glycoprotein Vi ◽  
Activated Platelets ◽  
Granule Secretion ◽  
And Function ◽  
Syndrome Protein

Abstract The role of the Wiskott-Aldrich syndrome protein (WASp) in platelet function is unclear because platelets that lack WASp function normally. WASp constitutively associates with WASp-interacting protein (WIP) in resting and activated platelets. The role of WIP in platelet function was investigated using mice that lack WIP or WASp. WIP knockout (KO) platelets lack WASp and thus are double deficient. WIP KO mice have a thrombocytopenia, similar to WASp KO mice, resulting in part from enhanced platelet clearance. Most WIP KO, but not WASp KO, mice evolved platelet-associated immunoglobulins (Ig) of the IgA class, which normalize their platelet survival but diminish their glycoprotein VI (GPVI) responses. Protein tyrosine phosphorylation, including that of phospholipase C-γ2, and calcium mobilization are impaired in IgA-presenting WIP KO platelets stimulated through GPVI, resulting in defects in α-granule secretion, integrin αIIbβ3 activation, and actin assembly. The anti-GPVI antibody JAQ1 induces the irreversible loss of GPVI from circulating platelets in wild-type mice, but not in WIP KO mice that bear high levels of platelet-associated IgAs. Together, the data indicate that platelet-associated IgAs negatively modulate GPVI signaling and function in WIP KO mice.

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