scholarly journals Differential roles of α-, β-, and γ-actin in axon growth and collateral branch formation in motoneurons

2017 ◽  
Vol 216 (3) ◽  
pp. 793-814 ◽  
Author(s):  
Mehri Moradi ◽  
Rajeeve Sivadasan ◽  
Lena Saal ◽  
Patrick Lüningschrör ◽  
Benjamin Dombert ◽  
...  
Keyword(s):  
Actin Polymerization ◽  
Axon Growth ◽  
Actin Dynamics ◽  
Collateral Branch ◽  
Actin Isoforms ◽  
Spatial Regulation ◽  
Presynaptic Differentiation ◽  
Primary Mouse ◽  
Branch Formation ◽  
Specific Contribution

Axonal branching and terminal arborization are fundamental events during the establishment of synaptic connectivity. They are triggered by assembly of actin filaments along axon shafts giving rise to filopodia. The specific contribution of the three actin isoforms, Actα, Actβ, and Actγ, to filopodia stability and dynamics during this process is not well understood. Here, we report that Actα, Actβ, and Actγ isoforms are expressed in primary mouse motoneurons and their transcripts are translocated into axons. shRNA-mediated depletion of Actα reduces axonal filopodia dynamics and disturbs collateral branch formation. Knockdown of Actβ reduces dynamic movements of growth cone filopodia and impairs presynaptic differentiation. Ablation of Actβ or Actγ leads to compensatory up-regulation of the two other isoforms, which allows maintenance of total actin levels and preserves F-actin polymerization. Collectively, our data provide evidence for specific roles of different actin isoforms in spatial regulation of actin dynamics and stability in axons of developing motoneurons.

scholarly journals DSCR1 is required for both axonal growth cone extension and steering

2016 ◽  
Vol 213 (4) ◽  
pp. 451-462 ◽  
Author(s):  
Wei Wang ◽  
Asit Rai ◽  
Eun-Mi Hur ◽  
Zeev Smilansky ◽  
Karen T. Chang ◽  
...  
Keyword(s):  
Nervous System ◽  
Protein Synthesis ◽  
Growth Cone ◽  
Actin Polymerization ◽  
Axon Growth ◽  
Axon Outgrowth ◽  
Actin Dynamics ◽  
Local Protein Synthesis ◽  
Axon Extension ◽  
Local Protein

Local information processing in the growth cone is essential for correct wiring of the nervous system. As an axon navigates through the developing nervous system, the growth cone responds to extrinsic guidance cues by coordinating axon outgrowth with growth cone steering. It has become increasingly clear that axon extension requires proper actin polymerization dynamics, whereas growth cone steering involves local protein synthesis. However, molecular components integrating these two processes have not been identified. Here, we show that Down syndrome critical region 1 protein (DSCR1) controls axon outgrowth by modulating growth cone actin dynamics through regulation of cofilin activity (phospho/dephospho-cofilin). Additionally, DSCR1 mediates brain-derived neurotrophic factor–induced local protein synthesis and growth cone turning. Our study identifies DSCR1 as a key protein that couples axon growth and pathfinding by dually regulating actin dynamics and local protein synthesis.

scholarly journals Amphiphysin 1 Is Important for Actin Polymerization during Phagocytosis

2007 ◽  
Vol 18 (11) ◽  
pp. 4669-4680 ◽  
Author(s):  
Hiroshi Yamada ◽  
Emiko Ohashi ◽  
Tadashi Abe ◽  
Norihiro Kusumi ◽  
Shun-AI Li ◽  
...  
Keyword(s):  
Rna Interference ◽  
Sertoli Cells ◽  
Small Interfering Rna ◽  
Actin Polymerization ◽  
Actin Dynamics ◽  
Polymerization Process ◽  
Amphiphysin 1 ◽  
Phagocytic Cups ◽  
Interfering Rna ◽  
Constitutively Active

Amphiphysin 1 is involved in clathrin-mediated endocytosis. In this study, we demonstrate that amphiphysin 1 is essential for cellular phagocytosis and that it is critical for actin polymerization. Phagocytosis in Sertoli cells was induced by stimulating phosphatidylserine receptors. This stimulation led to the formation of actin-rich structures, including ruffles, phagocytic cups, and phagosomes, all of which showed an accumulation of amphiphysin 1. Knocking out amphiphysin 1 by RNA interference in the cells resulted in the reduction of ruffle formation, actin polymerization, and phagocytosis. Phagocytosis was also drastically decreased in amph 1 (−/−) Sertoli cells. In addition, phosphatidylinositol-4,5-bisphosphate–induced actin polymerization was decreased in the knockout testis cytosol. The addition of recombinant amphiphysin 1 to the cytosol restored the polymerization process. Ruffle formation in small interfering RNA-treated cells was recovered by the expression of constitutively active Rac1, suggesting that amphiphysin 1 functions upstream of the protein. These findings support that amphiphysin 1 is important in the regulation of actin dynamics and that it is required for phagocytosis.

scholarly journals Tropomyosin inhibits ADF/cofilin-dependent actin filament dynamics

2002 ◽  
Vol 156 (6) ◽  
pp. 1065-1076 ◽  
Author(s):  
Shoichiro Ono ◽  
Kanako Ono
Keyword(s):  
Actin Filament ◽  
Actin Filaments ◽  
Actin Polymerization ◽  
Biological Properties ◽  
Actin Dynamics ◽  
Background Suppression ◽  
Thin Filaments ◽  
Actin Organization ◽  
C Elegans ◽  
Filament Dynamics

Tropomyosin binds to actin filaments and is implicated in stabilization of actin cytoskeleton. We examined biochemical and cell biological properties of Caenorhabditis elegans tropomyosin (CeTM) and obtained evidence that CeTM is antagonistic to ADF/cofilin-dependent actin filament dynamics. We purified CeTM, actin, and UNC-60B (a muscle-specific ADF/cofilin isoform), all of which are derived from C. elegans, and showed that CeTM and UNC-60B bound to F-actin in a mutually exclusive manner. CeTM inhibited UNC-60B–induced actin depolymerization and enhancement of actin polymerization. Within isolated native thin filaments, actin and CeTM were detected as major components, whereas UNC-60B was present at a trace amount. Purified UNC-60B was unable to interact with the native thin filaments unless CeTM and other associated proteins were removed by high-salt extraction. Purified CeTM was sufficient to restore the resistance of the salt-extracted filaments from UNC-60B. In muscle cells, CeTM and UNC-60B were localized in different patterns. Suppression of CeTM by RNA interference resulted in disorganized actin filaments and paralyzed worms in wild-type background. However, in an ADF/cofilin mutant background, suppression of CeTM did not worsen actin organization and worm motility. These results suggest that tropomyosin is a physiological inhibitor of ADF/cofilin-dependent actin dynamics.

scholarly journals F-actin-dependent Translocation of the Rap1 GDP/GTP Exchange Factor RasGRP2

2004 ◽  
Vol 279 (19) ◽  
pp. 20435-20446 ◽  
Author(s):  
Mariía J. Caloca ◽  
José L. Zugaza ◽  
Miguel Vicente-Manzanares ◽  
Francisco Sánchez-Madrid ◽  
Xosé R. Bustelo
Keyword(s):  
Signal Transduction ◽  
Amino Acids ◽  
Subcellular Distribution ◽  
Actin Polymerization ◽  
Biological Effects ◽  
Actin Dynamics ◽  
Exchange Factor ◽  
Common Structure ◽  
Direct Association ◽  
P21 Activated Kinase

RasGRPs constitute a new group of diacylglycerol-dependent GDP/GTP exchange factors that activate Ras subfamily GTPases. Despite a common structure, Ras-GRPs diverge in their GTPase specificity, subcellular distribution, and downstream biological effects. The more divergent family member is RasGRP2, a Rap1-specific exchange factor with low affinity toward diacylglycerol. The regulation of RasGRP2 during signal transduction has remained elusive up to now. In this report, we show that the subcellular localization of Ras-GRP2 is highly dependent on actin dynamics. Thus, the induction of F-actin by cytoskeletal regulators such as Vav, Vav2, Dbl, and Rac1 leads to the shift of RasGRP2 from the cytosol to membrane ruffles and its co-localization with F-actin. Treatment of cells with cytoskeletal disrupting drugs abolishes this effect, leading to an abnormal localization of RasGRP2 in cytoplasmic clusters of actin. The use of Rac1 effector mutants indicates that the RasGRP2 translocation is linked exclusively to actin polymerization and is independent of other pathways such as p21-activated kinase JNK, or superoxide production. Biochemical experiments demonstrate that the translocation of RasGRP2 to membrane ruffles is mediated by the direct association of this protein with F-actin, a property contained within its 150 first amino acids. Finally, we show that the RasGRP2/F-actin interaction promotes the regionalized activation of Rap1 in juxtamembrane areas of the cell. These results reveal a novel function of the actin cytoskeleton in mediating the spatial activation of Ras subfamily GTPases through the selective recruitment of GDP/GTP exchange factors.

scholarly journals Side-binding proteins modulate actin filament dynamics

2014 ◽  
Author(s):  
Alvaro H. Crevenna ◽  
Marcelino Arciniega ◽  
Aurelie Dupont ◽  
Kaja Kowalska ◽  
Oliver Lange ◽  
...  
Keyword(s):  
Actin Filament ◽  
Brownian Dynamics ◽  
Actin Polymerization ◽  
Actin Dynamics ◽  
Central Feature ◽  
Filament Dynamics ◽  
Filament Structure ◽  
The Rich ◽  
Dynamics Simulations ◽  
Pointed End

Actin filament dynamics govern many key physiological processes from cell motility to tissue morphogenesis. A central feature of actin dynamics is the capacity of the filament to polymerize and depolymerize at its ends in response to cellular conditions. It is currently thought that filament kinetics can be described by a single rate constant for each end. Here, using direct visualization of single actin filament elongation, we show that actin polymerization kinetics at both filament ends are strongly influenced by proteins that bind to the lateral filament surface. We also show that the less dynamic end, called the pointed-end, has a non-elongating state that dominates the observed filament kinetic asymmetry. Estimates of filament flexibility and Brownian dynamics simulations suggest that the observed kinetic diversity arises from structural alteration. Tuning filament kinetics by exploiting the natural malleability of the actin filament structure may be a ubiquitous mechanism to generate the rich variety of observed cellular actin dynamics.

scholarly journals EGF stimulates an increase in actin nucleation and filament number at the leading edge of the lamellipod in mammary adenocarcinoma cells

1998 ◽  
Vol 111 (2) ◽  
pp. 199-211 ◽  
Author(s):  
A.Y. Chan ◽  
S. Raft ◽  
M. Bailly ◽  
J.B. Wyckoff ◽  
J.E. Segall ◽  
...  
Keyword(s):  
Actin Polymerization ◽  
De Novo ◽  
Leading Edge ◽  
Mammary Adenocarcinoma ◽  
Actin Dynamics ◽  
Actin Nucleation ◽  
Nucleation Sites ◽  
Nucleation Activity ◽  
Pointed End ◽  
Stimulation Of

Stimulation of metastatic MTLn3 cells with EGF causes the rapid extension of lamellipods, which contain a zone of F-actin at the leading edge. In order to establish the mechanism for accumulation of F-actin at the leading edge and its relationship to lamellipod extension in response to EGF, we have studied the kinetics and location of EGF-induced actin nucleation activity in MTLn3 cells and characterized the actin dynamics at the leading edge by measuring the changes at the pointed and barbed ends of actin filaments upon EGF stimulation of MTLn3 cells. The major result of this study is that stimulation of MTLn3 cells with EGF causes a transient increase in actin nucleation activity resulting from the appearance of free barbed ends very close to the leading edge of extending lamellipods. In addition, cytochalasin D causes a significant decrease in the total F-actin content in EGF-stimulated cells, indicating that both actin polymerization and depolymerization are stimulated by EGF. Pointed end incorporation of rhodamine-labeled actin by the EGF stimulated cells is 2.12+/−0.47 times higher than that of control cells. Since EGF stimulation causes an increase in both barbed and pointed end incorporation of rhodamine-labeled actin in the same location, the EGF-stimulated nucleation sites are more likely due either to severing of pre-existing filaments or de novo nucleation of filaments at the leading edge thereby creating new barbed and pointed ends. The timing and location of EGF-induced actin nucleation activity in MTLn3 cells can account for the observed accumulation of F-actin at the leading edge and demonstrate that this F-actin rich zone is the primary actin polymerization zone after stimulation.

scholarly journals Specificity and Redundancy of Profilin 1 and 2 Function in Brain Development and Neuronal Structure

Cells ◽  
2021 ◽  
Vol 10 (9) ◽  
pp. 2310
Author(s):  
Marina Di Domenico ◽  
Melanie Jokwitz ◽  
Walter Witke ◽  
Pietro Pilo Boyl
Keyword(s):  
Actin Polymerization ◽  
Conditional Knockout ◽  
Brain Morphology ◽  
Actin Dynamics ◽  
Neuronal Structure ◽  
Knockout Mouse Model ◽  
Conditional Knockout Mouse ◽  
Neuronal Stem Cell ◽  
Neuronal Precursors ◽  
Redundant Functions

Profilin functions have been discussed in numerous cellular processes, including actin polymerization. One puzzling aspect is the concomitant expression of more than one profilin isoform in most tissues. In neuronal precursors and in neurons, profilin 1 and profilin 2 are co-expressed, but their specific and redundant functions in brain morphogenesis are still unclear. Using a conditional knockout mouse model to inactivate both profilins in the developing CNS, we found that threshold levels of profilin are necessary for the maintenance of the neuronal stem-cell compartment and the generation of the differentiated neurons, irrespective of the specific isoform. During embryonic development, profilin 1 is more abundant than profilin 2; consequently, modulation of profilin 1 levels resulted in a more severe phenotype than depletion of profilin 2. Interestingly, the relevance of the isoforms was reversed in the postnatal brain. Morphology of mature neurons showed a stronger dependence on profilin 2, since this is the predominant isoform in neurons. Our data highlight redundant functions of profilins in neuronal precursor expansion and differentiation, as well as in the maintenance of pyramidal neuron dendritic arborization. The specific profilin isoform is less relevant; however, a threshold profilin level is essential. We propose that the common activity of profilin 1 and profilin 2 in actin dynamics is responsible for the observed compensatory effects.

scholarly journals A new method for direct detection of the sites of actin polymerization in intact cells and its application to differentiated vascular smooth muscle

2010 ◽  
Vol 299 (5) ◽  
pp. C988-C993 ◽  
Author(s):  
Hak Rim Kim ◽  
Paul C. Leavis ◽  
Philip Graceffa ◽  
Cynthia Gallant ◽  
Kathleen G. Morgan
Keyword(s):  
Smooth Muscle ◽  
Vascular Smooth Muscle ◽  
Actin Polymerization ◽  
Direct Detection ◽  
New Method ◽  
Actin Dynamics ◽  
Tat Peptide ◽  
Isolated Cells ◽  
Intact Cells ◽  
Permeabilized Cells

Here we report and validate a new method, suitable broadly, for use in differentiated cells and tissues, for the direct visualization of actin polymerization under physiological conditions. We have designed and tested different versions of fluorescently labeled actin, reversibly attached to the protein transduction tag TAT, and have introduced this novel reagent into intact differentiated vascular smooth muscle cells (dVSMCs). A thiol-reactive version of the TAT peptide was synthesized by adding the amino acids glycine and cysteine to its NH2-terminus and forming a thionitrobenzoate adduct: viz. TAT-Cys-S-STNB. This peptide reacts readily with G-actin, and the complex is rapidly taken up by freshly enzymatically isolated dVSMC, as indicated by the fluorescence of a FITC tag on the TAT peptide. By comparing different versions of the construct, we determined that the optimal construct for biological applications is a nonfluorescently labeled TAT peptide conjugated to rhodamine-labeled actin. When TAT-Cys-S-STNB-tagged rhodamine actin (TSSAR) was added to live, freshly enzymatically isolated cells, we observed punctae of incorporated actin at the cortex of the cell. The punctae are indistinguishable from those we have previously reported to occur in the same cell type when rhodamine G-actin is added to permeabilized cells. Thus this new method allows the delivery of labeled G-actin into intact cells without disrupting the native state and will allow its further use to study the effect of physiological intracellular Ca2+ concentration transients and signal transduction on actin dynamics in intact cells.

scholarly journals Actin Cytoskeleton Regulates Calcium Dynamics and NFAT Nuclear Duration

2004 ◽  
Vol 24 (4) ◽  
pp. 1628-1639 ◽  
Author(s):  
Fabiola V. Rivas ◽  
James P. O'Keefe ◽  
Maria-Luisa Alegre ◽  
Thomas F. Gajewski
Keyword(s):  
T Cell ◽  
Actin Cytoskeleton ◽  
T Cell Activation ◽  
Actin Polymerization ◽  
Cell Activation ◽  
Cell Function ◽  
Immunological Synapse ◽  
Interleukin 2 ◽  
Actin Dynamics ◽  
Activated T Cells

ABSTRACT T-cell activation by antigen-presenting cells is accompanied by actin polymerization, T-cell receptor (TCR) capping, and formation of the immunological synapse. However, whether actin-dependent events are required for T-cell function is poorly understood. Herein, we provide evidence for an unexpected negative regulatory role of the actin cytoskeleton on TCR-induced cytokine production. Disruption of actin polymerization resulted in prolonged intracellular calcium elevation in response to anti-CD3, thapsigargin, or phorbol myristate acetate plus ionomycin, leading to persistent NFAT (nuclear factor of activated T cells) nuclear duration. These events were dominant, as the net effect of actin blockade was augmented interleukin 2 promoter activity. Increased surface expression of the plasma membrane Ca2+ ATPase was observed upon stimulation, which was inhibited by cytochalasin D, suggesting that actin polymerization contributes to calcium export. Our results imply a novel role for the actin cytoskeleton in modulating the duration of Ca2+-NFAT signaling and indicate that actin dynamics regulate features of T-cell activation downstream of receptor clustering.

scholarly journals More than a mere supply of monomers: G-Actin pools regulate actin dynamics in dendritic spines

2017 ◽  
Vol 216 (8) ◽  
pp. 2255-2257 ◽  
Author(s):  
Katalin Schlett
Keyword(s):  
Plasma Membrane ◽  
Dendritic Spines ◽  
Actin Polymerization ◽  
Synaptic Activity ◽  
Actin Dynamics ◽  
Actin Monomer ◽  
Cell Biol ◽  
Local Enrichment

Synaptic activity reshapes the morphology of dendritic spines via regulating F-actin arborization. In this issue, Lei et al. (2017. J. Cell Biol. https://doi.org/10.1083/jcb.201612042) reports a novel, G-actin–dependent regulation of actin polymerization within spine heads. They show that actin monomer levels are elevated in spines upon activity, with G-actin immobilized by the local enrichment of phosphatidylinositol (3,4,5)-triphosphate (PIP3) within the spine plasma membrane.

Sign in / Sign up

Export Citation Format

Share Document