scholarly journals Coactosin Promotes F-Actin Protrusion in Growth Cones Under Cofilin-Related Signaling Pathway

2021 ◽  
Vol 9 ◽  
Author(s):  
Xubin Hou ◽  
Motohiro Nozumi ◽  
Harukazu Nakamura ◽  
Michihiro Igarashi ◽  
Sayaka Sugiyama
Keyword(s):  
Growth Cone ◽  
Actin Polymerization ◽  
Leading Edge ◽  
Axonal Growth ◽  
Growth Cones ◽  
Optical Diffraction ◽  
Dependent Manner ◽  
Structured Illumination Microscopy ◽  
Axon Extension ◽  
Specific Distribution

During brain development, axon outgrowth and its subsequent pathfinding are reliant on a highly motile growth cone located at the tip of the axon. Actin polymerization that is regulated by actin-depolymerizing factors homology (ADF-H) domain-containing family drives the formation of lamellipodia and filopodia at the leading edge of growth cones for axon guidance. However, the precise localization and function of ADF-H domain-containing proteins involved in axon extension and retraction remain unclear. We have previously shown that transcripts and proteins of coactosin-like protein 1 (COTL1), an ADF-H domain-containing protein, are observed in neurites and axons in chick embryos. Coactosin overexpression analysis revealed that this protein was localized to axonal growth cones and involved in axon extension in the midbrain. We further examined the specific distribution of coactosin and cofilin within the growth cone using superresolution microscopy, structured illumination microscopy, which overcomes the optical diffraction limitation and is suitable to the analysis of cellular dynamic movements. We found that coactosin was tightly associated with F-actin bundles at the growth cones and that coactosin overexpression promoted the expansion of lamellipodia and extension of growth cones. Coactosin knockdown in oculomotor neurons resulted in an increase in the levels of the inactive, phosphorylated form of cofilin and dysregulation of actin polymerization and axonal elongation, which suggests that coactosin promoted axonal growth in a cofilin-dependent manner. Indeed, the application of a dominant-negative form of LIMK1, a downstream effector of GTPases, reversed the effect of coactosin knockdown on axonal growth by enhancing cofilin activity. Combined, our results indicate that coactosin functions promote the assembly of protrusive actin filament arrays at the leading edge for growth cone motility.

scholarly journals Xenopus cytoplasmic linker–associated protein 1 (XCLASP1) promotes axon elongation and advance of pioneer microtubules

2013 ◽  
Vol 24 (10) ◽  
pp. 1544-1558 ◽  
Author(s):  
Astrid Marx ◽  
William J. Godinez ◽  
Vasil Tsimashchuk ◽  
Peter Bankhead ◽  
Karl Rohr ◽  
...  
Keyword(s):  
Growth Cone ◽  
Binding Protein ◽  
Leading Edge ◽  
Growth Cones ◽  
Retrograde Flow ◽  
Spinal Cord Neurons ◽  
Actin Organization ◽  
Axon Elongation ◽  
Axon Extension ◽  
Potential Link

Dynamic microtubules (MTs) are required for neuronal guidance, in which axons extend directionally toward their target tissues. We found that depletion of the MT-binding protein Xenopus cytoplasmic linker–associated protein 1 (XCLASP1) or treatment with the MT drug Taxol reduced axon outgrowth in spinal cord neurons. To quantify the dynamic distribution of MTs in axons, we developed an automated algorithm to detect and track MT plus ends that have been fluorescently labeled by end-binding protein 3 (EB3). XCLASP1 depletion reduced MT advance rates in neuronal growth cones, very much like treatment with Taxol, demonstrating a potential link between MT dynamics in the growth cone and axon extension. Automatic tracking of EB3 comets in different compartments revealed that MTs increasingly slowed as they passed from the axon shaft into the growth cone and filopodia. We used speckle microscopy to demonstrate that MTs experience retrograde flow at the leading edge. Microtubule advance in growth cone and filopodia was strongly reduced in XCLASP1-depleted axons as compared with control axons, but actin retrograde flow remained unchanged. Instead, we found that XCLASP1-depleted growth cones lacked lamellipodial actin organization characteristic of protrusion. Lamellipodial architecture depended on XCLASP1 and its capacity to associate with MTs, highlighting the importance of XCLASP1 in actin–microtubule interactions.

scholarly journals Shootin1–cortactin interaction mediates signal–force transduction for axon outgrowth

2015 ◽  
Vol 210 (4) ◽  
pp. 663-676 ◽  
Author(s):  
Yusuke Kubo ◽  
Kentarou Baba ◽  
Michinori Toriyama ◽  
Takunori Minegishi ◽  
Tadao Sugiura ◽  
...  
Keyword(s):  
Actin Polymerization ◽  
Leading Edge ◽  
Growth Cones ◽  
Axon Outgrowth ◽  
Actin Binding ◽  
Retrograde Flow ◽  
Environmental Chemical ◽  
Mechanical Coupling ◽  
Cell Adhesions ◽  
Force Transduction

Motile cells transduce environmental chemical signals into mechanical forces to achieve properly controlled migration. This signal–force transduction is thought to require regulated mechanical coupling between actin filaments (F-actins), which undergo retrograde flow at the cellular leading edge, and cell adhesions via linker “clutch” molecules. However, the molecular machinery mediating this regulatory coupling remains unclear. Here we show that the F-actin binding molecule cortactin directly interacts with a clutch molecule, shootin1, in axonal growth cones, thereby mediating the linkage between F-actin retrograde flow and cell adhesions through L1-CAM. Shootin1–cortactin interaction was enhanced by shootin1 phosphorylation by Pak1, which is activated by the axonal chemoattractant netrin-1. We provide evidence that shootin1–cortactin interaction participates in netrin-1–induced F-actin adhesion coupling and in the promotion of traction forces for axon outgrowth. Under cell signaling, this regulatory F-actin adhesion coupling in growth cones cooperates with actin polymerization for efficient cellular motility.

scholarly journals Nerve growth cone lamellipodia contain two populations of actin filaments that differ in organization and polarity.

1992 ◽  
Vol 119 (5) ◽  
pp. 1219-1243 ◽  
Author(s):  
A K Lewis ◽  
P C Bridgman
Keyword(s):  
Actin Filament ◽  
Growth Cone ◽  
Actin Filaments ◽  
Membrane Surface ◽  
Leading Edge ◽  
Growth Cones ◽  
Differential Interference Contrast Microscopy ◽  
Nerve Growth ◽  
Filament Bundles ◽  
Two Populations

The organization and polarity of actin filaments in neuronal growth cones was studied with negative stain and freeze-etch EM using a permeabilization protocol that caused little detectable change in morphology when cultured nerve growth cones were observed by video-enhanced differential interference contrast microscopy. The lamellipodial actin cytoskeleton was composed of two distinct subpopulations: a population of 40-100-nm-wide filament bundles radiated from the leading edge, and a second population of branching short filaments filled the volume between the dorsal and ventral membrane surfaces. Together, the two populations formed the three-dimensional structural network seen within expanding lamellipodia. Interaction of the actin filaments with the ventral membrane surface occurred along the length of the filaments via membrane associated proteins. The long bundled filament population was primarily involved in these interactions. The filament tips of either population appeared to interact with the membrane only at the leading edge; this interaction was mediated by a globular Triton-insoluble material. Actin filament polarity was determined by decoration with myosin S1 or heavy meromyosin. Previous reports have suggested that the polarity of the actin filaments in motile cells is uniform, with the barbed ends toward the leading edge. We observed that the actin filament polarity within growth cone lamellipodia is not uniform; although the predominant orientation was with the barbed end toward the leading edge (47-56%), 22-25% of the filaments had the opposite orientation with their pointed ends toward the leading edge, and 19-31% ran parallel to the leading edge. The two actin filament populations display distinct polarity profiles: the longer filaments appear to be oriented predominantly with their barbed ends toward the leading edge, whereas the short filaments appear to be randomly oriented. The different length, organization and polarity of the two filament populations suggest that they differ in stability and function. The population of bundled long filaments, which appeared to be more ventrally located and in contact with membrane proteins, may be more stable than the population of short branched filaments. The location, organization, and polarity of the long bundled filaments suggest that they may be necessary for the expansion of lamellipodia and for the production of tension mediated by receptors to substrate adhesion molecules.

NrCAM, cerebellar granule cell receptor for the neuronal adhesion molecule F3, displays an actin-dependent mobility in growth cones

1999 ◽  
Vol 112 (18) ◽  
pp. 3015-3027 ◽  
Author(s):  
C. Faivre-Sarrailh ◽  
J. Falk ◽  
E. Pollerberg ◽  
M. Schachner ◽  
G. Rougon
Keyword(s):  
Growth Cone ◽  
Actin Filaments ◽  
Cerebellar Granule Cells ◽  
Leading Edge ◽  
Growth Cones ◽  
Inhibitory Effect ◽  
Immunoglobulin Superfamily ◽  
Cerebellar Granule ◽  
Axonal Elongation ◽  
Neuronal Adhesion

The neuronal adhesion glycoprotein F3 is a multifunctional molecule of the immunoglobulin superfamily that displays heterophilic binding activities. In the present study, NrCAM was identified as the functional receptor mediating the inhibitory effect of F3 on axonal elongation from cerebellar granule cells. F3Fc-conjugated microspheres binding to neuronal growth cones resulted from heterophilic interaction with NrCAM but not with L1. Time-lapse video-microscopy indicated that F3Fc beads bind at the leading edge and move retrogradely to reach the base of the growth cone within a lapse of 30–60 seconds. Such velocity (5.7 microm/minute) is consistent with a coupling between F3 receptors and the retrograde flow of actin filaments. When actin filaments were disrupted by cytochalasin B, the F3Fc beads remained immobile at the leading edge. The retrograde mobility appeared to be dependent on NrCAM clustering since it was induced upon binding with cross-linked but not dimeric F3Fc chimera. These data indicate that F3 may control growth cone motility by modulating the linkage of its receptor, NrCAM, to the cytoskeleton. They provide further insights into the mechanisms by which GPI-anchored adhesion molecules may exert an inhibitory effect on axonal elongation.

scholarly journals Unique Responses of Differentiating Neuronal Growth Cones to Inhibitory Cues Presented by Oligodendrocytes

1998 ◽  
Vol 142 (1) ◽  
pp. 191-202 ◽  
Author(s):  
A. Shibata ◽  
M.V. Wright ◽  
S. David ◽  
L. McKerracher ◽  
P.E. Braun ◽  
...  
Keyword(s):  
Growth Cone ◽  
Hippocampal Neurons ◽  
Dendritic Growth ◽  
System Development ◽  
Axonal Growth ◽  
Growth Cones ◽  
Nervous System Development ◽  
Environmental Cues ◽  
Growth Cone Collapse ◽  
Neuronal Growth Cones

During central nervous system development, neurons differentiate distinct axonal and dendritic processes whose outgrowth is influenced by environmental cues. Given the known intrinsic differences between axons and dendrites and that little is known about the response of dendrites to inhibitory cues, we tested the hypothesis that outgrowth of differentiating axons and dendrites of hippocampal neurons is differentially influenced by inhibitory environmental cues. A sensitive growth cone behavior assay was used to assess responses of differentiating axonal and dendritic growth cones to oligodendrocytes and oligodendrocyte- derived, myelin-associated glycoprotein (MAG). We report that >90% of axonal growth cones collapsed after contact with oligodendrocytes. None of the encounters between differentiating, MAP-2 positive dendritic growth cones and oligodendrocytes resulted in growth cone collapse. The insensitivity of differentiating dendritic growth cones appears to be acquired since they develop from minor processes whose growth cones are inhibited (nearly 70% collapse) by contact with oligodendrocytes. Recombinant MAG(rMAG)-coated beads caused collapse of 72% of axonal growth cones but only 29% of differentiating dendritic growth cones. Unlike their response to contact with oligodendrocytes, few growth cones of minor processes were inhibited by rMAG-coated beads (20% collapsed). These results reveal the capability of differentiating growth cones of the same neuron to partition the complex molecular terrain they navigate by generating unique responses to particular inhibitory environmental cues.

scholarly journals Actions of cytochalasins on the organization of actin filaments and microtubules in a neuronal growth cone.

1988 ◽  
Vol 107 (4) ◽  
pp. 1505-1516 ◽  
Author(s):  
P Forscher ◽  
S J Smith
Keyword(s):  
Growth Cone ◽  
Spatial Organization ◽  
Leading Edge ◽  
Growth Cones ◽  
Actin Networks ◽  
Antibody Staining ◽  
Flow Perfusion ◽  
Rapid Flow ◽  
Lamellar Region ◽  
Neuronal Growth Cone

Actions of cytochalasin B (CB) on cytoskeletons and motility of growth cones from cultured Aplysia neurons were studied using a rapid flow perfusion chamber and digital video light microscopy. Living growth cones were observed using differential interference contrast optics and were also fixed at various time points to assay actin filament (F-actin) and microtubule distributions. Treatment with CB reversibly blocked motility and eliminated most of the phalloidin-stainable F-actin from the leading lamella. The loss of F-actin was nearly complete within 2-3 min of CB application and was largely reversed within 5-6 min of CB removal. The loss and recovery of F-actin were found to occur with a very distinctive spatial organization. Within 20-30 s of CB application, F-actin networks receded from the entire peripheral margin of the lamella forming a band devoid of F-actin. This band widened as F-actin receded at rates of 3-6 microns/min. Upon removal of CB, F-actin began to reappear within 20-30 s. The initial reappearance of F-actin took two forms: a coarse isotropic matrix of F-actin bundles throughout the lamella, and a denser matrix along the peripheral margin. The denser peripheral matrix then expanded in width, extending centrally to replace the coarse matrix at rates again between 3-6 microns/min. These results suggest that actin normally polymerizes at the leading edge and then flows rearward at a rate between 3-6 microns/min. CB treatment was also observed to alter the distribution of microtubules, assayed by antitubulin antibody staining. Normally, microtubules are restricted to the neurite shaft and a central growth cone domain. Within approximately 5 min after CB application, however, microtubules began extending into the lamellar region, often reaching the peripheral margin. Upon removal of CB, the microtubules were restored to their former central localization. The timing of these microtubule redistributions is consistent with their being secondary to effects of CB on lamellar F-actin.

scholarly journals Curvature sensing steers actin-driven cell migration

2021 ◽  
Author(s):  
Ewa Sitarska ◽  
Silvia Dias Almeida ◽  
Marianne Beckwith ◽  
Julian Stopp ◽  
Yannick Schwab ◽  
...  
Keyword(s):  
Cell Migration ◽  
Actin Polymerization ◽  
Leading Edge ◽  
Membrane Curvature ◽  
Cell Periphery ◽  
Dependent Manner ◽  
Complex Environments ◽  
Time Resolved ◽  
Curvature Sensing ◽  
Membrane Topography

Cell migration is fundamental for the immune response, development, and morphogenesis. For navigation through complex and ever-changing environments, migrating cells require a balance between a stable leading-edge, which is necessary for directional migration, and some unstable features to enable the required dynamic behaviors. The leading edge is often composed of actin-driven protrusions including lamellipodia and ruffles with continuously changing membrane curvature. Whether their membrane topography affects the cell's leading edge and motion persistence in complex environments remains unknown. To study this, we combined a theoretical analysis with machine learning-based segmentation for time-resolved TIRF microscopy, membrane topography analysis from electron microscopy images and microfluidics. We discovered that cell motion persistence and directionality, in both freely moving and environmentally-constrained cells, strongly depend on the curvature-sensing protein Snx33. Specifically, Snx33 promotes leading edge instabilities by locally inhibiting WAVE2- driven actin polymerization in a curvature-dependent manner. Snx33 knockout cells migrate faster and are more persistent during unobstructed migration, but fail when a change in direction is required. Thus, Snx33 is key for steering cell motility in complex environments by facilitating contact inhibition of locomotion and promoting efficient turning. These results identify cell surface topography as an organizing principle at the cell periphery that directs cell migration.

scholarly journals Nanoscopic Clustering of Neuroligin-3 and Neuroligin-4X Regulates Growth Cone Organization and Size

2019 ◽  
Author(s):  
Nicholas J. F. Gatford ◽  
P. J. Michael Deans ◽  
Rodrigo R.R. Duarte ◽  
George Chennell ◽  
Pooja Raval ◽  
...  
Keyword(s):  
Growth Cone ◽  
Synapse Formation ◽  
Super Resolution ◽  
Leading Edge ◽  
Growth Cones ◽  
Autism Spectrum ◽  
Adhesion Proteins ◽  
Human Neurons ◽  
Spectrum Condition ◽  
P21 Activated Kinase

AbstractThe cell-adhesion proteins neuroligin-3 and neuroligin-4X (NLGN3/4X) have well described roles in synapse formation. NLGN3/4X are also expressed highly during neurodevelopment. However, the role these proteins play during this period is unknown. Here we show that NLGN3/4X localized to the leading edge of growth cones where itpromoted neuritogenesis in immature human neurons. Super-resolution microscopyrevealed that NLGN3/4X clustering induced growth cone enlargement and influenced actin filament organization. Critically, these morphological effects were not induced by Autism spectrum condition (ASC)-associated NLGN3/4X variants. Finally, actin regulators p21-activated kinase 1 (PAK1) and cofilin were found to be activated by NLGN3/4X and involved in mediating the effects of these adhesion proteins on actin filaments, growth cones, and neuritogenesis. These data reveal a novel role for NLGN3 and NLGN4X in the development of neuronal architecture, which may be altered in the presence of ASD-associated variants.

scholarly journals Radixin Is Involved in Lamellipodial Stability during Nerve Growth Cone Motility

1999 ◽  
Vol 10 (5) ◽  
pp. 1511-1520 ◽  
Author(s):  
Leslie Castelo ◽  
Daniel G. Jay
Keyword(s):  
Growth Cone ◽  
Critical Role ◽  
Leading Edge ◽  
Growth Cones ◽  
Forward Movement ◽  
Nerve Growth ◽  
Nerve Growth Cone

Immunocytochemistry and in vitro studies have suggested that the ERM (ezrin-radixin-moesin) protein, radixin, may have a role in nerve growth cone motility. We tested the in situ role of radixin in chick dorsal root ganglion growth cones by observing the effects of its localized and acute inactivation. Microscale chromophore-assisted laser inactivation (micro-CALI) of radixin in growth cones causes a 30% reduction of lamellipodial area within the irradiated region whereas all control treatments did not affect lamellipodia. Micro-CALI of radixin targeted to the middle of the leading edge often split growth cones to form two smaller growth cones during continued forward movement (>80%). These findings suggest a critical role for radixin in growth cone lamellipodia that is similar to ezrin function in pseudopodia of transformed fibroblasts. They are consistent with radixin linking actin filaments to each other or to the membrane during motility.

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