Membrane Tension, Myosin Force, and Actin Turnover Maintain Actin Treadmill in the Nerve Growth Cone

Craig, Erin M.; Goor, David Van; Forscher, Paul; Mogilner, Alex

doi:10.1016/j.bpj.2012.03.003

Cited by 69 publications

(71 citation statements)

References 56 publications

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“…The actin-capping drug cytochalasin D, the monomeric actin-sequestering drug latrunculin A, and the actinstabilizing agent jasplakinolide all dramatically diminished the fraction of rearward-moving actin-mEos2 molecules (21%, 7%, and 2%, respectively) ( Fig. S5), consistent with a major role of actin tread-milling in generating the flow (2,3,5). Myosin-light chain (MLC)-GFP was localized in the intermediary region of the growth cone, and MLC-Eos2 trajectories were mostly confined in this area (Fig.…”

Section: Significancesupporting

confidence: 56%

“…Growth cones translate extracellular signals into directional migration through a coordinated regulation of cytoskeleton, adhesion, and membrane processes (1). At the cytoskeletal level, motility is generated by polarized actin treadmilling, which, together with myosin contraction, generates a continuous retrograde actin flow from the periphery to the base of growth cones (2)(3)(4)(5)(6)(7). At the membrane level, adhesion proteins form dynamic bonds with immobilized extracellular ligands, allowing step-by-step locomotion (8).…”

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confidence: 99%

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Two-tiered coupling between flowing actin and immobilized N -cadherin/catenin complexes in neuronal growth cones

Garcia

Leduc

Lagardère

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

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Neuronal growth cones move forward by dynamically connecting actin-based motility to substrate adhesion, but the mechanisms at the individual molecular level remain unclear. We cultured primary neurons on N-cadherin-coated micropatterned substrates, and imaged adhesion and cytoskeletal proteins at the ventral surface of growth cones using single particle tracking combined to photoactivated localization microscopy (sptPALM). We demonstrate transient interactions in the second time scale between flowing actin filaments and immobilized N-cadherin/catenin complexes, translating into a local reduction of the actin retrograde flow. Normal actin flow on micropatterns was rescued by expression of a dominant negative N-cadherin construct competing for the coupling between actin and endogenous N-cadherin. Fluorescence recovery after photobleaching (FRAP) experiments confirmed the differential kinetics of actin and N-cadherin, and further revealed a 20% actin population confined at N-cadherin micropatterns, contributing to local actin accumulation. Computer simulations with relevant kinetic parameters modeled N-cadherin and actin turnover well, validating this mechanism. Such a combination of short-and long-lived interactions between the motile actin network and spatially restricted adhesive complexes represents a two-tiered clutch mechanism likely to sustain dynamic environment sensing and provide the force necessary for growth cone migration.growth cone | actin flow | N-cadherin adhesion | micropatterned substrates | single-molecule tracking G rowth cones are motile structures at the extremity of axons responsible for path finding and neurite extension during nervous system development and repair. Growth cones translate extracellular signals into directional migration through a coordinated regulation of cytoskeleton, adhesion, and membrane processes (1). At the cytoskeletal level, motility is generated by polarized actin treadmilling, which, together with myosin contraction, generates a continuous retrograde actin flow from the periphery to the base of growth cones (2-7). At the membrane level, adhesion proteins form dynamic bonds with immobilized extracellular ligands, allowing step-by-step locomotion (8).The molecular clutch model postulates that the mechanical coupling between ligand-bound transmembrane adhesion receptors and the actin flow allows traction forces to be transmitted to the substrate, resulting in local diminution of the retrograde flow and forward progression (9-11). Optical tweezers and flexible substrate experiments using microspheres coated with adhesion molecules revealed clutch-like mechanisms for integrins (12, 13), Ig cell adhesion molecules (14, 15), and cadherins (16,17). However, the mechanism of clutch engagement at the individual molecular level remains elusive. For integrin-based adhesion, single-molecule tracking experiments suggested that talin and vinculin could switch between a state bound to flowing actin and a state bound to immobilized integrins (18, 19). For cadherin-based adhesion,...

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Section: Significancesupporting

confidence: 56%

mentioning

confidence: 99%

Two-tiered coupling between flowing actin and immobilized N -cadherin/catenin complexes in neuronal growth cones

Garcia

Leduc

Lagardère

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

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“…The amplitude of forces exerted on N-cadherin-coated substrates is lower than that on FN (Ladoux et al, 2010). Furthermore, the actin treadmilling speed is higher on N-cadherin than on FN, which could be interpreted as a lower degree of coupling of Ncadherin to actin filaments (Craig et al, 2012;Giannone et al, 2009). As a result, cadherin adhesions would not initiate or tolerate the increased internal cell tension required for actin reorganization.…”

Section: Discussionmentioning

confidence: 97%

“…It might result from the continuous flow and severing of the branched network of actin filaments that elongate at the lamellipodial leading edge (Craig et al, 2012). On FN substrates, following the reinforcement of focal adhesions that are subjected to sustained increased contractility, cells shift their actin organization from an isotropic actomyosin belt to stress fibers, initializing cell polarization (Prager-Khoutorsky et al, 2011;Zamir et al, 2000).…”

Section: Discussionmentioning

confidence: 99%

Adhesive interactions of N-cadherin limit the recruitment of microtubules to cell–cell contacts through organization of actomyosin

Plestant

Strale

Seddiki

et al. 2014

Journal of Cell Science

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Adhesive interactions of cadherins induce crosstalk between adhesion complexes and the actin cytoskeleton, allowing strengthening of adhesions and cytoskeletal organization. The underlying mechanisms are not completely understood, and microtubules (MTs) might be involved, as for integrin-mediated cell-extracellular-matrix adhesions. Therefore, we investigated the relationship between N-cadherin and MTs by analyzing the influence of N-cadherin engagement on MT distribution and dynamics. MTs progressed less, with a lower elongation rate, towards cadherin adhesions than towards focal adhesions. Increased actin treadmilling and the presence of an actomyosin contractile belt, suggested that actin relays inhibitory signals from cadherin adhesions to MTs. The reduced rate of MT elongation, associated with reduced recruitment of end-binding (EB) proteins to plus ends, was alleviated by expression of truncated N-cadherin, but was only moderately affected when actomyosin was disrupted. By contrast, destabilizing actomyosin fibers allowed MTs to enter the adhesion area, suggesting that tangential actin bundles impede MT growth independently of MT dynamics. Blocking MT penetration into the adhesion area strengthened cadherin adhesions. Taken together, these results establish a crosstalk between N-cadherin, Factin and MTs. The opposing effects of cadherin and integrin engagement on actin organization and MT distribution might induce bias of the MT network during cell polarization.

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“…In the lamellipodium, from which filopodia often emanate, the retrograde flow originates from actin treadmilling due to actin depolymerization at the rear and polymerization at the front of the lamellipodium. This retrograde flow is further amplified by the motor activity of myosins (20)(21)(22)(23). In neurons, the filopodial shaft is deeply anchored in the growth cone and filopodial dynamics depends on the balance between actin polymerization at the filopodial tip and its retrograde flow (19).…”

mentioning

confidence: 99%

Filopodial retraction force is generated by cortical actin dynamics and controlled by reversible tethering at the tip

Bornschlögl

Roméro

Vestergaard

et al. 2013

Proc. Natl. Acad. Sci. U.S.A.

136

109

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Filopodia are dynamic, finger-like plasma membrane protrusions that sense the mechanical and chemical surroundings of the cell. Here, we show in epithelial cells that the dynamics of filopodial extension and retraction are determined by the difference between the actin polymerization rate at the tip and the retrograde flow at the base of the filopodium. Adhesion of a bead to the filopodial tip locally reduces actin polymerization and leads to retraction via retrograde flow, reminiscent of a process used by pathogens to invade cells. Using optical tweezers, we show that filopodial retraction occurs at a constant speed against counteracting forces up to 50 pN. Our measurements point toward retrograde flow in the cortex together with frictional coupling between the filopodial and cortical actin networks as the main retraction-force generator for filopodia. The force exerted by filopodial retraction, however, is limited by the connection between filopodial actin filaments and the membrane at the tip. Upon mechanical rupture of the tip connection, filopodia exert a passive retraction force of 15 pN via their plasma membrane. Transient reconnection at the tip allows filopodia to continuously probe their surroundings in a load-and-fail manner within a welldefined force range.cytoskeleton | mechanics | membrane-tension | pulling | lamellipodium

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Membrane Tension, Myosin Force, and Actin Turnover Maintain Actin Treadmill in the Nerve Growth Cone

Cited by 69 publications

References 56 publications

Two-tiered coupling between flowing actin and immobilized N -cadherin/catenin complexes in neuronal growth cones

Two-tiered coupling between flowing actin and immobilized N -cadherin/catenin complexes in neuronal growth cones

Adhesive interactions of N-cadherin limit the recruitment of microtubules to cell–cell contacts through organization of actomyosin

Filopodial retraction force is generated by cortical actin dynamics and controlled by reversible tethering at the tip

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