Direct measurement of force generation by actin filament polymerization using an optical trap

Footer, Matthew J.; Kerssemakers, Jacob; Theriot, Julie A.; Dogterom, Marileen

doi:10.1073/pnas.0607052104

Cited by 335 publications

(381 citation statements)

References 44 publications

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“…We make the further assumption (described previously) that actin filament protrusion is mechanically stalled by the membrane tension T at the sides of the front of the lamellipodium (l~+x=2). The force acting on each filament at the sides must therefore be approximately equal to the force required to stall a single actin filament 28 , f stall , which has been measured 29,30 , so…”

Section: Nature | Vol 453 | 22 May 2008mentioning

confidence: 99%

Mechanism of shape determination in motile cells

Keren

Pincus

Allen³

et al. 2008

Nature

Self Cite

678

816

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The shape of motile cells is determined by many dynamic processes spanning several orders of magnitude in space and time, from local polymerization of actin monomers at subsecond timescales to global, cell-scale geometry that may persist for hours. Understanding the mechanism of shape determination in cells has proved to be extremely challenging due to the numerous components involved and the complexity of their interactions. Here we harness the natural phenotypic variability in a large population of motile epithelial keratocytes from fish (Hypsophrys nicaraguensis) to reveal mechanisms of shape determination. We find that the cells inhabit a low-dimensional, highly correlated spectrum of possible functional states. We further show that a model of actin network treadmilling in an inextensible membrane bag can quantitatively recapitulate this spectrum and predict both cell shape and speed. Our model provides a simple biochemical and biophysical basis for the observed morphology and behaviour of motile cells.Cell shape emerges from the interaction of many constituent elements-notably, the cytoskeleton, the cell membrane and cellsubstrate adhesions-that have been studied in great detail at the molecular level 1-3 ; however, the mechanism by which global morphology is generated and maintained at the cellular scale is not understood. Many studies have characterized the morphological effects of perturbing various cytoskeletal and other cellular components (for example, ref. 4); yet, there have been no comprehensive efforts to try to understand cell shape from first principles. Here we address this issue in the context of motile epithelial keratocytes derived from fish skin. Fish keratocytes are among the fastest moving animal cells, and their motility machinery is characterized by extremely rapid molecular dynamics and turnover [5][6][7][8] . At the same time, keratocytes are able to maintain nearly constant speed and direction during movement over many cell lengths. Their shapes, consisting of a bulbous cell body at the rear attached to a broad, thin lamellipodium at the front and sides, are simple, stereotyped and notoriously temporally persistent 9,10 . The molecular dynamism of these cells, combined with the persistence of their global shape and behaviour, make them an ideal model system for investigating the mechanisms of cell shape determination.The relative simplicity of keratocytes has inspired extensive experimental and theoretical investigations into this cell type 5-17 , considerably advancing the understanding of cell motility. A notable example is the graded radial extension (GRE) model 12 , which was an early attempt to link the mechanism of motility at the molecular level with overall cell geometry. The GRE model proposed that local cell extension (either protrusion or retraction) occurs perpendicular to the cell edge, and that the magnitude of this extension is graded from a maximum near the cell midline to a minimum towards the sides. Although this phenomenological model has been shown experimentally ...

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Section: Nature | Vol 453 | 22 May 2008mentioning

confidence: 99%

Mechanism of shape determination in motile cells

Keren

Pincus

Allen³

et al. 2008

Nature

Self Cite

678

816

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“…To the best of our knowledge, only one set of experiments has reported stall force for a single growing actin filament -a value of ≈ 1pN [41]. In another set of experiments on a few actin filaments growing in parallel, Footer et al [49] reported a collective stall force of ≈ 1 pN, which is very similar to the load required to stall a single filament. In this experiment, the interpretation of filament cooperativity while pushing together against the barrier is, however, complicated by the fact that individual actin filaments may buckle before reaching their stall force, and hence, collectively the bundle may be unable to reach its full potential for force generation.…”

Section: Introductionmentioning

confidence: 62%

“…A number of experiments on force generation by an assembly of biofilaments have been reported in the literature [36,39,[41][42][43][44][45][46][47][48][49]. In view of the overall content and objective of our paper we will focus only on the experiments related to single and parallel bundles of growing biofilaments.…”

Section: Introductionmentioning

confidence: 99%

Sufficient conditions for the additivity of stall forces generated by multiple filaments or motors

Bameta

Das

et al. 2017

Phys. Rev. E

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Molecular motors and cytoskeletal filaments work collectively most of the time under opposing forces. This opposing force may be due to cargo carried by motors or resistance coming from the cell membrane pressing against the cytoskeletal filaments. Some recent studies have shown that the collective maximum force (stall force) generated by multiple cytoskeletal filaments or molecular motors may not always be just a simple sum of the stall forces of the individual filaments or motors. To understand this excess or deficit in the collective force, we study a broad class of models of both cytoskeletal filaments and molecular motors. We argue that the stall force generated by a group of filaments or motors is additive, that is, the stall force of N number of filaments (motors) is N times the stall force of one filament (motor), when the system is in equilibrium at stall. Conversely, we show that this additive property typically does not hold true when the system is not at equilibrium at stall. We thus present a novel and unified understanding of the existing models exhibiting such nonaddivity, and generalise our arguments by developing new models that demonstrate this phenomena. We also propose a quantity similar to thermodynamic efficiency to easily predict this deviation from stall-force additivity for filament and motor collectives.

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“…The bending shape of single F-actin filaments observed by fluorescence spectroscopy, was precisely exploited to measure for the first time the typical polymerization force generated by single living actin filaments [15,16]. In the optical trap experiment of Footer et al [5], a bundle of about ten filaments, with their seeds glued to a trapped colloidal particle, push with their active side (barbed end) on a fixed rigid wall. The polymerization force they progressively develop to reach equilibrium is inferred by measuring the colloidal displacement in the trap.…”

Section: Introductionmentioning

confidence: 99%

“…In vitro experiments on biofilaments, like actin and tubulin, to measure in supercritical conditions either the forcevelocity relationship in detectable non-zero velocity conditions [3,4], or the approach to stalling where the applied load effectively stops the net polymerization of living filaments and the stalling force is effectively measured [5], have been deviced. To simplify the analysis and to concentrate on the fundamental process of force generation by polymerizing filaments, the experiments deal with bundles of parallel filaments hitting an orthogonal moving wall, a network having strong analogy with the structure of actin filopedia [1].…”

Section: Introductionmentioning

confidence: 99%

A semi-flexible model prediction for the polymerization force exerted by a living F-actin filament on a fixed wall

Pierleoni

Ciccotti

Ryckaert

2015

The Journal of Chemical Physics

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We consider a single living semi-flexible filament with persistence length p in chemical equilibrium with a solution of free monomers at fixed monomer chemical potential µ1 and fixed temperature T . While one end of the filament is chemically active with single monomer (de)polymerization steps, the other end is grafted normally to a rigid wall to mimick a rigid network from which the filament under consideration emerges. A second rigid wall, parallel to the grafting wall, is fixed at distance L << p from the filament seed. In supercritical conditions where monomer density ρ1 is higher than the critical density ρ1c, the filament tends to polymerize and impinges onto the second surface which, in suitable conditions (non-escaping filament regime) stops the filament growth. We first establish the grand-potential Ω(µ1, T, L) of this system treated as an ideal reactive mixture and derive some general properties, in particular the filament size distribution and the force exerted by the living filament on the obstacle wall. We apply this formalism to the semi-flexible, living, over a compression range much narrower than the size d of a monomer. Employing this universal form for living filaments, we find that the average force exerted by a living filament on a wall at distance L is in practice L independent and very close to the value of the stalling force F H s = (kBT /d) ln(ρ1) predicted by Hill, this expression being strictly valid in the rigid filament limit. The average filament force results from the product of the cumulative size fraction

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Direct measurement of force generation by actin filament polymerization using an optical trap

Cited by 335 publications

References 44 publications

Mechanism of shape determination in motile cells

Mechanism of shape determination in motile cells

Sufficient conditions for the additivity of stall forces generated by multiple filaments or motors

A semi-flexible model prediction for the polymerization force exerted by a living F-actin filament on a fixed wall

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