Myosin II contributes to cell-scale actin network treadmilling through network disassembly

Wilson, Cyrus A.; Tsuchida, Mark A.; Allen, Greg M.; Barnhart, Erin L.; Applegate, Kathryn T.; Yam, Patricia T.; Ji, Li; Keren, Kinneret; Danuser, Gaudenz; Theriot, Julie A.

doi:10.1038/nature08994

Cited by 355 publications

(400 citation statements)

References 29 publications

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“…2 D-E, Movie S1). Similar flow patterns were observed in whole keratocytes, with the important difference that in whole-cells a myosin II-dependent inward actin flow was prominent near the rear boundary (7,16). This difference is likely due to the higher quantity of myosin II in cells (9) compared to fragments (13), which are typically generated from lamellipodia that are relatively poor in myosin (9).…”

Section: Resultssupporting

confidence: 53%

“…Myosin contraction can contribute to rear retraction as well as promote motility by enhancing actin disassembly (7) and inducing forward directed fluid flow (27). However, it is not required; knockouts and inhibition experiments have shown that although myosin II typically promotes movement and increases cell speed, it is not essential for motility in many cell types including keratocytes (5,7,27), Acanthamoeba (28), and Dictyostelium discoideum (29). Here we show that myosin II has a negligible role in rear retraction in keratocyte fragments (Fig.…”

Section: Discussionmentioning

confidence: 99%

“…Actin dynamics are measured as in ref. 7, and actin distribution is measured by fixation and phalloidin staining as in ref. 5.…”

Section: Methodsmentioning

confidence: 99%

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Actin disassembly clock determines shape and speed of lamellipodial fragments

Ofer

Mogilner

Keren

2011

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

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113

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A central challenge in motility research is to quantitatively understand how numerous molecular building blocks self-organize to achieve coherent shape and movement on cellular scales. A classic example of such self-organization is lamellipodial motility in which forward translocation is driven by a treadmilling actin network. Actin polymerization has been shown to be mechanically restrained by membrane tension in the lamellipodium. However, it remains unclear how membrane tension is determined, what is responsible for retraction and shaping of the rear boundary, and overall how actin-driven protrusion at the front is coordinated with retraction at the rear. To answer these questions, we utilize lamellipodial fragments from fish epithelial keratocytes which lack a cell body but retain the ability to crawl. The absence of the voluminous cell body in fragments simplifies the relation between lamellipodial geometry and cytoskeletal dynamics. We find that shape and speed are highly correlated over time within individual fragments, whereby faster crawling is accompanied by larger front-to-rear lamellipodial length. Furthermore, we find that the actin network density decays exponentially from front-to-rear indicating a constant net disassembly rate. These findings lead us to a simple hypothesis of a disassembly clock mechanism in which rear position is determined by where the actin network has disassembled enough for membrane tension to crush it and haul it forward. This model allows us to directly relate membrane tension with actin assembly and disassembly dynamics and elucidate the role of the cell membrane as a global mechanical regulator which coordinates protrusion and retraction.cell motility | keratocyte fragments U nderstanding the large-scale coordination of molecular processes into coherent behavior at the cellular level is one of the central challenges in cell biology. Actin-based motility involves numerous molecular players with complex interactions (1) that span a wide-range of scales from the molecular level to the cellular one. Despite substantial progress in characterizing the molecular details involved (2), we still do not understand the remarkable self-organization of these molecular components into a moving cell. The complex interplay between biochemical reactions and biophysical forces plays a central role in this selforganization. In particular, the mechanical feedback between the cell membrane and the dynamic actin network has been shown to have a substantial effect on cell protrusion (3, 4), and a significant role in coordinating protrusion over cellular scales (5, 6). However, quantitative understanding of the coupling between the cell membrane and the motility machinery is still lacking; it is unknown what determines membrane tension and how membrane tension is related to the protrusion and retraction dynamics. Elucidating this dynamic interplay is essential for understanding how overall cell morphology and movement emerge from the underlying molecular processes.Fish epithelial keratocytes are...

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Section: Resultssupporting

confidence: 53%

Section: Discussionmentioning

confidence: 99%

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Actin disassembly clock determines shape and speed of lamellipodial fragments

Ofer

Mogilner

Keren

2011

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

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113

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“…Inhibition of myosin II also caused a mild, although statistically significant reduction in GzmA-mediated death. Given the role of myosin II in regulating actin dynamics in migrating cells, 27 we hypothesise that myosin may also influence the function of the actin network during athetosis. We also discovered that a high proportion of cells undergoing GzmA-mediated alternate death had altered MTOC localisation; however, microtubule-disrupting agents had no effect, indicating that the actin cytoskeleton has the more critical role.…”

Section: Discussionmentioning

confidence: 99%

Mouse granzyme A induces a novel death with writhing morphology that is mechanistically distinct from granzyme B-induced apoptosis

et al. 2013

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Human and mouse granzyme (Gzm)B both induce target cell apoptosis in concert with pore-forming perforin (Pfp); however the mechanisms by which other Gzms induce non-apoptotic death remain controversial and poorly characterised. We used timelapse microscopy to document, quantitatively and in real time, the death of target cells exposed to primary natural killer (NK) cells from mice deficient in key Gzms. We found that in the vast majority of cases, NK cells from wild-type mice induced classic apoptosis. However, NK cells from syngeneic Gzm B-deficient mice induced a novel form of cell death characterised by slower kinetics and a pronounced, writhing, 'worm-like' morphology. Dying cells initially contracted but did not undergo membrane blebbing, and annexin-V staining was delayed until the onset of secondary necrosis. As it is different from any cell death process previously reported, we tentatively termed this cell death 'athetosis'. Two independent lines of evidence showed this alternate form of death was due to Gzm A: first, cell death was revealed in the absence of Gzm B, but was completely lost when the NK cells were deficient in both Gzm A and B; second, the athetotic morphology was precisely reproduced when recombinant mouse Gzm A was delivered by an otherwise innocuous dose of recombinant Pfp. Gzm A-mediated athetosis did not require caspase activation, early mitochondrial disruption or generation of reactive oxygen species, but did require an intact actin cytoskeleton and was abolished by latrunculin B and mycalolide B. This work defines an authentic role for mouse Gzm A in granule-induced cell death by cytotoxic lymphocytes.

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“…Notably, a recent in vitro study of actin networks bundled with fascin showed that myosin could spontaneously disassemble contractile structures (40). There may be an interplay between myosin force production and force-induced dissociation of cross-link proteins that controls the lifetime of cellular contractile structures (52,53). Finally, assembly of cellular actomyosin condensates into ordered structures such as contractile rings depends not only on myosin contractility but also on forces produced by actin disassembly (23,54).…”

Section: Discussionmentioning

confidence: 99%

Active multistage coarsening of actin networks driven by myosin motors

Silva

Depken

Stuhrmann

et al. 2011

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

214

174

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In cells, many vital processes involve myosin-driven motility that actively remodels the actin cytoskeleton and changes cell shape. Here we study how the collective action of myosin motors organizes actin filaments into contractile structures in a simplified model system devoid of biochemical regulation. We show that this self-organization occurs through an active multistage coarsening process. First, motors form dense foci by moving along the actin network structure followed by coalescence. Then the foci accumulate actin filaments in a shell around them. These actomyosin condensates eventually cluster due to motor-driven coalescence. We propose that the physical origin of this multistage aggregation is the highly asymmetric load response of actin filaments: they can support large tensions but buckle easily under piconewton compressive loads. Because the motor-generated forces well exceed this threshold, buckling is induced on the connected actin network that resists motor-driven filament sliding. We show how this buckling can give rise to the accumulation of actin shells around myosin foci and subsequent coalescence of foci into superaggregates. This new physical mechanism provides an explanation for the formation and contractile dynamics of disordered condensed actomyosin states observed in vivo.active gels | molecular motors | nonequilibrium | soft condensed matter C ells undergo dramatic changes in shape and internal organization during vital processes such as migration and division. These changes involve remodeling of the cytoskeleton partly driven by collective physical interactions between molecular motors and cytoskeletal filaments. The motors use adenosine triphosphate (ATP) as fuel and hydrolyze it to actively generate forces and move along filaments (1). Multiheaded motors or complexes of motors may cross-link neighboring filaments and generate relative motion between them. Kinesin and dynein motors interact with microtubules to form the mitotic spindle, which is responsible for chromosome segregation (2). Myosin motors interact with filamentous actin (F-actin) to form complex arrays such as the contractile ring driving cell division (3, 4) and contractile networks that drive cell migration (4) and polarizing cortical flows (5, 6).To identify the biophysical processes underlying cytoskeletal organization, many in vitro model systems of purified motors and filaments that lack biochemical regulation have been recently developed. It is known that kinesins can organize microtubules into polarity sorted asters, as well as vortex or bundle states (7-10). These structures resemble physiological arrays such as the mitotic spindle (11). In contrast to microtubules, purified F-actin does not form well-defined structures when motors are added. Actin-myosin II solutions remain disordered at high levels of ATP (12-14) and generate dense condensates that appear internally unstructured if the ATP level is lowered or when the actin filaments are cross-linked (15-18). Interestingly, similar dense condensates appear in...

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Myosin II contributes to cell-scale actin network treadmilling through network disassembly

Cited by 355 publications

References 29 publications

Actin disassembly clock determines shape and speed of lamellipodial fragments

Actin disassembly clock determines shape and speed of lamellipodial fragments

Mouse granzyme A induces a novel death with writhing morphology that is mechanistically distinct from granzyme B-induced apoptosis

Active multistage coarsening of actin networks driven by myosin motors

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