Myosin-10 and actin filaments are essential for mitotic spindle function

Woolner, Sarah; O’Brien, Lori L.; Wiese, Christiane; Bement, William M.

doi:10.1083/jcb.200804062

Cited by 199 publications

(312 citation statements)

References 44 publications

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“…They convert the chemical energy derived from ATP hydrolysis into directed motion along actin and are responsible for a wide variety of intracellular motilities and for muscle contraction. Mammalian myosin-10 was originally found to localize to the tips of filopodia (3,4) and is known to be important for filopodial formation and extension (4-6), phagocytosis (7), cell migration (8), and mitotic spindle length maintenance and anchoring (9). It is an ∼240-kDa protein consisting of an N-terminal, consensus motor region (which binds actin, hydrolyses ATP, and produces force and movement), a light chain binding domain (LCBD) with three IQ motifs (10), and a stable single alpha-helix (SAH) followed by a tail region containing, an antiparallel coiled-coil (APCC) region, a proline, glutamate, serine, and threonine rich (PEST) domain, three pleckstrin homology (PH) domains, a myosin tail homology 4 (MyTH4) domain, and a band 4.1, ezrin, radixin, moesin (FERM) domain (Fig.…”

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confidence: 99%

Myosin-10 produces its power-stroke in two phases and moves processively along a single actin filament under low load

Takagi

Farrow

Billington

et al. 2014

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

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Myosin-10 is an actin-based molecular motor that participates in essential intracellular processes such as filopodia formation/ extension, phagocytosis, cell migration, and mitotic spindle maintenance. To study this motor protein's mechano-chemical properties, we used a recombinant, truncated form of myosin-10 consisting of the first 936 amino acids, followed by a GCN4 leucine zipper motif, to force dimerization. Negative-stain electron microscopy reveals that the majority of molecules are dimeric with a head-to-head contour distance of ∼50 nm. In vitro motility assays show that myosin-10 moves actin filaments smoothly with a velocity of ∼310 nm/s. Steady-state and transient kinetic analysis of the ATPase cycle shows that the ADP release rate (∼13 s −1 ) is similar to the maximum ATPase activity (∼12-14 s −1 ) and therefore contributes to rate limitation of the enzymatic cycle. Single molecule optical tweezers experiments show that under intermediate load (∼0.5 pN), myosin-10 interacts intermittently with actin and produces a power stroke of ∼17 nm, composed of an initial 15-nm and subsequent 2-nm movement. At low optical trap loads, we observed staircaselike processive movements of myosin-10 interacting with the actin filament, consisting of up to six ∼35-nm steps per binding interaction. We discuss the implications of this load-dependent processivity of myosin-10 as a filopodial transport motor.actomyosin | stable single alpha-helix | optical trapping | myosin X | myosin-5a

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mentioning

confidence: 99%

Myosin-10 produces its power-stroke in two phases and moves processively along a single actin filament under low load

Takagi

Farrow

Billington

et al. 2014

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

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“…Myosin X is found at the mitotic and meiotic spindle, where it sets the orientation of the spindle relative to the substrate and influences spindle length (3,4). However, myosin X is more commonly found in interphase cells at the tips of filopodia (5).…”

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confidence: 99%

Structured Post-IQ Domain Governs Selectivity of Myosin X for Fascin-Actin Bundles

Nagy

Rock

2010

Journal of Biological Chemistry

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Without guidance cues, cytoskeletal motors would traffic components to the wrong destination with disastrous consequences for the cell. Recently, we identified a motor protein, myosin X, that identifies bundled actin filaments for transport. These bundles direct myosin X to a unique destination, the tips of cellular filopodia. Because the structural and kinetic features that drive bundle selection are unknown, we employed a domain-swapping approach with the nonselective myosin V to identify the selectivity module of myosin X. We found a surprising role of the myosin X tail region (post-IQ) in supporting long runs on bundles. Moreover, the myosin X head is adapted for initiating processive runs on bundles. We found that the tail is structured and biases the orientation of the two myosin X heads because a targeted insertion that introduces flexibility in the tail abolishes selectivity. Together, these results suggest how myosin motors may manage to read cellular addresses.The essential role of cytoskeletal motor proteins in organizing cellular compartments and cargoes is widely accepted (1). However, many of the molecular details of the overall transport and organization process are poorly understood. These essential features include how motors engage cargoes at their source, how motors are activated once they engage cargo, how they choose the correct set of cytoskeletal tracks, how they disengage from cargo at their destination, and how they return to sources of cargo. Clearly, errors at any of these stages would lead to faults in cellular organization and misplaced cargoes.To begin to address these questions we have focused on a particular motor protein, myosin X, due to its ability to navigate to a precise location within the cell (2). Myosin X is found at the mitotic and meiotic spindle, where it sets the orientation of the spindle relative to the substrate and influences spindle length (3, 4). However, myosin X is more commonly found in interphase cells at the tips of filopodia (5). These long, slender projections at the leading edge of migrating cells are involved in cell motility and environmental sensing. Myosin X transports components such as Ena/VASP and integrins that are found in the filopodial tip complex (2, 6). The early work by Berg et al. (5) demonstrated that myosin X reaches filopodial tips under its own power. Recently, we found that myosin X identifies filopodia by recognizing the fascin-bundled actin filaments found in the filopodial core (7). Myosin X takes long processive runs along these tightly bundled actin filaments while largely ignoring isolated actin filaments found throughout the cell. In contrast, myosin V does not distinguish between single actin filaments and bundled actin filaments when taking processive runs.Here, we seek to identify the "selectivity module" that allows myosin X to recognize bundled actin filaments. Our first approach is to swap similar domains between the selective myosin X and the nonselective myosin V. In this approach, we are aided by the fact that both m...

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“…Given that Myo10 interacts with TPX2 22 and ADD1 42, this prompted us to examine whether TPX2 interacts with ADD1 through Myo10. Indeed, we found that TPX2 interacted with ADD1 in mitosis, but the interaction was not detected in interphase (Fig 4A).…”

Section: Resultsmentioning

confidence: 99%

“…Myo10 is the ADD1 binding partner on mitotic spindles 42 and its role in spindle positioning and spindle length control is F‐actin‐dependent 22. In fact, the spindle F‐actin has been shown to surround mitotic spindles during early Xenopus embryo mitosis 22. Arp2/3 protein complex, which is involved in the nucleation and assembly of branched actin filaments, has been shown to participate in the formation of the spindle F‐actin 52, 53.…”

Section: Discussionmentioning

confidence: 99%

Adducin‐1 is essential for spindle pole integrity through its interaction with TPX2

et al. 2018

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Bipolar spindle assembly is necessary to ensure the proper progression of cell division. Loss of spindle pole integrity leads to multipolar spindles and aberrant chromosomal segregation. However, the mechanism underlying the maintenance of spindle pole integrity remains unclear. In this study, we show that the actin‐binding protein adducin‐1 (ADD1) is phosphorylated at S726 during mitosis. S726‐phosphorylated ADD1 localizes to centrosomes, wherein it organizes into a rosette‐like structure at the pericentriolar material. ADD1 depletion causes centriole splitting and therefore results in multipolar spindles during mitosis, which can be restored by re‐expression of ADD1 and the phosphomimetic S726D mutant but not by the S726A mutant. Moreover, the phosphorylation of ADD1 at S726 is crucial for its interaction with TPX2, which is essential for spindle pole integrity. Together, our findings unveil a novel function of ADD1 in maintaining spindle pole integrity through its interaction with TPX2.

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Myosin-10 and actin filaments are essential for mitotic spindle function

Abstract: Abbreviations used in this paper: HMM, heavy meromyosin; LatB, latrunculin B; MO, morpholino; Myo10, myosin-10.The online version of this paper contains supplemental material.

Cited by 199 publications

References 44 publications

Myosin-10 produces its power-stroke in two phases and moves processively along a single actin filament under low load

Myosin-10 produces its power-stroke in two phases and moves processively along a single actin filament under low load

Structured Post-IQ Domain Governs Selectivity of Myosin X for Fascin-Actin Bundles

Adducin‐1 is essential for spindle pole integrity through its interaction with TPX2

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