Arp2/3 Complex and Cofilin Modulate Binding of Tropomyosin to Branched Actin Networks

Hsiao, Jennifer Ying; Goins, Lauren M; Petek, Natalie A.; Mullins, R. Dyche

doi:10.1016/j.cub.2015.04.038

Cited by 53 publications

(56 citation statements)

References 40 publications

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“…Expression of only one Tm isoform has previously been documented in vivo (Hanke et al, 1987). Recently three isoforms were studied in S2 cells (Goins and Mullins, 2015; Hsiao et al, 2015), however most remained uncharacterized, and it was unclear whether the atypical isoforms were expressed or required.…”

Section: Resultsmentioning

confidence: 99%

An Atypical Tropomyosin in Drosophila with Intermediate Filament-like Properties

et al. 2016

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A longstanding mystery has been the absence of cytoplasmic intermediate filaments (IFs) from Drosophila, despite their importance in other organisms. In the course of characterizing the in vivo expression and functions of Drosophila Tropomyosin (Tm) isoforms, we discovered an essential but unusual product of the Tm1 locus, Tm1-I/C, which resembles an IF protein in some respects. Like IFs, Tm1-I/C spontaneously forms filaments in vitro, which are intermediate in diameter between F-actin and microtubules. Like IFs, but unlike canonical Tms, Tm1-I/C contains N- and C-terminal low complexity domains flanking a central coiled coil. In vivo, Tm1-I/C forms cytoplasmic filaments that do not associate with F-actin or canonical Tms. Tm1-I/C is essential for collective border cell migration, in epithelial cells for proper cytoarchitecture, and in the germline for formation of germ plasm. These results suggest that flies have evolved a distinctive type of cytoskeletal filament from Tm.

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Section: Resultsmentioning

confidence: 99%

An Atypical Tropomyosin in Drosophila with Intermediate Filament-like Properties

et al. 2016

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“…One explanation is that after nucleating at a single site of F-actin (e.g. the filament ends), nascent tropomyosin chains grow uniformly from that point (Johnson et al 2014; Hsiao et al 2015; Gunning et al 2015). Alternatively, tropomyosin may initially bind at multiple sites, and followed either by a stochastic mechanism of nucleation/annihilation or by means of axial diffusion, gaps are filled between growing tropomyosin chains on F-actin (Holmes and Lehman 2008; Schmidt et al 2015).…”

Section: Resultsmentioning

confidence: 99%

Electrostatic interaction map reveals a new binding position for tropomyosin on F-actin

Rynkiewicz

Schott

Orzechowski

et al. 2015

J Muscle Res Cell Motil

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Azimuthal movement of tropomyosin around the F-actin thin filament is responsible for muscle activation and relaxation. Recently a model of αα-tropomyosin, derived from molecular-mechanics and electron microscopy of different contractile states, showed that tropomyosin is rather stiff and pre-bent to present one specific face to F-actin during azimuthal transitions. However, a new model based on cryo-EM of troponin- and myosin-free filaments proposes that the interacting-face of tropomyosin can differ significantly from that in the original model. Because resolution was insufficient to assign tropomyosin side-chains, the interacting-face could not be unambiguously determined. Here, we use structural analysis and energy landscapes to further examine the proposed models. The observed bend in seven crystal structures of tropomyosin is much closer in direction and extent to the original model than to the new model. Additionally, we computed the interaction map for repositioning tropomyosin over the F-actin surface, but now extended over a much larger surface than previously (using the original interacting-face). This map shows two energy minima— one corresponding to the “blocked-state” as in the original model, and the other related by a simple 24 Å translation of tropomyosin parallel to the F-actin axis. The tropomyosinactin complex defined by the second minimum fits perfectly into the recent cryo-EM density, without requiring any change in the interacting-face. Together, these data suggest that movement of tropomyosin between regulatory states does not require interacting-face rotation. Further, they imply that thin filament assembly may involve an interplay between initially seeded tropomyosin molecules growing from distinct binding-site regions on actin.

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“…Human fascin and both Human and C. elegans α-actinin were labeled with either Cy5-Monomaleimide (GE Healthcare) or TMR-6-Maleimide (Life Technologies, Grand Island, NY). Skeletal muscle tropomyosin was purified from rabbit muscle acetone powder (Pel Freez Biologicals) and labeled as described [46]. Proteins containing SNAP fusions were labeled with SNAP-surface-549 or SNAP-surface-647 (New England BioLabs).…”

Section: Methodsmentioning

confidence: 99%

Fascin- and α-Actinin-Bundled Networks Contain Intrinsic Structural Features that Drive Protein Sorting

et al. 2016

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SUMMARY Cells assemble and maintain functionally distinct actin cytoskeleton networks with various actin filament organizations and dynamics through the coordinated action of different sets of actin binding proteins. The biochemical and functional properties of diverse actin binding proteins, both alone and in combination, have been increasingly well studied. Conversely, how different sets of actin binding proteins properly sort to distinct actin filament networks in the first place is not nearly as well understood. Actin binding protein sorting is critical for the self-organization of diverse dynamic actin cytoskeleton networks within a common cytoplasm. Using in vitro reconstitution techniques including biomimetic assays and single molecule multi-color TIRF microscopy, we discovered that sorting of the prominent actin bundling proteins fascin and α-actinin to distinct networks is an intrinsic behavior, free of complicated cellular signaling cascades. When mixed, fascin and α-actinin mutually exclude each other by promoting their own recruitment and inhibiting recruitment of the other, resulting in the formation of distinct fascin- or α-actinin-bundled domains. Subdiffraction-resolution light microscopy and negative staining electron microscopy revealed that fascin domains are densely packed, while α-actinin domains consist of widely spaced parallel actin filaments. Importantly, other actin binding proteins such as fimbrin and espin show high specificity between these two bundle types within the same reaction. Here we directly observe that fascin and α-actinin intrinsically segregate to discrete bundled domains that are specifically recognized by other actin binding proteins.

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Arp2/3 Complex and Cofilin Modulate Binding of Tropomyosin to Branched Actin Networks

Cited by 53 publications

References 40 publications

An Atypical Tropomyosin in Drosophila with Intermediate Filament-like Properties

An Atypical Tropomyosin in Drosophila with Intermediate Filament-like Properties

Electrostatic interaction map reveals a new binding position for tropomyosin on F-actin

Fascin- and α-Actinin-Bundled Networks Contain Intrinsic Structural Features that Drive Protein Sorting

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