Faculty Opinions recommendation of Cytokinesis requires localized β-actin filament production by an actin isoform specific nucleator.

Most nonmuscle cells of higher vertebrates contain two different actin isoforms, beta- and gamma-cytoplasmic actin. The beta-isoform is with few exceptions the predominant isoform in nonmuscle cells and tissues. Perturbation of the beta:gamma ratio has been shown to affect the organization of bundled actin filaments indicating that the beta- and gamma-genes encode functionally distinct cytoarchitectural information. In the present study we localized by immunostaining beta- and gamma-actin in chicken auditory hair cells. These highly specialized cells serve as model system for studying certain developmental and structural aspects of a complex actin filament system with high architectural precision. We show that gamma-actin is the predominant actin isoform in auditory hair cells with an apparent beta:gamma ratio of approximately 1:2. gamma-Actin is not sorted and occurs in all three actin assemblies of the hair border, i.e. the cores of sensory hairs (stereocilia), the subjacent gel-like actin filament meshwork (cuticular plate) and the zonula adherens ring. In contrast to gamma-actin, the beta-isoform is specifically sorted to the actin filament core bundle of stereocilia that is extensively crosslinked by fimbrin. In view of recent studies showing that L-plastin, the leukocyte homolog of fimbrin, has a higher binding affinity for beta-actin than for gamma-actin, a mechanism is proposed for how hair cells might restrict formation of actin filament bundles to a single cellular site (i.e. the stereocilia). The limited level of expression of beta-actin in hair cells may help to prevent ectopic bundle formation in other cellular compartments.

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Cytokinesis requires localized β-actin filament production by an actin isoform specific nucleator

Nature Communications ◽

10.1038/s41467-017-01231-x ◽

2017 ◽

Vol 8 (1) ◽

Cited By ~ 23

Author(s):

A. Chen ◽

P. D. Arora ◽

C. A. McCulloch ◽

A. Wilde

Keyword(s):

Actin Filament ◽

Actin Isoform

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Inhibition of polar actin assembly by astral microtubules is required for cytokinesis

Nature Communications ◽

10.1038/s41467-021-22677-0 ◽

2021 ◽

Vol 12 (1) ◽

Author(s):

Anan Chen ◽

Luisa Ulloa Severino ◽

Thomas C. Panagiotou ◽

Trevor F. Moraes ◽

Darren A. Yuen ◽

...

Keyword(s):

Cell Division ◽

Actin Cytoskeleton ◽

Actin Filament ◽

Cell Cortex ◽

Actin Assembly ◽

Actin Network ◽

Contractile Ring ◽

Actin Networks ◽

Membrane Blebbing ◽

Actin Isoform

AbstractDuring cytokinesis, the actin cytoskeleton is partitioned into two spatially distinct actin isoform specific networks: a β-actin network that generates the equatorial contractile ring, and a γ-actin network that localizes to the cell cortex. Here we demonstrate that the opposing regulation of the β- and γ-actin networks is required for successful cytokinesis. While activation of the formin DIAPH3 at the cytokinetic furrow underlies β-actin filament production, we show that the γ-actin network is specifically depleted at the cell poles through the localized deactivation of the formin DIAPH1. During anaphase, CLIP170 is delivered by astral microtubules and displaces IQGAP1 from DIAPH1, leading to formin autoinhibition, a decrease in cortical stiffness and localized membrane blebbing. The contemporaneous production of a β-actin contractile ring at the cell equator and loss of γ-actin from the poles is required to generate a stable cytokinetic furrow and for the completion of cell division.

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Tropomyosin isoforms segregate into distinct clusters on single actin filaments

10.1101/2021.07.23.453476 ◽

2021 ◽

Author(s):

Peyman Obeidy ◽

Tom Sobey ◽

Philip R. Nicovich ◽

Adelle C. F. Coster ◽

Elvis Pandzic

Keyword(s):

Amino Acids ◽

Actin Filament ◽

Actin Filaments ◽

Automated Image Analysis ◽

Tropomyosin Isoforms ◽

Low Concentrations ◽

Image Analysis Algorithm ◽

Actin Isoform ◽

Spatio Temporal ◽

Assembly Intermediate

Tropomyosins (Tpm) are rod-shaped proteins that interact head-to-tail to form a continuous polymer along both sides of most cellular actin filaments. Head-to-tail interaction between adjacent Tpm molecules and the formation of an overlap complex between them leads to the assembly of actin filaments with one type of Tpm isoform in time and space. Variations in the affinity of tropomyosin isoforms for different actin structures are proposed as a potential sorting mechanism. However, the detailed mechanisms of spatio-temporal sorting of Tpms remain elusive. In this study, we investigated the early intermediates during actin-tropomyosin filament assembly, using skeletal/cardiac Tpm isoform (Tpm1.1) and a cytoskeletal isoform (Tpm1.6) that differ only in the last 27 amino acids. We investigated how the muscle isoform Tpm1.1 and the cytoskeletal isoform Tpm1.6 nucleate domains on the actin filament and tested whether (1) recruitment is affected by the actin isoform (muscle vs cytoskeletal) and (2) whether there is specificity in recruiting the same isoform to a domain at these early stages. To address these questions, actin filaments were exposed to low concentrations of fluorescent tropomyosins in solution. The filaments were immobilized onto glass coverslips and the pattern of decoration was visualized by TIRF microscopy. We show that at the early assembly stage, tropomyosins formed multiple distinct fluorescent domains (here termed "cluster") on the actin filaments. An automated image analysis algorithm was developed and validated to identify clusters and estimate the number of tropomyosins in each cluster. The analysis showed that tropomyosin isoform sorting onto an actin filament is unlikely to be driven by a preference for nucleating on the corresponding muscle or cytoskeletal actin isoforms but rather is facilitated by a higher probability of incorporating the same tropomyosin isoforms into an early assembly intermediate. We showed that the 27 amino acids at the end of each tropomyosin seem to provide enough molecular information for attachment of the same tropomyosin isoforms adjacent to each other on an actin filament. This results in the formation of homogeneous clusters composed of the same isoform rather than clusters with mixed isoforms.

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Toward an alignment of the actin molecule within the actin filament

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100075981 ◽

1983 ◽

Vol 41 ◽

pp. 450-451

Author(s):

P.R. Smith ◽

W.E. Fowler ◽

U. Aebi

Keyword(s):

Actin Filament ◽

Diffraction Data ◽

Quantitative Estimate ◽

Molecular Model ◽

Quantitative Comparison ◽

Regulatory Proteins ◽

Essential Information ◽

X Ray ◽

Maximum Resolution ◽

Lack Of Information

An understanding of the specific interactions of actin with regulatory proteins has been limited by the lack of information about the structure of the actin filament. Molecular actin has been studied in actin-DNase I complexes by single crystal X-ray analysis, to a resolution of about 0.6nm, and in the electron microscope where two dimensional actin sheets have been reconstructed to a maximum resolution of 1.5nm. While these studies have shown something of the structure of individual actin molecules, essential information about the orientation of actin in the filament is still unavailable.The work of Egelman & DeRosier has, however, suggested a method which could be used to provide an initial quantitative estimate of the orientation of actin within the filament. This method involves the quantitative comparison of computed diffraction data from single actin filaments with diffraction data derived from synthetic filaments constructed using the molecular model of actin as a building block. Their preliminary work was conducted using a model consisting of two juxtaposed spheres of equal size.

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The 3-Dimensional Molecular Structure of the Actin Filament Revisited

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100119223 ◽

1985 ◽

Vol 43 ◽

pp. 480-481

Author(s):

U. Aebi ◽

R. Millonig ◽

H. Salvo

Keyword(s):

Actin Filament ◽

Supramolecular Assembly ◽

Specimen Preparation ◽

X Ray Diffraction ◽

Single Filament ◽

X Ray ◽

3 Dimensional ◽

Filament Diameter ◽

Preparation Conditions ◽

Apparent Diameter

To date, most 3-D reconstructions of undecorated actin filaments have been obtained from actin filament paracrystal data (for refs, see 1,2). However, due to the fact that (a) the paracrystals may be several filament layers thick, and (b) adjacent filaments may sustantially interdigitate, these reconstructions may be subject to significant artifacts. None of these reconstructions has permitted unambiguous tracing or orientation of the actin subunits within the filament. Furthermore, measured values for the maximal filament diameter both determined by EM and by X-ray diffraction analysis, vary between 6 and 10 nm. Obviously, the apparent diameter of the actin filament revealed in the EM will critically depend on specimen preparation, since it is a rather flexible supramolecular assembly which can easily be bent or distorted. To resolve some of these ambiguities, we have explored specimen preparation conditions which may preserve single filaments sufficiently straight and helically ordered to be suitable for single filament 3-D reconstructions, possibly revealing molecular detail.

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