scholarly journals Heterodimeric capping protein is required for stereocilia length and width regulation

2017 ◽  
Vol 216 (11) ◽  
pp. 3861-3881 ◽  
Author(s):  
Matthew R. Avenarius ◽  
Jocelyn F. Krey ◽  
Rachel A. Dumont ◽  
Clive P. Morgan ◽  
Connor B. Benson ◽  
...  
Keyword(s):  
Hair Cell ◽  
Hair Cells ◽  
Actin Filaments ◽  
Actin Polymerization ◽  
Vestibular Function ◽  
In Utero ◽  
In Utero Electroporation ◽  
Interacting Protein ◽  
Capping Protein ◽  
Dimension Control

Control of the dimensions of actin-rich processes like filopodia, lamellipodia, microvilli, and stereocilia requires the coordinated activity of many proteins. Each of these actin structures relies on heterodimeric capping protein (CAPZ), which blocks actin polymerization at barbed ends. Because dimension control of the inner ear’s stereocilia is particularly precise, we studied the CAPZB subunit in hair cells. CAPZB, present at ∼100 copies per stereocilium, concentrated at stereocilia tips as hair cell development progressed, similar to the CAPZB-interacting protein TWF2. We deleted Capzb specifically in hair cells using Atoh1-Cre, which eliminated auditory and vestibular function. Capzb-null stereocilia initially developed normally but later shortened and disappeared; surprisingly, stereocilia width decreased concomitantly with length. CAPZB2 expressed by in utero electroporation prevented normal elongation of vestibular stereocilia and irregularly widened them. Together, these results suggest that capping protein participates in stereocilia widening by preventing newly elongating actin filaments from depolymerizing.

On the Legacy of Genetically Altered Mouse Models to Explore Vestibular Function: Distribution of Vestibular Hair Cell Phenotypes in the Otoferlin-Null Mouse

2019 ◽  
Vol 128 (6_suppl) ◽  
pp. 125S-133S ◽  
Author(s):  
Terry J. Prins ◽  
Johnny J. Saldate ◽  
Gerald S. Berke ◽  
Larry F. Hoffman
Keyword(s):  
Hair Cell ◽  
Hair Cells ◽  
Vestibular Function ◽  
Null Mouse ◽  
Type I ◽  
Vestibular Hypofunction ◽  
Cellular Changes ◽  
Sensory Epithelia ◽  
Vestibular Sensory Epithelia ◽  
Cell Phenotypes

Objectives: Early in his career, David Lim recognized the scientific impact of genetically anomalous mice exhibiting otoconia agenesis as models of drastically compromised vestibular function. While these studies focused on the mutant pallid mouse, contemporary genetic tools have produced other models with engineered functional modifications. Lim and colleagues foresaw the need to analyze vestibular epithelia from pallid mice to verify the absence of downstream consequences that might be secondary to the altered load represented by otoconial agenesis. More generally, however, such comparisons also contribute to an understanding of the susceptibility of labyrinthine sensory epithelia to more widespread cellular changes associated with what may appear as isolated modifications. Methods: Our laboratory utilizes a model of vestibular hypofunction produced through genetic alteration, the otoferlin-null mouse, which has been shown to exhibit severely compromised stimulus-evoked neurotransmitter release in type I hair cells of the utricular striola. The present study, reminiscent of early investigations of Lim and colleagues that explored the utility of a genetically altered mouse to explore its utility as a model of vestibular hypofunction, endeavored to compare the expression of the hair cell marker oncomodulin in vestibular epithelia from wild-type and otoferlin-null mice. Results: We found that levels of oncomodulin expression were much greater in type I than type II hair cells, though were similar across the 3 genotypes examined (ie, including heterozygotes). Conclusion: These findings support the notion that modifications resulting in a specific component of vestibular hypofunction are not accompanied by widespread morphologic and cellular changes in the vestibular sensory epithelia.

scholarly journals The Huntingtin-interacting protein SETD2/HYPB is an actin lysine methyltransferase

2020 ◽  
Vol 6 (40) ◽  
pp. eabb7854 ◽  
Author(s):  
Riyad N. H. Seervai ◽  
Rahul K. Jangid ◽  
Menuka Karki ◽  
Durga Nand Tripathi ◽  
Sung Yun Jung ◽  
...  
Keyword(s):  
Cell Migration ◽  
Actin Filaments ◽  
Actin Polymerization ◽  
Scaffolding Protein ◽  
Actin Binding ◽  
Set Domain ◽  
Interacting Protein ◽  
Lysine Methyltransferase ◽  
Huntingtin Interacting Protein ◽  
In Cells

The methyltransferase SET domain–containing 2 (SETD2) was originally identified as Huntingtin (HTT) yeast partner B. However, a SETD2 function associated with the HTT scaffolding protein has not been elucidated, and no linkage between HTT and methylation has yet been uncovered. Here, we show that SETD2 is an actin methyltransferase that trimethylates lysine-68 (ActK68me3) in cells via its interaction with HTT and the actin-binding adapter HIP1R. ActK68me3 localizes primarily to the insoluble F-actin cytoskeleton in cells and regulates actin polymerization/depolymerization dynamics. Disruption of the SETD2-HTT-HIP1R axis inhibits actin methylation, causes defects in actin polymerization, and impairs cell migration. Together, these data identify SETD2 as a previously unknown HTT effector regulating methylation and polymerization of actin filaments and provide new avenues for understanding how defects in SETD2 and HTT drive disease via aberrant cytoskeletal methylation.

scholarly journals Actin filaments, stereocilia, and hair cells of the bird cochlea. I. Length, number, width, and distribution of stereocilia of each hair cell are related to the position of the hair cell on the cochlea.

1983 ◽  
Vol 96 (3) ◽  
pp. 807-821 ◽  
Author(s):  
L G Tilney ◽  
J C Saunders
Keyword(s):  
Hair Cell ◽  
Surface Area ◽  
Hair Cells ◽  
Actin Filaments ◽  
Hexagonal Lattice ◽  
Frequency Discrimination ◽  
Sensory Epithelium ◽  
Scanning Microscopy ◽  
Apical Surface ◽  
Length Width

Located on the sensory epithelium of the sickle-shaped cochlea of a 7- to 10-d-old chick are approximately 5,000 hair cells. When the apical surface of these cell is examined by scanning microscopy, we find that the length, number, width, and distribution of the stereocilia on each hair cell are predetermined. Thus, a hair cell located at the distal end of the cochlea has 50 stereocilia, the longest of which are 5.5 microns in length and 0.12 microns in width, while those at the proximal end number 300 and are maximally 1.5 microns in length and 0.2 micron in width. In fact, if we travel along the cochlea from its distal to proximal end, we see that the stereocilia on successive hair cells gradually increase in number and width, yet decrease in length. Also, if we look transversely across the cochlea where adjacent hair cells have the same length and number of stereocilia (they are the same distance from the distal end of the cochlea), we find that the stereocilia of successive hair cells become thinner and that the apical surface area of the hair cell proper, not including the stereocilia, decreases from a maximum of 80 microns2 to 15 microns2. Thus, if we are told the length of the longest stereocilium on a hair cell and the width of that stereocilium, we can pinpoint the position of that hair cell on the cochlea in two axes. Likewise, if we are told the number of stereocilia and the apical surface of a hair cell, we can pinpoint the location of that cell in two axes. The distribution of the stereocilia on the apical surface of the cell is also precisely determined. More specifically, the stereocilia are hexagonally packed and this hexagonal lattice is precisely positioned relative to the kinocilium. Because of the precision with which individual hair cells regulate the length, width, number, and distribution of their cell extensions, we have a magnificent object with which to ask questions about how actin filaments that are present within the cell are regulated. Equally interesting is that the gradient in stereociliary length, number, width, and distribution may play an important role in frequency discrimination in the cochlea. This conclusion is amplified by the information presented in the accompanying paper (Tilney, L.G., E.H. Egelman, D.J. DeRosier, and J.C. Saunders, 1983, J. Cell Biol., 96:822-834) on the packing of actin filaments in this stereocilia.

scholarly journals Actin filaments, stereocilia, and hair cells of the bird cochlea. V. How the staircase pattern of stereociliary lengths is generated

1988 ◽  
Vol 106 (2) ◽  
pp. 355-365 ◽  
Author(s):  
LG Tilney ◽  
MS Tilney ◽  
DA Cotanche
Keyword(s):  
Hair Cell ◽  
Hair Cells ◽  
Actin Filaments ◽  
Developmental Time ◽  
Final Phase ◽  
Set Up

The stereocilia on each hair cell are arranged into rows of ascending height, resulting in what we refer to as a "staircase-like" profile. At the proximal end of the cochlea the length of the tallest row of stereocilia in the staircase is 1.5 micron, with the shortest row only 0.3 micron. As one proceeds towards the distal end of the cochlea the length of the stereocilia progressively increases so that at the extreme distal end the length of the tallest row of the staircase is 5.5 micron and the shortest row is 2 micron. During development hair cells form their staircases in four phases of growth separated from each other by developmental time. First, stereocilia sprout from the apical surfaces of the hair cells (8-10-d embryos). Second (10-12-d embryos), what will be the longest row of the staircase begins to elongate. As the embryo gets older successive rows of stereocilia initiate elongation. Thus the staircase is set up by the sequential initiation of elongation of stereociliary rows located at increased distances from the row that began elongation. Third (12-17-d embryos), all the stereocilia in the newly formed staircase elongate until those located on the first step of the staircase have reached the prescribed length. In the final phase (17-d embryos to hatchlings) there is a progressive cessation of elongation beginning with the shortest step and followed by taller and taller rows with the tallest step stopping last. Thus, to obtain a pattern of stereocilia in rows of increasing height what transpires are progressive go signals followed by a period when all the stereocilia grow and ending with progressive stop signals. We discuss how such a sequence could be controlled.

scholarly journals The actin filament content of hair cells of the bird cochlea is nearly constant even though the length, width, and number of stereocilia vary depending on the hair cell location.

1988 ◽  
Vol 107 (6) ◽  
pp. 2563-2574 ◽  
Author(s):  
L G Tilney ◽  
M S Tilney
Keyword(s):  
Hair Cell ◽  
Actin Filament ◽  
Cross Sections ◽  
Hair Cells ◽  
Actin Filaments ◽  
Membrane Surface ◽  
Scanning Electron Micrographs ◽  
Length Width ◽  
Filament Content ◽  
Filament Bundle

By direct counts off scanning electron micrographs, we determined the number of stereocilia per hair cell of the chicken cochlea as a function of the position of the hair cell on the cochlea. Micrographs of thin cross sections of stereociliary bundles located at known positions on the cochlea were enlarged and the total number of actin filaments per stereocilium was counted and recorded. By comparing the counts of filament number with measurements of actin filament bundle width of the same stereocilium, we were able to relate actin filament bundle width to filament number with an error margin (r2) of 16%. Combining this data with data already published or in the process of publication from our laboratory on the length and width of stereocilia, we were able to calculate the total length of actin filaments present in stereociliary bundles of hair cells located at a variety of positions on the cochlea. We found that stereociliary bundles of hair cells contain 80,000-98,000 micron of actin filament, i.e., the concentration of actin is constant in all hair cells with a range of values that is less than our error in measurement and/or biological variation, the greatest variation being in relating the diameters of the stereocilia to filament number. We also calculated the membrane surface needed to cover the stereocilia of hair cells located throughout the cochlea. The values (172-192 micron 2) are also constant. The implications of our observation that the total amount of actin is constant even though the length, width, and number of stereocilia per hair cell vary are discussed.

scholarly journals High Rates of Actin Filament Turnover in Budding Yeast and Roles for Actin in Establishment and Maintenance of Cell Polarity Revealed Using the Actin Inhibitor Latrunculin-A

1997 ◽  
Vol 137 (2) ◽  
pp. 399-416 ◽  
Author(s):  
Kathryn R. Ayscough ◽  
Joel Stryker ◽  
Navin Pokala ◽  
Miranda Sanders ◽  
Phil Crews ◽  
...  
Keyword(s):  
Cell Surface ◽  
Actin Cytoskeleton ◽  
Cell Polarity ◽  
Actin Filaments ◽  
Three Dimensional ◽  
Surface Growth ◽  
Interacting Protein ◽  
Capping Protein ◽  
Latrunculin A

We report that the actin assembly inhibitor latrunculin-A (LAT-A) causes complete disruption of the yeast actin cytoskeleton within 2–5 min, suggesting that although yeast are nonmotile, their actin filaments undergo rapid cycles of assembly and disassembly in vivo. Differences in the LAT-A sensitivities of strains carrying mutations in components of the actin cytoskeleton suggest that tropomyosin, fimbrin, capping protein, Sla2p, and Srv2p act to increase actin cytoskeleton stability, while End3p and Sla1p act to decrease stability. Identification of three LAT-A resistant actin mutants demonstrated that in vivo effects of LAT-A are due specifically to impairment of actin function and implicated a region on the three-dimensional actin structure as the LAT-A binding site. LAT-A was used to determine which of 19 different proteins implicated in cell polarity development require actin to achieve polarized localization. Results show that at least two molecular pathways, one actindependent and the other actin-independent, underlie polarity development. The actin-dependent pathway localizes secretory vesicles and a putative vesicle docking complex to sites of cell surface growth, providing an explanation for the dependence of polarized cell surface growth on actin function. Unexpectedly, several proteins that function with actin during cell polarity development, including an unconventional myosin (Myo2p), calmodulin, and an actin-interacting protein (Bud6/Aip3p), achieved polarized localization by an actin-independent pathway, revealing interdependence among cell polarity pathways. Finally, transient actin depolymerization caused many cells to abandon one bud site or mating projection and to initiate growth at a second site. Thus, actin filaments are also required for maintenance of an axis of cell polarity.

scholarly journals Conserved and Divergent Principles of Planar Polarity Revealed by Hair Cell Development and Function

2021 ◽  
Vol 15 ◽  
Author(s):  
Michael R. Deans
Keyword(s):  
Hair Cell ◽  
Inner Ear ◽  
Hair Cells ◽  
Model Organism ◽  
Vestibular Function ◽  
Sensory Receptor ◽  
Apical Surface ◽  
Planar Polarity ◽  
Vestibular Hair Cells ◽  
Sensory Epithelia

Planar polarity describes the organization and orientation of polarized cells or cellular structures within the plane of an epithelium. The sensory receptor hair cells of the vertebrate inner ear have been recognized as a preeminent vertebrate model system for studying planar polarity and its development. This is principally because planar polarity in the inner ear is structurally and molecularly apparent and therefore easy to visualize. Inner ear planar polarity is also functionally significant because hair cells are mechanosensors stimulated by sound or motion and planar polarity underlies the mechanosensory mechanism, thereby facilitating the auditory and vestibular functions of the ear. Structurally, hair cell planar polarity is evident in the organization of a polarized bundle of actin-based protrusions from the apical surface called stereocilia that is necessary for mechanosensation and when stereociliary bundle is disrupted auditory and vestibular behavioral deficits emerge. Hair cells are distributed between six sensory epithelia within the inner ear that have evolved unique patterns of planar polarity that facilitate auditory or vestibular function. Thus, specialized adaptations of planar polarity have occurred that distinguish auditory and vestibular hair cells and will be described throughout this review. There are also three levels of planar polarity organization that can be visualized within the vertebrate inner ear. These are the intrinsic polarity of individual hair cells, the planar cell polarity or coordinated orientation of cells within the epithelia, and planar bipolarity; an organization unique to a subset of vestibular hair cells in which the stereociliary bundles are oriented in opposite directions but remain aligned along a common polarity axis. The inner ear with its complement of auditory and vestibular sensory epithelia allows these levels, and the inter-relationships between them, to be studied using a single model organism. The purpose of this review is to introduce the functional significance of planar polarity in the auditory and vestibular systems and our contemporary understanding of the developmental mechanisms associated with organizing planar polarity at these three cellular levels.

scholarly journals Regulation of Sodium Channel Activity by Capping of Actin Filaments

2003 ◽  
Vol 14 (4) ◽  
pp. 1709-1716 ◽  
Author(s):  
Ekaterina V. Shumilina ◽  
Yuri A. Negulyaev ◽  
Elena A. Morachevskaya ◽  
Horst Hinssen ◽  
Sofia Yu Khaitlina
Keyword(s):  
Plasma Membrane ◽  
Sodium Channel ◽  
Actin Filaments ◽  
Actin Polymerization ◽  
Actin Dynamics ◽  
Channel Activity ◽  
Capping Protein ◽  
Channel Inactivation ◽  
Actin Capping Protein ◽  
Actin Capping

Ion transport in various tissues can be regulated by the cortical actin cytoskeleton. Specifically, involvement of actin dynamics in the regulation of nonvoltage-gated sodium channels has been shown. Herein, inside-out patch clamp experiments were performed to study the effect of the heterodimeric actin capping protein CapZ on sodium channel regulation in leukemia K562 cells. The channels were activated by cytochalasin-induced disruption of actin filaments and inactivated by G-actin under ionic conditions promoting rapid actin polymerization. CapZ had no direct effect on channel activity. However, being added together with G-actin, CapZ prevented actin-induced channel inactivation, and this effect occurred at CapZ/actin molar ratios from 1:5 to 1:100. When actin was allowed to polymerize at the plasma membrane to induce partial channel inactivation, subsequent addition of CapZ restored the channel activity. These results can be explained by CapZ-induced inhibition of further assembly of actin filaments at the plasma membrane due to the modification of actin dynamics by CapZ. No effect on the channel activity was observed in response to F-actin, confirming that the mechanism of channel inactivation does not involve interaction of the channel with preformed filaments. Our data show that actin-capping protein can participate in the cytoskeleton-associated regulation of sodium transport in nonexcitable cells.

scholarly journals Actin-capping protein promotes microtubule stability by antagonizing the actin activity of mDia1

2012 ◽  
Vol 23 (20) ◽  
pp. 4032-4040 ◽  
Author(s):  
Francesca Bartolini ◽  
Nagendran Ramalingam ◽  
Gregg G. Gundersen
Keyword(s):  
Actin Filaments ◽  
Actin Polymerization ◽  
Dependent Manner ◽  
Proliferating Cells ◽  
Capping Protein ◽  
Microtubule Stability ◽  
Selective Stabilization ◽  
Actin Capping Protein ◽  
Actin Capping ◽  
Interfering Rna

In migrating fibroblasts, RhoA and its effector mDia1 regulate the selective stabilization of microtubules (MTs) polarized in the direction of migration. The conserved formin homology 2 domain of mDia1 is involved both in actin polymerization and MT stabilization, and the relationship between these two activities is unknown. We found that latrunculin A (LatA) and jasplakinolide, actin drugs that release mDia1 from actin filament barbed ends, stimulated stable MT formation in serum-starved fibroblasts and caused a redistribution of mDia1 onto MTs. Knockdown of mDia1 by small interfering RNA (siRNA) prevented stable MT induction by LatA, whereas blocking upstream Rho or integrin signaling had no effect. In search of physiological regulators of mDia1, we found that actin-capping protein induced stable MTs in an mDia1-dependent manner and inhibited the translocation of mDia on the ends of growing actin filaments. Knockdown of capping protein by siRNA reduced stable MT levels in proliferating cells and in starved cells stimulated with lysophosphatidic acid. These results show that actin-capping protein is a novel regulator of MT stability that functions by antagonizing mDia1 activity toward actin filaments and suggest a novel form of actin–MT cross-talk in which a single factor acts sequentially on actin and MTs.

scholarly journals Yeast actin patches are networks of branched actin filaments

2004 ◽  
Vol 166 (5) ◽  
pp. 629-635 ◽  
Author(s):  
Michael E. Young ◽  
John A. Cooper ◽  
Paul C. Bridgman
Keyword(s):  
Actin Filaments ◽  
Actin Polymerization ◽  
Branch Ratio ◽  
Actin Assembly ◽  
Cortical Actin ◽  
Capping Protein ◽  
Negative Stain ◽  
Actin Structure ◽  
Negative Stain Electron Microscopy ◽  
Actin Patch

Cortical actin patches are the most prominent actin structure in budding and fission yeast. Patches assemble, move, and disassemble rapidly. We investigated the mechanisms underlying patch actin assembly and motility by studying actin filament ultrastructure within a patch. Actin patches were partially purified from Saccharomyces cerevisiae and examined by negative-stain electron microscopy (EM). To identify patches in the EM, we correlated fluorescence and EM images of GFP-labeled patches. Patches contained a network of actin filaments with branches characteristic of Arp2/3 complex. An average patch contained 85 filaments. The average filament was only 50-nm (20 actin subunits) long, and the filament to branch ratio was 3:1. Patches lacking Sac6/fimbrin were unstable, and patches lacking capping protein were relatively normal. Our results are consistent with Arp2/3 complex-mediated actin polymerization driving yeast actin patch assembly and motility, as described by a variation of the dendritic nucleation model.

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