scholarly journals The tension mounts: Stress fibers as force-generating mechanotransducers

2013 ◽  
Vol 200 (1) ◽  
pp. 9-19 ◽  
Author(s):  
Keith Burridge ◽  
Erika S. Wittchen
Keyword(s):  
Tissue Culture ◽  
Physical Environment ◽  
Actin Filaments ◽  
Striated Muscle ◽  
Myosin Ii ◽  
Stress Fibers ◽  
New Work ◽  
Mechanical Tension ◽  
And Behavior ◽  
In Cells

Stress fibers (SFs) are often the most prominent cytoskeletal structures in cells growing in tissue culture. Composed of actin filaments, myosin II, and many other proteins, SFs are force-generating and tension-bearing structures that respond to the surrounding physical environment. New work is shedding light on the mechanosensitive properties of SFs, including that these structures can respond to mechanical tension by rapid reinforcement and that there are mechanisms to repair strain-induced damage. Although SFs are superficially similar in organization to the sarcomeres of striated muscle, there are intriguing differences in their organization and behavior, indicating that much still needs to be learned about these structures.

scholarly journals Specification of Actin Filament Function and Molecular Composition by Tropomyosin Isoforms

2003 ◽  
Vol 14 (3) ◽  
pp. 1002-1016 ◽  
Author(s):  
Nicole S. Bryce ◽  
Galina Schevzov ◽  
Vicki Ferguson ◽  
Justin M. Percival ◽  
Jim J.-C. Lin ◽  
...  
Keyword(s):  
Cell Size ◽  
Functional Properties ◽  
Actin Filament ◽  
Actin Filaments ◽  
Myosin Ii ◽  
Molecular Composition ◽  
Cellular Migration ◽  
Stress Fibers ◽  
Human Homologue ◽  
Tropomyosin Isoforms

The specific functions of greater than 40 vertebrate nonmuscle tropomyosins (Tms) are poorly understood. In this article we have tested the ability of two Tm isoforms, TmBr3 and the human homologue of Tm5 (hTM5NM1), to regulate actin filament function. We found that these Tms can differentially alter actin filament organization, cell size, and shape. hTm5NM1was able to recruit myosin II into stress fibers, which resulted in decreased lamellipodia and cellular migration. In contrast, TmBr3 transfection induced lamellipodial formation, increased cellular migration, and reduced stress fibers. Based on coimmunoprecipitation and colocalization studies, TmBr3 appeared to be associated with actin-depolymerizing factor/cofilin (ADF)-bound actin filaments. Additionally, the Tms can specifically regulate the incorporation of other Tms into actin filaments, suggesting that selective dimerization may also be involved in the control of actin filament organization. We conclude that Tm isoforms can be used to specify the functional properties and molecular composition of actin filaments and that spatial segregation of isoforms may lead to localized specialization of actin filament function.

scholarly journals Crawling from soft to stiff matrix polarizes the cytoskeleton and phosphoregulates myosin-II heavy chain

2012 ◽  
Vol 199 (4) ◽  
pp. 669-683 ◽  
Author(s):  
Matthew Raab ◽  
Joe Swift ◽  
P.C. Dave P. Dingal ◽  
Palak Shah ◽  
Jae-Won Shin ◽  
...  
Keyword(s):  
Stem Cells ◽  
Myosin Ii ◽  
Stress Fibers ◽  
Site Specific ◽  
Soft Matrix ◽  
3D Collagen ◽  
The Difference ◽  
Rigid Surfaces ◽  
Tissue Matrix ◽  
In Cells

On rigid surfaces, the cytoskeleton of migrating cells is polarized, but tissue matrix is normally soft. We show that nonmuscle MIIB (myosin-IIB) is unpolarized in cells on soft matrix in 2D and also within soft 3D collagen, with rearward polarization of MIIB emerging only as cells migrate from soft to stiff matrix. Durotaxis is the tendency of cells to crawl from soft to stiff matrix, and durotaxis of primary mesenchymal stem cells (MSCs) proved more sensitive to MIIB than to the more abundant and persistently unpolarized nonmuscle MIIA (myosin-IIA). However, MIIA has a key upstream role: in cells on soft matrix, MIIA appeared diffuse and mobile, whereas on stiff matrix, MIIA was strongly assembled in oriented stress fibers that MIIB then polarized. The difference was caused in part by elevated phospho-S1943–MIIA in MSCs on soft matrix, with site-specific mutants revealing the importance of phosphomoderated assembly of MIIA. Polarization is thus shown to be a highly regulated compass for mechanosensitive migration.

scholarly journals Different contributions of nonmuscle myosin IIA and IIB to the organization of stress fiber subtypes in fibroblasts

2018 ◽  
Vol 29 (8) ◽  
pp. 911-922 ◽  
Author(s):  
Masahiro Kuragano ◽  
Taro Q. P. Uyeda ◽  
Keiju Kamijo ◽  
Yota Murakami ◽  
Masayuki Takahashi
Keyword(s):  
Actin Filaments ◽  
Myosin Ii ◽  
Contractile Force ◽  
Stress Fiber ◽  
Stress Fibers ◽  
Proper Function ◽  
Nonmuscle Myosin ◽  
Nonmuscle Myosin Ii ◽  
Exogenous Expression ◽  
Main Component

Stress fibers (SFs) are contractile, force-generating bundled structures that can be classified into three subtypes, namely ventral SFs (vSFs), transverse arcs (TAs), and dorsal SFs. Nonmuscle myosin II (NMII) is the main component of SFs. This study examined the roles of the NMII isoforms NMIIA and NMIIB in the organization of each SF subtype in immortalized fibroblasts. Knockdown (KD) of NMIIA (a major isoform) resulted in loss of TAs from the lamella and caused the lamella to lose its flattened shape. Exogenous expression of NMIIB rescued this defect in TA formation. However, the TAs that formed on exogenous NMIIB expression in NMIIA-KD cells and the remaining TAs in NMIIB-KD cells, which mainly consisted of NMIIB and NMIIA, respectively, failed to rescue the defect in lamellar flattening. These results indicate that both isoforms are required for the proper function of TAs in lamellar flattening. KD of NMIIB resulted in loss of vSFs from the central region of the cell body, and this defect was not rescued by exogenous expression of NMIIA, indicating that NMIIA cannot replace the function of NMIIB in vSF formation. Moreover, we raised the possibility that actin filaments in vSFs are in a stretched conformation.

scholarly journals Plectin sidearms mediate interaction of intermediate filaments with microtubules and other components of the cytoskeleton.

1996 ◽  
Vol 135 (4) ◽  
pp. 991-1007 ◽  
Author(s):  
T M Svitkina ◽  
A B Verkhovsky ◽  
G G Borisy
Keyword(s):  
Transgenic Mice ◽  
Mammalian Cell ◽  
Intermediate Filaments ◽  
Myosin Ii ◽  
Immunogold Labeling ◽  
Stress Fibers ◽  
Cross Links ◽  
Membrane Components ◽  
In Cells

By immunogold labeling, we demonstrate that "millipede-like" structures seen previously in mammalian cell cytoskeletons after removal of actin by treatment with gelsolin are composed of the cores of vimentin IFs with sidearms containing plectin. These plectin sidearms connect IFs to microtubules, the actin-based cytoskeleton and possibly membrane components. Plectin binding to microtubules was significantly increased in cells from transgenic mice lacking IFs and was reversed by microinjection of exogenous vimentin. These results suggest the existence of a pool of plectin which preferentially associates with IFs but may also be competed for by microtubules. The association of IFs with microtubules did not show a preference for Glu-tubulin. Nor did it depend upon the presence of MAP4 since plectin links were retained after specific immunodepletion of MAP4. The association of IFs with stress fibers survived actin depletion by gelsolin suggesting that myosin II minifilaments or components closely associated with them may play a role as plectin targets. Our results provide direct structural evidence for the hypothesis that plectin cross-links elements of the cytoskeleton thus leading to integration of the cytoplasm.

scholarly journals Retrograde Flow and Myosin II Activity within the Leading Cell Edge Deliver F-Actin to the Lamella to Seed the Formation of Graded Polarity Actomyosin II Filament Bundles in Migrating Fibroblasts

2008 ◽  
Vol 19 (11) ◽  
pp. 5006-5018 ◽  
Author(s):  
Tom W. Anderson ◽  
Andrew N. Vaughan ◽  
Louise P. Cramer
Keyword(s):  
Actin Filaments ◽  
Myosin Ii ◽  
Stress Fibers ◽  
Live Cells ◽  
Retrograde Flow ◽  
Mechanism Of Formation ◽  
Unknown Mechanism ◽  
Cell Edge ◽  
Cell Protrusion ◽  
Filament Bundles

In migrating fibroblasts actomyosin II bundles are graded polarity (GP) bundles, a distinct organization to stress fibers. GP bundles are important for powering cell migration, yet have an unknown mechanism of formation. Electron microscopy and the fate of photobleached marks show actin filaments undergoing retrograde flow in filopodia, and the lamellipodium are structurally and dynamically linked with stationary GP bundles within the lamella. An individual filopodium initially protrudes, but then becomes separated from the tip of the lamellipodium and seeds the formation of a new GP bundle within the lamella. In individual live cells expressing both GFP-myosin II and RFP-actin, myosin II puncta localize to the base of an individual filopodium an average 28 s before the filopodium seeds the formation of a new GP bundle. Associated myosin II is stationary with respect to the substratum in new GP bundles. Inhibition of myosin II motor activity in live cells blocks appearance of new GP bundles in the lamella, without inhibition of cell protrusion in the same timescale. We conclude retrograde F-actin flow and myosin II activity within the leading cell edge delivers F-actin to the lamella to seed the formation of new GP bundles.

scholarly journals A novel cytoskeletal structure involved in purse string wound closure and cell polarity maintenance.

1993 ◽  
Vol 121 (3) ◽  
pp. 565-578 ◽  
Author(s):  
W M Bement ◽  
P Forscher ◽  
M S Mooseker
Keyword(s):  
Actin Filaments ◽  
Laser Scanning ◽  
Wound Repair ◽  
Immunofluorescence Microscopy ◽  
Myosin Ii ◽  
Epithelial Cell Line ◽  
Purse String ◽  
Cytoskeletal Structure ◽  
Junction Protein ◽  
In Cells

The process of wound repair in monolayers of the intestinal epithelial cell line, Caco-2BBe, was analyzed by a combination of time-lapse differential interference contrast (DIC) video and immunofluorescence microscopy, and laser scanning confocal immunofluorescence microscopy (LSCIM). DIC video analysis revealed that stab wounds made in Caco-2BBe monolayers healed by two distinct processes: (a) Extension of lamellipodia into the wounds; and (b) Purse string closure of the wound by distinct arcs or rings formed by cells bordering the wound. The arcs and rings which effected purse string closure appeared sharp and sheer in DIC, spanned between two and eight individual cells along the wound border, and contracted in a concerted fashion. Immunofluorescence analysis of the wounds demonstrated that the arcs and rings contained striking accumulations of actin filaments, myosin-II, villin, and tropomyosin. In contrast, arcs and rings contained no apparent enrichment of microtubules, brush border myosin-I immunogens, or myosin-V. LSCIM analysis confirmed the localization of actin filaments, myosin-II, villin, and tropomyosin in arcs and rings at wound borders. ZO-1 (a tight junction protein), also accumulated in arcs and rings around wounds, despite the fact that cell-cell contacts are absent at wound borders. Sucrase-isomaltase, an apically-localized integral membrane protein, maintained an apical localization in cells where arcs or rings were formed, but was found in lamellipodia extending into wounds in cells where arcs failed to form. Time-course, LSCIM quantification of actin, myosin II, and ZO-1 revealed that accumulation of these proteins within arcs and rings at the wound edge began within 5 minutes and peaked within 30-60 minutes of wounding. Actin filaments, myosin-II, and ZO-1 achieved 10-, 3-, and 4-fold enrichments, respectively, relative to cell edges which did not border wounds. The results demonstrate that wounded Caco-2BBe monolayers assemble a novel cytoskeletal structure at the borders of wounds. The results further suggest that this structure plays at least two roles in wound repair; first, mediation of concerted, purse string movement of cells into the area of the wound and second, maintenance of apical/basolateral polarity in cells which border the wound.

scholarly journals Muscle-specific stress fibers give rise to sarcomeres in cardiomyocytes

eLife ◽  
2018 ◽  
Vol 7 ◽  
Author(s):  
Aidan M Fenix ◽  
Abigail C Neininger ◽  
Nilay Taneja ◽  
Karren Hyde ◽  
Mike R Visetsouk ◽  
...  
Keyword(s):  
Actin Filaments ◽  
Myosin Ii ◽  
Stress Fibers ◽  
Myosin Filament ◽  
Cardiac Myosin ◽  
Human Cardiomyocytes ◽  
Filament Assembly ◽  
Muscle Stress ◽  
Sarcomere Assembly ◽  
Muscle Myosin

The sarcomere is the contractile unit within cardiomyocytes driving heart muscle contraction. We sought to test the mechanisms regulating actin and myosin filament assembly during sarcomere formation. Therefore, we developed an assay using human cardiomyocytes to monitor sarcomere assembly. We report a population of muscle stress fibers, similar to actin arcs in non-muscle cells, which are essential sarcomere precursors. We show sarcomeric actin filaments arise directly from muscle stress fibers. This requires formins (e.g., FHOD3), non-muscle myosin IIA and non-muscle myosin IIB. Furthermore, we show short cardiac myosin II filaments grow to form ~1.5 μm long filaments that then ‘stitch’ together to form the stack of filaments at the core of the sarcomere (i.e., the A-band). A-band assembly is dependent on the proper organization of actin filaments and, as such, is also dependent on FHOD3 and myosin IIB. We use this experimental paradigm to present evidence for a unifying model of sarcomere assembly.

scholarly journals 2TA4-09 Specific subsets of actin filaments in cells have a high affinity for S1 of myosin II.(The 47th Annual Meeting of the Biophysical Society of Japan)

2009 ◽  
Vol 49 (supplement) ◽  
pp. S44
Author(s):  
Taro Uyeda ◽  
Akira Nagasaki ◽  
Akira Ichihara ◽  
Takashi Hirano
Keyword(s):  
Annual Meeting ◽  
Actin Filaments ◽  
Myosin Ii ◽  
High Affinity ◽  
Biophysical Society ◽  
In Cells

Cytoplasmic Tubules in Cells Infected with Laryngotracheitis Virus

1969 ◽  
Vol 27 ◽  
pp. 376-377
Author(s):  
A. M. Watrach
Keyword(s):  
Tissue Culture ◽  
Small Proportion ◽  
Chicken Embryo ◽  
Culture Cells ◽  
Tissue Culture Cells ◽  
Morphologic Characteristics ◽  
Infectious Laryngotracheitis ◽  
Laryngotracheitis Virus ◽  
In Cells

During a study of the development of infectious laryngotracheitis (LT) virus in tissue culture cells, unusual tubular formations were found in the cytoplasm of a small proportion of the affected cells. It is the purpose of this report to describe the morphologic characteristics of the tubules and to discuss their possible association with the development of virus.The source and maintenance of the strain of LT virus have been described. Prior to this study, the virus was passed several times in chicken embryo kidney (CEK) tissue culture cells.

Orientation of actin filaments during motion in motility assay

1995 ◽  
Vol 53 ◽  
pp. 52-53
Author(s):  
J. Borejdo ◽  
S. Burlacu
Keyword(s):  
Actin Filaments ◽  
Thin Filament ◽  
Striated Muscle ◽  
Myosin Head ◽  
Classical Method ◽  
Polarization Of Fluorescence ◽  
Single Protein ◽  
Intracellular Organelles ◽  
Change Orientation ◽  
Active Entity

Polarization of fluorescence is a classical method to assess orientation or mobility of macromolecules. It has been a common practice to measure polarization of fluorescence through a microscope to characterize orientation or mobility of intracellular organelles, for example anisotropic bands in striated muscle. Recently, we have extended this technique to characterize single protein molecules. The scientific question concerned the current problem in muscle motility: whether myosin heads or actin filaments change orientation during contraction. The classical view is that the force-generating step in muscle is caused by change in orientation of myosin head (subfragment-1 or SI) relative to the axis of thin filament. The molecular impeller which causes this change resides at the interface between actin and SI, but it is not clear whether only the myosin head or both SI and actin change orientation during contraction. Most studies assume that observed orientational change in myosin head is a reflection of the fact that myosin is an active entity and actin serves merely as a passive "rail" on which myosin moves.

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