Intracellular Memory of Stress Fiber Orientation: Focal Adhesions Store It and Microtubules Erase It During Disassembly–Reassembly Process of Stress Fibers

2009 ◽  
pp. 649-650
Author(s):  
Y. F. Yang ◽  
K. Nagayama ◽  
T. Matsumoto
Keyword(s):  
Fiber Orientation ◽  
Focal Adhesions ◽  
Stress Fiber ◽  
Stress Fibers

On the Intracellullar Memory of Stress Fiber Orientation: Effects of Microtubules and Focal Adhesions on the Repolymerization Process of Stress Fibers

2008 ◽  
Author(s):  
Yunfeng Yang ◽  
Kazuaki Nagayama ◽  
Takeo Matsumoto
Keyword(s):  
Fiber Orientation ◽  
Cell Movement ◽  
Focal Adhesions ◽  
Stress Fiber ◽  
Cyclic Stretch ◽  
Stress Fibers ◽  
Cellular Functions ◽  
Division 1 ◽  
Key Features ◽  
Strain Magnitude

Stress fibers (SFs) play essential roles in various cellular functions such as cell movement, shape maintenance and cell division [1]. One of their key features is that they dynamically change their structures in response to mechanical environment to which they are exposed [2]. For example, cultured endothelial cells exposed to cyclic stretch preferentially reorganize their actin stress fibers to the direction in which the strain magnitude of the fibers become minimum [3].

scholarly journals Calcium-Sensing Receptor Stimulation in Cultured Glomerular Podocytes Induces TRPC6-Dependent Calcium Entry and RhoA Activation

2017 ◽  
Vol 43 (5) ◽  
pp. 1777-1789 ◽  
Author(s):  
Lei Zhang ◽  
Tianrong Ji ◽  
Qin Wang ◽  
Kexin Meng ◽  
Rui Zhang ◽  
...  
Keyword(s):  
Protective Action ◽  
Focal Adhesions ◽  
Stress Fiber ◽  
Fiber Formation ◽  
Stress Fibers ◽  
Dependent Manner ◽  
Calcium Sensing Receptor ◽  
Rhoa Activity ◽  
Rhoa Activation ◽  
Calcium Sensing

Background/Aims: Recent studies provided compelling evidence that stimulation of the calcium sensing receptor (CaSR) exerts direct renoprotective action at the glomerular podocyte level. This protective action may be attributed to the RhoA-dependent stabilization of the actin cytoskeleton. However, the underlying mechanisms remain unclear. Methods: In the present study, an immortalized human podocyte cell line was used. Fluo-3 fluorescence was utilized to determine intracellular Ca2+ concentration ([Ca2+]i), and western blotting was used to measure canonical transient receptor potential 6 (TRPC6) protein expression and RhoA activity. Stress fibers were detected by FITC-phalloidin. Results: Activating CaSR with a high extracellular Ca2+ concentration ([Ca2+]o) or R-568 (a type II CaSR agonist) induces an increase in the [Ca2+]i in a dose-dependent manner. This increase in [Ca2+]i is phospholipase C (PLC)-dependent and is smaller in the absence of extracellular Ca2+ than in the presence of 0.5 mM [Ca2+]o. The CaSR activation-induced [Ca2+]i increase is attenuated by the pharmacological blockage of TRPC6 channels or siRNA targeting TRPC6. These data suggest that TRPC6 is involved in CaSR activation-induced Ca2+ influx. Consistent with a previous study, CaSR stimulation results in an increase in RhoA activity. However, the knockdown of TRPC6 significantly abolished the RhoA activity increase induced by CaSR stimulation, suggesting that TRPC6-dependent Ca2+ entry is required for RhoA activation. The activated RhoA is involved in the formation of stress fibers and focal adhesions in response to CaSR stimulation because siRNA targeting RhoA attenuated the increase in the stress fiber mediated by CaSR stimulation. Moreover, this effect of CaSR activation on the formation of stress fibers is also abolished by the knockdown of TRPC6. Conclusion: TRPC6 is involved in the regulation of stress fiber formation and focal adhesions via the RhoA pathway in response to CaSR activation. This may explain the direct protective action of CaSR agonists.

Indomethacin Delays Gastric Restitution: Association with the Inhibition of Focal Adhesion Kinase and Tensin Phosphorylation and Reduced Actin Stress Fibers

2002 ◽  
Vol 227 (6) ◽  
pp. 412-424 ◽  
Author(s):  
Imre L. Szabó ◽  
Rama Pai ◽  
Michael K. Jones ◽  
George R. Ehring ◽  
Hirofumi Kawanaka ◽  
...  
Keyword(s):  
Cell Migration ◽  
Focal Adhesion ◽  
Focal Adhesions ◽  
Stress Fiber ◽  
Fiber Formation ◽  
Stress Fibers ◽  
Actin Stress Fiber ◽  
Stress Fiber Formation ◽  
Actin Stress Fiber Formation

Repair of superficial gastric mucosal injury is accomplished by the process of restitution—migration of epithelial cells to restore continuity of the mucosal surface. Actin filaments, focal adhesions, and focal adhesion kinase (FAK) play crucial roles in cell motility essential for restitution. We studied whether epidermal growth factor (EGF) and/or indomethacin (IND) affect cell migration, actin stress fiber formation, and/or phosphorylation of FAK and tensin in wounded gastric monolayers. Human gastric epithelial monolayers (MKN 28 cells) were wounded and treated with either vehicle or 0.5 mM IND for 16 hr followed by EGF. EGF treatment significantly stimulated cell migration and actin stress fiber formation, and increased FAK localization to focal adhesions, and phosphorylation of FAK and tensin, whereas IND inhibited all these at the baseline and EGF-stimulated conditions. IND-induced inhibition of FAK phosphorylation preceded changes in actin polymerization, indicating that actin depolymerization might be the consequence of decreased FAK activity. In in vivo experiments, rats received either vehicle or IND (5 mg/kg i.g.), and 3 min later, they received water or 5% hypertonic NaCl; gastric mucosa was obtained at 1, 4, and 8 hr after injury. Four and 8 hr after hypertonic injury, FAK phosphorylation was induced in gastric mucosa compared with controls. IND pretreatment significantly delayed epithelial restitution in vivo, and reduced FAK phosphorylation and recruitment to adhesion points, as well as actin stress fiber formation in migrating surface epithelial cells. Our study indicates that FAK, tensin, and actin stress fibers are likely mediators of EGF-stimulated cell migration in wounded human gastric monolayers and potential targets for IND-induced inhibition of restitution.

Stress fiber organization regulated by MLCK and Rho-kinase in cultured human fibroblasts

2001 ◽  
Vol 280 (6) ◽  
pp. C1669-C1679 ◽  
Author(s):  
Kazuo Katoh ◽  
Yumiko Kano ◽  
Mutsuki Amano ◽  
Kozo Kaibuchi ◽  
Keigi Fujiwara
Keyword(s):  
Kinase Inhibitors ◽  
Fluorescent Protein ◽  
Human Fibroblasts ◽  
Rho Kinase ◽  
Focal Adhesions ◽  
Stress Fiber ◽  
Stress Fibers ◽  
Fiber System ◽  
Rho Kinase Inhibitors ◽  
Calmodulin Inhibitors

To understand the roles of Rho-kinase and myosin light chain kinase (MLCK) for the contraction and organization of stress fibers, we treated cultured human foreskin fibroblasts with several MLCK, Rho-kinase, or calmodulin inhibitors and analyzed F-actin organization in the cells. Some cells were transfected with green fluorescent protein (GFP)-labeled actin, and the effects of inhibitors were also studied in these living cells. The Rho-kinase inhibitors Y-27632 and HA1077 caused disassembly of stress fibers and focal adhesions in the central portion of the cell within 1 h. However, stress fibers located in the periphery of the cell were not severely affected by the Rho-kinase inhibitors. When these cells were washed with fresh medium, the central stress fibers and focal adhesions gradually reformed, and within 3 h the cells were completely recovered. ML-7 and KT5926 are specific MLCK inhibitors and caused disruption and/or shortening of peripheral stress fibers, leaving the central fibers relatively intact even though their number was reduced. The calmodulin inhibitors W-5 and W-7 gave essentially the same results as the MLCK inhibitors. The MLCK and calmodulin inhibitors, but not the Rho-kinase inhibitors, caused cells to lose the spread morphology, indicating that the peripheral fibers play a major role in keeping the flattened state of the cell. When stress fiber models were reactivated, the peripheral fibers contracted before the central fibers. Thus our study shows that there are at least two different stress fiber systems in the cell. The central stress fiber system is dependent more on the activity of Rho-kinase than on that of MLCK, while the peripheral stress fiber system depends on MLCK.

scholarly journals LIMCH1 regulates nonmuscle myosin-II activity and suppresses cell migration

2017 ◽  
Vol 28 (8) ◽  
pp. 1054-1065 ◽  
Author(s):  
Yu-Hung Lin ◽  
Yen-Yi Zhen ◽  
Kun-Yi Chien ◽  
I-Ching Lee ◽  
Wei-Chi Lin ◽  
...  
Keyword(s):  
Cell Migration ◽  
Hela Cells ◽  
Myosin Ii ◽  
Focal Adhesions ◽  
Stress Fiber ◽  
Stress Fibers ◽  
Actin Stress Fiber ◽  
Nonmuscle Myosin ◽  
Head Region ◽  
Nonmuscle Myosin Ii

Nonmuscle myosin II (NM-II) is an important motor protein involved in cell migration. Incorporation of NM-II into actin stress fiber provides a traction force to promote actin retrograde flow and focal adhesion assembly. However, the components involved in regulation of NM-II activity are not well understood. Here we identified a novel actin stress fiber–associated protein, LIM and calponin-homology domains 1 (LIMCH1), which regulates NM-II activity. The recruitment of LIMCH1 into contractile stress fibers revealed its localization complementary to actinin-1. LIMCH1 interacted with NM-IIA, but not NM-IIB, independent of the inhibition of myosin ATPase activity with blebbistatin. Moreover, the N-terminus of LIMCH1 binds to the head region of NM-IIA. Depletion of LIMCH1 attenuated myosin regulatory light chain (MRLC) diphosphorylation in HeLa cells, which was restored by reexpression of small interfering RNA–resistant LIMCH1. In addition, LIMCH1-depleted HeLa cells exhibited a decrease in the number of actin stress fibers and focal adhesions, leading to enhanced cell migration. Collectively, our data suggest that LIMCH1 plays a positive role in regulation of NM-II activity through effects on MRLC during cell migration.

scholarly journals Force-dependent activation of actin elongation factor mDia1 protects the cytoskeleton from mechanical damage and facilitates stress fiber repair

2021 ◽  
Author(s):  
Fernando R. Valencia ◽  
Eduardo Sandoval ◽  
Jian Liu ◽  
Sergey V. Plotnikov
Keyword(s):  
Cell Mechanics ◽  
Actin Polymerization ◽  
Safety Valve ◽  
Mechanical Damage ◽  
Elongation Factor ◽  
Focal Adhesions ◽  
Stress Fiber ◽  
Stress Fibers ◽  
Mechanical Tension ◽  
Contractile Forces

ABSTRACTPlasticity of cell mechanics, which relies heavily on the spatiotemporal regulation of the actomyosin cytoskeleton, safeguards cells against mechanical damage. Yet, mechanisms of adaptive change in cell mechanics remain elusive. Here, we report a new mechanism whereby mechanically activated actin elongation factor mDia1 controls the dynamics of actin polymerization at focal adhesions, force bearing linkages between the actin cytoskeleton and extracellular matrix. By combining live-cell imaging with mathematical modelling, we show that actin polymerization at focal adhesions exhibits pulsatile dynamics where the spikes of mDia1 activity are triggered by cell-generated contractile forces. We show that suppression of mDia1-mediated actin polymerization at focal adhesions results in two-fold increase in mechanical tension on the stress fibers. This elevated tension leads to an increased frequency of spontaneous stress fiber damage and decreased efficiency of zyxin-mediated stress fiber repair. We conclude that tension-controlled actin polymerization at focal adhesions acts as a safety valve dampening excessive mechanical tension on the actin cytoskeleton and safeguarding stress fibers against mechanical damage.SUMMARYValencia et al. reports that stress fiber elongation at focal adhesion requires mDia1 activity, furthermore contractile forces trigger mDia1-dependent actin polymerization. mDia1-mediated actin polymerization acts as a safety valve to dampen mechanical stress and protect the cell from damage.

Actin stress fiber dynamics in laterally confined cells

2019 ◽  
Vol 11 (5) ◽  
pp. 175-185 ◽  
Author(s):  
Andreas Müller ◽  
Sandra Müller ◽  
Veselin Nasufovic ◽  
Hans-Dieter Arndt ◽  
Tilo Pompe
Keyword(s):  
Fiber Orientation ◽  
Stress Fiber ◽  
Stress Fibers ◽  
Time Interval ◽  
Actin Stress Fiber ◽  
Micro Patterning ◽  
Actin Fiber ◽  
Depth Analysis ◽  
Fiber Dynamics ◽  
Actin Stress Fibers

Abstract Multiple cellular processes are affected by spatial constraints from the extracellular matrix and neighboring cells. In vitro experiments using defined micro-patterning allow for in-depth analysis and a better understanding of how these constraints impact cellular behavior and functioning. Herein we focused on the analysis of actin cytoskeleton dynamics as a major determinant of mechanotransduction mechanisms in cells. We seeded primary human umbilical vein endothelial cells onto stripe-like cell-adhesive micro-patterns with varying widths and then monitored and quantified the dynamic reorganization of actin stress fibers, including fiber velocities, orientation and density, within these live cells using the cell permeable F-actin marker SiR-actin. Although characteristic parameters describing the overall stress fiber architecture (average orientation and density) were nearly constant throughout the observation time interval of 60 min, we observed permanent transport and turnover of individual actin stress fibers. Stress fibers were more strongly oriented along stripe direction with decreasing stripe width, (5° on 20 μm patterns and 10° on 40 μm patterns), together with an overall narrowing of the distribution of fiber orientation. Fiber dynamics was characterized by a directed movement from the cell edges towards the cell center, where fiber dissolution frequently took place. By kymograph analysis, we found median fiber velocities in the range of 0.2 μm/min with a weak dependence on pattern width. Taken together, these data suggest that cell geometry determines actin fiber orientation, while it also affects actin fiber transport and turnover.

scholarly journals Septins promote stress fiber–mediated maturation of focal adhesions and renal epithelial motility

2014 ◽  
Vol 207 (2) ◽  
pp. 225-235 ◽  
Author(s):  
Lee Dolat ◽  
John L. Hunyara ◽  
Jonathan R. Bowen ◽  
Eva Pauline Karasmanis ◽  
Maha Elgawly ◽  
...  
Keyword(s):  
G Proteins ◽  
Tumor Metastasis ◽  
Focal Adhesions ◽  
Stress Fiber ◽  
Stress Fibers ◽  
Actin Network ◽  
Septin 9 ◽  
The Stability ◽  
2D And 3D ◽  
Hepatocyte Growth

Organogenesis and tumor metastasis involve the transformation of epithelia to highly motile mesenchymal-like cells. Septins are filamentous G proteins, which are overexpressed in metastatic carcinomas, but their functions in epithelial motility are unknown. Here, we show that a novel network of septin filaments underlies the organization of the transverse arc and radial (dorsal) stress fibers at the leading lamella of migrating renal epithelia. Surprisingly, septin depletion resulted in smaller and more transient and peripheral focal adhesions. This phenotype was accompanied by a highly disorganized lamellar actin network and rescued by the actin bundling protein α-actinin-1. We show that preassembled actin filaments are cross-linked directly by Septin 9 (SEPT9), whose expression is increased after induction of renal epithelial motility with the hepatocyte growth factor. Significantly, SEPT9 overexpression enhanced renal cell migration in 2D and 3D matrices, whereas SEPT9 knockdown decreased migration. These results suggest that septins promote epithelial motility by reinforcing the cross-linking of lamellar stress fibers and the stability of nascent focal adhesions.

scholarly journals ARF1 Mediates Paxillin Recruitment to Focal Adhesions and Potentiates Rho-stimulated Stress Fiber Formation in Intact and Permeabilized Swiss 3T3 Fibroblasts

1998 ◽  
Vol 143 (7) ◽  
pp. 1981-1995 ◽  
Author(s):  
J.C. Norman ◽  
D. Jones ◽  
S.T. Barry ◽  
M.R. Holt ◽  
S. Cockcroft ◽  
...  
Keyword(s):  
Focal Adhesion ◽  
Focal Adhesions ◽  
Stress Fiber ◽  
Fiber Formation ◽  
Stress Fibers ◽  
3T3 Fibroblasts ◽  
Stress Fiber Formation ◽  
Swiss 3T3 ◽  
Swiss 3T3 Fibroblasts ◽  
Actin Stress Fibers

Focal adhesion assembly and actin stress fiber formation were studied in serum-starved Swiss 3T3 fibroblasts permeabilized with streptolysin-O. Permeabilization in the presence of GTPγS stimulated rho-dependent formation of stress fibers, and the redistribution of vinculin and paxillin from a perinuclear location to focal adhesions. Addition of GTPγS at 8 min after permeabilization still induced paxillin recruitment to focal adhesion–like structures at the ends of stress fibers, but vinculin remained in the perinuclear region, indicating that the distributions of these two proteins are regulated by different mechanisms. Paxillin recruitment was largely rho-independent, but could be evoked using constitutively active Q71L ADP-ribosylation factor (ARF1), and blocked by NH2-terminally truncated Δ17ARF1. Moreover, leakage of endogenous ARF from cells was coincident with loss of GTPγS- induced redistribution of paxillin to focal adhesions, and the response was recovered by addition of ARF1. The ability of ARF1 to regulate paxillin recruitment to focal adhesions was confirmed by microinjection of Q71LARF1 and Δ17ARF1 into intact cells. Interestingly, these experiments showed that V14RhoA- induced assembly of actin stress fibers was potentiated by Q71LARF1. We conclude that rho and ARF1 activate complimentary pathways that together lead to the formation of paxillin-rich focal adhesions at the ends of prominent actin stress fibers.

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