scholarly journals Focal adhesion kinase signaling is decreased in polyamine-depleted IEC-6 cells

2001 ◽  
Vol 281 (2) ◽  
pp. C475-C485 ◽  
Author(s):  
Ramesh M. Ray ◽  
Mary Jane Viar ◽  
Shirley A. McCormack ◽  
Leonard R. Johnson
Keyword(s):  
Extracellular Matrix ◽  
Focal Adhesion Kinase ◽  
Focal Adhesion ◽  
Focal Adhesions ◽  
Stress Fiber ◽  
Fiber Formation ◽  
Integrin Signaling ◽  
Actin Stress Fiber ◽  
Stress Fiber Formation ◽  
And Migration

Polyamines are essential to the migration of epithelial cells in the intestinal mucosa. Cells depleted of polyamines do not attach as rapidly to the extracellular matrix and do not form the actin stress fibers essential for migration. Because both attachment and stress fiber formation depend on integrin signaling and the formation of focal adhesions, we examined these and related processes in polyamine-depleted IEC-6 cells. There was general decreased tyrosine phosphorylation of focal adhesion kinase (FAK), and, specifically, decreased phosphorylation of Tyr-925, the paxillin binding site. In control cells, FAK phosphorylation was rapid after attachment to the extracellular matrix, while attached cells depleted of polyamines had significantly delayed phosphorylation. FAK activity was also significantly inhibited in polyamine-depleted cells as was the phosphorylation of paxillin. Polyamine-depleted cells failed to spread normally after attachment, and immunocytochemistry showed little colocalization of FAK and actin compared with controls. Focal adhesion complex formation was greatly reduced in the absence of polyamines. These data suggest that defective integrin signaling may, at least in part, account for the decreased rates of attachment, actin stress fiber formation, spreading, and migration observed in polyamine-depleted cells.

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.

scholarly journals B-RAF Regulation of Rnd3 Participates in Actin Cytoskeletal and Focal Adhesion Organization

2008 ◽  
Vol 19 (2) ◽  
pp. 498-508 ◽  
Author(s):  
R. Matthew Klein ◽  
Laurie S. Spofford ◽  
Ethan V. Abel ◽  
Arisa Ortiz ◽  
Andrew E. Aplin
Keyword(s):  
Actin Cytoskeleton ◽  
Focal Adhesion ◽  
Constitutive Expression ◽  
Focal Adhesions ◽  
Stress Fiber ◽  
Fiber Formation ◽  
Mitogen Activated Protein Kinase ◽  
Actin Stress Fiber ◽  
Lim Kinase ◽  
Stress Fiber Formation

The actin cytoskeleton controls multiple cellular functions, including cell morphology, movement, and growth. Accumulating evidence indicates that oncogenic activation of the mitogen-activated protein kinase kinase/extracellular signal-regulated kinase 1/2 (MEK/ERK1/2) pathway is accompanied by actin cytoskeletal reorganization. However, the signaling events contributing to actin cytoskeleton remodeling mediated by aberrant ERK1/2 activation are largely unknown. Mutant B-RAF is found in a variety of cancers, including melanoma, and it enhances activation of the MEK/ERK1/2 pathway. We show that targeted knockdown of B-RAF with small interfering RNA or pharmacological inhibition of MEK increased actin stress fiber formation and stabilized focal adhesion dynamics in human melanoma cells. These effects were due to stimulation of the Rho/Rho kinase (ROCK)/LIM kinase-2 signaling pathway, cumulating in the inactivation of the actin depolymerizing/severing protein cofilin. The expression of Rnd3, a Rho antagonist, was attenuated after B-RAF knockdown or MEK inhibition, but it was enhanced in melanocytes expressing active B-RAF. Constitutive expression of Rnd3 suppressed the actin cytoskeletal and focal adhesion effects mediated by B-RAF knockdown. Depletion of Rnd3 elevated cofilin phosphorylation and stress fiber formation and reduced cell invasion. Together, our results identify Rnd3 as a regulator of cross talk between the RAF/MEK/ERK and Rho/ROCK signaling pathways, and a key contributor to oncogene-mediated reorganization of the actin cytoskeleton and focal adhesions.

GEF-H1 is involved in agonist-induced human pulmonary endothelial barrier dysfunction

2006 ◽  
Vol 290 (3) ◽  
pp. L540-L548 ◽  
Author(s):  
Anna A. Birukova ◽  
Djanybek Adyshev ◽  
Boris Gorshkov ◽  
Gary M. Bokoch ◽  
Konstantin G. Birukov ◽  
...  
Keyword(s):  
Critical Role ◽  
Focal Adhesions ◽  
Stress Fiber ◽  
Small Gtpase ◽  
Fiber Formation ◽  
Cell Contact ◽  
Actin Stress Fiber ◽  
Barrier Dysfunction ◽  
Actin Remodeling ◽  
Stress Fiber Formation

Endothelial cell (EC) permeability is precisely controlled by cytoskeletal elements [actin filaments, microtubules (MT), intermediate filaments] and cell contact protein complexes (focal adhesions, adherens junctions, tight junctions). We have recently shown that the edemagenic agonist thrombin caused partial MT disassembly, which was linked to activation of small GTPase Rho, Rho-mediated actin remodeling, cell contraction, and dysfunction of lung EC barrier. GEF-H1 is an MT-associated Rho-specific guanosine nucleotide (GDP/GTP) exchange factor, which in MT-unbound state stimulates Rho activity. In this study we tested hypothesis that GEF-H1 may be a key molecule involved in Rho activation, myosin light chain phosphorylation, actin remodeling, and EC barrier dysfunction associated with partial MT disassembly. Our results show that depletion of GEF-H1 or expression of dominant negative GEF-H1 mutant significantly attenuated permeability increase, actin stress fiber formation, and increased MLC and MYPT1 phosphorylation induced by thrombin or MT-depolymerizing agent nocodazole. In contrast, expression of wild-type or activated GEF-H1 mutants dramatically enhanced thrombin and nocodazole effects on stress fiber formation and cell retraction. These results show a critical role for the GEF-H1 in the Rho activation caused by MT disassembly and suggest GEF-H1 as a key molecule involved in cross talk between MT and actin cytoskeleton in agonist-induced Rho-dependent EC barrier regulation.

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.

scholarly journals Neuronal migration is mediated by inositol hexakisphosphate kinase 1 via α-actinin and focal adhesion kinase

2017 ◽  
Vol 114 (8) ◽  
pp. 2036-2041 ◽  
Author(s):  
Chenglai Fu ◽  
Jing Xu ◽  
Weiwei Cheng ◽  
Tomas Rojas ◽  
Alfred C. Chin ◽  
...  
Keyword(s):  
Focal Adhesion Kinase ◽  
Focal Adhesion ◽  
Neuronal Migration ◽  
Intracellular Localization ◽  
Focal Adhesions ◽  
Fiber Formation ◽  
Inositol Hexakisphosphate ◽  
Stress Fiber Formation ◽  
And Function ◽  
Pharmacologic Inhibition

Inositol hexakisphosphate kinase 1 (IP6K1), which generates 5-diphosphoinositol pentakisphosphate (5-IP7), physiologically mediates numerous functions. We report thatIP6K1deletion leads to brain malformation and abnormalities of neuronal migration. IP6K1 physiologically associates with α-actinin and localizes to focal adhesions.IP6K1deletion disrupts α-actinin’s intracellular localization and function. TheIP6K1deleted cells display substantial decreases of stress fiber formation and impaired cell migration and spreading. Regulation of α-actinin by IP6K1 requires its kinase activity. Deletion ofIP6K1abolishes α-actinin tyrosine phosphorylation, which is known to be regulated by focal adhesion kinase (FAK). FAK phosphorylation is substantially decreased inIP6K1deleted cells. 5-IP7, a product of IP6K1, promotes FAK autophosphorylation. Pharmacologic inhibition of IP6K by TNP [N2-(m-Trifluorobenzyl), N6-(p-nitrobenzyl)purine] recapitulates the phenotype ofIP6K1deletion. These findings establish that IP6K1 physiologically regulates neuronal migration by binding to α-actinin and influencing phosphorylation of both FAK and α-actinin through its product 5-IP7.

scholarly journals Genetic ablation of zyxin causes Mena/VASP mislocalization, increased motility, and deficits in actin remodeling

2006 ◽  
Vol 172 (5) ◽  
pp. 771-782 ◽  
Author(s):  
Laura M. Hoffman ◽  
Christopher C. Jensen ◽  
Susanne Kloeker ◽  
C.-L. Albert Wang ◽  
Masaaki Yoshigi ◽  
...  
Keyword(s):  
Extracellular Matrix ◽  
Focal Adhesions ◽  
Stress Fiber ◽  
Fiber Formation ◽  
Actin Stress Fiber ◽  
Adhesion Protein ◽  
Genetic Ablation ◽  
Associated Proteins ◽  
Dependent Cell ◽  
Focal Adhesion Protein

Focal adhesions are specialized regions of the cell surface where integrin receptors and associated proteins link the extracellular matrix to the actin cytoskeleton. To define the cellular role of the focal adhesion protein zyxin, we characterized the phenotype of fibroblasts in which the zyxin gene was deleted by homologous recombination. Zyxin-null fibroblasts display enhanced integrin-dependent adhesion and are more migratory than wild-type fibroblasts, displaying reduced dependence on extracellular matrix cues. We identified differences in the profiles of 75- and 80-kD tyrosine-phosphorylated proteins in the zyxin-null cells. Tandem array mass spectrometry identified both modified proteins as isoforms of the actomyosin regulator caldesmon, a protein known to influence contractility, stress fiber formation, and motility. Zyxin-null fibroblasts also show deficits in actin stress fiber remodeling and exhibit changes in the molecular composition of focal adhesions, most notably by severely reduced accumulation of Ena/VASP proteins. We postulate that zyxin cooperates with Ena/VASP proteins and caldesmon to influence integrin-dependent cell motility and actin stress fiber remodeling.

Sign in / Sign up

Export Citation Format

Share Document