Endothelial paxillin and focal adhesion kinase (FAK) play a critical role in neutrophil transmigration

2012 ◽  
Vol 42 (2) ◽  
pp. 436-446 ◽  
Author(s):  
Sean A. Parsons ◽  
Ritu Sharma ◽  
Dawn L. Roccamatisi ◽  
Hong Zhang ◽  
Björn Petri ◽  
...  
Keyword(s):  
Focal Adhesion Kinase ◽  
Focal Adhesion ◽  
Critical Role

Focal Adhesion Kinase Inactivation Reduces the Development of Acute Leukemia and Partially Rescues Hematopoietic Stem Cell Defects in Pten-Knockout Mice

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 864-864
Author(s):  
Dewen You ◽  
Andrew Volk ◽  
Clare Sun ◽  
Junping Xin ◽  
Geunhyoung Ha ◽  
...  
Keyword(s):  
Focal Adhesion Kinase ◽  
Phosphatase Activity ◽  
Focal Adhesion ◽  
Protein Phosphatase ◽  
Molecular Mechanisms ◽  
Conflicts Of Interest ◽  
Critical Role ◽  
Hematopoietic Stem ◽  
Hematopoietic System ◽  
Polycytidylic Acid

Abstract Abstract 864 Phosphatase and tensin homolog on chromosome 10 (Pten) is a tumor suppressor which possesses both lipid and protein phosphatase activities. Mutations and epigenetic inactivations of the Pten gene are commonly detected in a large number of tissue malignancies, including leukemias and lymphomas. Studies using Hematopoietic Pten-knockout in adult mice (Pten−/−) have demonstrated that Pten plays a critical role in maintaining the homeostasis of bone marrow (BM) hematopoiesis. Pten inactivation promotes the proliferation and peripheral mobilization of BM hematopoietic stem cells (HSCs). Pten−/− mice develop myeloproliferative disorders (MPD) within days, followed by acute leukemic transformation. Most previous studies attributed such phenotypic changes observed in Pten−/− mice to excessive activation of the PI3K/AKT/mTOR signal, a consequence of the loss of Pten's lipid phosphatase activity. However, the role of Pten's protein phosphatase activity in the regulation of HSCs and leukemogenesis is not well studied. Focal adhesion kinase (Fak) is a critical substrate for the protein phosphatase activity of Pten. Dysregulation of Fak has been observed in many cancers, including acute myeloid leukemias (AML) and acute lymphocytic leukemias (ALL). Therefore, we postulated that Fak might play a pivotal role in the development and progression of leukemia following Pten deletion. To test this hypothesis, we generated Mx1-Cre+Ptenfl/flFakfl/fl mice (an interferon-inducible Pten and Fak compound-knockout, Pten−/−Fak−/−) in which both the Pten and Fak genes in the hematopoietic system are deleted upon injection of polyinosinic-polycytidylic acid (pI-pC). Our results showed that the genetic inactivation of Fak can partially rescue HSC defects associated with Pten deficiency. We found that peripheral mobilization of HSCs in Pten−/−Fak−/− mice is significantly reduced compared to Pten−/− mice. As a consequence, more long-term HSCs (LT-HSCs) are preserved in the BM of Pten−/−Fak−/− mice compared to Pten−/− mice. Transplantation studies suggested that the hematopoietic reconstitutive capacity of Pten−/−Fak−/− HSCs is significantly improved compared to Pten−/− HSCs. Although Fak deletion fails to prevent the development of MPD in Pten−/− mice, Fak deletion does significantly reduce the frequency of AML/ALL, also significantly delays the onset of AML/ALL in comparison to Pten−/− mice. This study suggests that Fak might be a potential target for preventing the MPD-to-AML/ALL transformation and therefore blocking the Fak activity may hold a promise for a novel anti-leukemia therapy. The molecular mechanisms underlying the phenotype restoration of Pten−/− mice by Fak deletion in the hematopoietic system are actively being studied in our laboratory. Disclosures: No relevant conflicts of interest to declare.

Abstract P154: Focal Adhesion Kinase Plays a Critical Role in Upregulating Fibrogenesis in Load-Induced Cardiac Hypertrophy

2011 ◽  
Vol 109 (suppl_1) ◽  
Author(s):  
Ana Paula Dalla Costa ◽  
Carolina F Clemente ◽  
Thais H Theizen ◽  
José Roberto Souza ◽  
Leandro Cardoso ◽  
...  
Keyword(s):  
Cardiac Hypertrophy ◽  
Focal Adhesion Kinase ◽  
Myocardial Fibrosis ◽  
Focal Adhesion ◽  
Cardiac Fibroblasts ◽  
Heart Diseases ◽  
Smooth Muscle Actin ◽  
Critical Role ◽  
Matrix Metalloproteinase 2 ◽  
Activated Fibroblasts

Myocardial fibrosis is maladaptive, accelerating the evolution of diseased hearts to failure. The pathogenesis of myocardial fibrosis is critically dependent on complex processes of activation (i.e. enhanced proliferation, production and secretion of soluble factors, collagen and matrix metalloproteinases) and terminal differentiation of cardiac fibroblasts into myofibroblasts, resultant from the mobilization of numerous signaling molecules by physical and humoral stimuli. Noting that Focal Adhesion Kinase (FAK) is activated in areas of ongoing myocardial fibrosis, we sought to examine whether it is a critical mediator of fibrogenesis in load-induced hypertrophic hearts. Isolated fibroblasts from hypertrophic hearts of mice subjected to transverse aortic constriction (TAC; 1 to 8 weeks) were highly activated as recognized by markers that indicate enhanced proliferation (nuclear Ki67), production of collagen and matrix metalloproteinase-2 (MMP-2) and differentiation into myofibroblasts (expression of α-smooth muscle actin - α-SMA). In these cells, FAK was upregulated, as also were its dowstream pathways Src/ERK1/2 and PI3K/AKT/mTOR. Depletion of FAK (∼80%) after treatment with small interfering RNA (siRNA-FAK) markedly attenuated cardiac hypertrophy and fibrosis, and significantly reduced the number of activated fibroblasts harvested from overloaded hearts. Restoration of FAK function by overexpressing a full-length FAK construct in these cells, selectively enhanced the activity of the downstream PI3K/AKT/mTOR and rescued the activated phenotype of fibroblasts. Transfection with an inactive FAK mutant (Tyr397 substituted by phenylalanine) did not rescue the activated phenotype of fibroblasts harvested from overloaded hearts depleted of FAK. However, cells harvested from overloaded hearts depleted of FAK and treated with the mTOR activating aminoacid leucine showed typical phenotype of activated fibroblasts. These findings uncover a role for FAK in regulating the signaling cascade PI3K/AKT/mTOR in cardiac fibroblasts, which seems to be critical for the pathogenesis of myocardial fibrosis in hypertrophic hearts. Targeting this pathway may provide a novel strategy for treating hypertrophic heart diseases.

scholarly journals Resealing of endothelial junctions by focal adhesion kinase

2007 ◽  
Vol 292 (1) ◽  
pp. L334-L342 ◽  
Author(s):  
Sadiqa K. Quadri ◽  
Jahar Bhattacharya
Keyword(s):  
Focal Adhesion Kinase ◽  
Focal Adhesion ◽  
Time Course ◽  
Critical Role ◽  
Adhesion Formation ◽  
Adherens Junction ◽  
Barrier Properties ◽  
Cell Periphery ◽  
Continuous Presence ◽  
E Cadherin

Endothelial cell (EC) junctions determine vascular barrier properties and are subject to transient opening to allow liquid flux from blood to tissue. Although EC junctions open in the presence of permeability-enhancing factors, including oxidants, the mechanisms by which they reseal remain inadequately understood. To model opening and resealing of EC junctions in the presence of an oxidant, we quantified changes in H2O2-induced transendothelial resistance (TER) in monolayers of rat lung microvascular EC. During a 30-min exposure, H2O2 (100 μM) decreased TER for an initial ∼10 min, indicating junctional opening. Subsequently, despite continuous presence of H2O2, TER recovered to baseline, indicating the activation of junctional resealing mechanisms. These bimodal TER transients matched the time course of loss and then gain of E-cadherin at EC junctions. The timing of the TER decrease matched the onset of focal adhesion formation, while F-actin increase at the cell periphery occurred with a time course that complemented the recovery of peripheral E-cadherin. In monolayers expressing a focal adhesion kinase (FAK) mutant (del-FAK) that inhibits FAK activity, the initial H2O2-induced junctional opening was present, although the subsequent junctional recovery was blocked. Expression of transfected E-cadherin was evident at the cell periphery of wild-type but not del-FAK-expressing EC. E-cadherin overexpression in del-FAK-expressing EC failed to effect major rescue of the junctional resealing response. These findings indicate that in oxidant-induced EC junction opening, FAK plays a critical role in remodeling the adherens junction to reseal the barrier.

scholarly journals Integrin-dependent translocation of phosphoinositide 3-kinase to the cytoskeleton of thrombin-activated platelets involves specific interactions of p85 alpha with actin filaments and focal adhesion kinase.

1995 ◽  
Vol 129 (3) ◽  
pp. 831-842 ◽  
Author(s):  
C Guinebault ◽  
B Payrastre ◽  
C Racaud-Sultan ◽  
H Mazarguil ◽  
M Breton ◽  
...  
Keyword(s):  
Focal Adhesion Kinase ◽  
Tyrosine Phosphorylation ◽  
Focal Adhesion ◽  
Actin Filaments ◽  
Actin Polymerization ◽  
Subcellular Fraction ◽  
Critical Role ◽  
Physical Interaction ◽  
Activated Platelets ◽  
Phosphoinositide 3 Kinase

Thrombin-induced accumulation of phosphatidylinositol 3,4-bisphosphate (PtdIns(3,4)P2) but not of PtdIns(3,4,5,)P3 is strongly correlated with the relocation to the cytoskeleton of 29% of the p85 alpha regulatory subunit of phosphoinositide 3-kinase (PtdIns 3-kinase) and is accompanied by a significant increase in PtdIns 3-kinase activity in this subcellular fraction. Actually, PtdIns(3,4)P2 accumulation and PtdIns 3-kinase, pp60c-src, and p125FAK translocations as well as aggregation were concomitant events occurring with a distinct lag after actin polymerization. The accumulation of PtdIns(3,4)P2 and the relocalization of PtdIns 3-kinase to the cytoskeleton were both dependent on tyrosine phosphorylation, integrin signaling, and aggregation. Furthermore, although p85 alpha was detected in anti-phosphotyrosine immunoprecipitates obtained from the cytoskeleton of thrombin-activated platelets, we failed to demonstrate tyrosine phosphorylation of cytoskeletal p85 alpha. Tyrphostin treatment clearly reduced its presence in this subcellular fraction, suggesting a physical interaction of p85 alpha with a phosphotyrosyl protein. These data led us to investigate the proteins that are able to interact with PtdIns 3-kinase in the cytoskeleton. We found an association of this enzyme with actin filaments: this interaction was spontaneously restored after one cycle of actin depolymerization-repolymerization in vitro. This association with F-actin appeared to be at least partly indirect, since we demonstrated a thrombin-dependent interaction of p85 alpha with a proline-rich sequence of the tyrosine-phosphorylated cytoskeletal focal adhesion kinase, p125FAK. In addition, we show that PtdIns 3-kinase is significantly activated by the p125FAK proline-rich sequence binding to the src homology 3 domain of p85 alpha subunit. This interaction may represent a new mechanism for PtdIns 3-kinase activation at very specific areas of the cell and indicates that the focal contact-like areas linked to the actin filaments play a critical role in signaling events that occur upon ligand engagement of alpha IIb/beta 3 integrin and platelet aggregation evoked by thrombin.

scholarly journals v-Crk Activates the Phosphoinositide 3-Kinase/AKT Pathway by Utilizing Focal Adhesion Kinase and H-Ras

2002 ◽  
Vol 22 (20) ◽  
pp. 7015-7023 ◽  
Author(s):  
Tsuyoshi Akagi ◽  
Kazutaka Murata ◽  
Tomoyuki Shishido ◽  
Hidesaburo Hanafusa
Keyword(s):  
Focal Adhesion Kinase ◽  
Focal Adhesion ◽  
Critical Role ◽  
Sh3 Domain ◽  
Sh2 Domain ◽  
Dominant Negative ◽  
Regulatory Subunit ◽  
Akt Pathway ◽  
Dominant Negative Mutant ◽  
Phosphoinositide 3 Kinase

ABSTRACT v-Crk, an oncogene product of avian sarcoma virus CT10, efficiently transforms chicken embryo fibroblasts (CEF). We have recently reported that constitutive activation of the phosphoinositide 3-kinase (PI3K)/AKT pathway plays a critical role in the v-Crk-induced transformation of CEF. In the present study we investigated the molecular mechanism by which v-Crk activates the PI3K/AKT pathway. First, we found that v-Crk promotes the association of the p85 regulatory subunit of PI3K with focal adhesion kinase (FAK) by inducing the phosphorylation of the Y397 residue in FAK. This FAK phosphorylation needs activation of the Src family tyrosine kinase(s) for which the v-Crk SH2 domain is responsible. v-Crk was unable to activate the PI3K/AKT pathway in FAK-null cells, indicating the functional importance of FAK. In addition, we found that H-Ras is also required for the activation of the PI3K/AKT pathway. The v-Crk-induced activation of AKT was greatly enhanced by the overexpression of H-Ras or its guanine nucleotide exchange factor mSOS, which binds to the v-Crk SH3 domain, whereas a dominant-negative mutant of H-Ras almost completely suppressed this activation. Furthermore, we showed that v-Crk stimulates the interaction of H-Ras with the Ras binding domain in the PI3K p110 catalytic subunit. Our data indicated that the v-Crk-induced activation of PI3K/AKT pathway was cooperatively achieved by two distinct interactions. One is the interaction of p85 with tyrosine-phosphorylated FAK promoted by the v-Crk SH2 domain, and another is the interaction of p110 with H-Ras dictated by the v-Crk SH3 domain.

Stimulation of cell migration by overexpression of focal adhesion kinase and its association with Src and Fyn

1996 ◽  
Vol 109 (7) ◽  
pp. 1787-1794 ◽  
Author(s):  
L.A. Cary ◽  
J.F. Chang ◽  
J.L. Guan
Keyword(s):  
Cell Migration ◽  
Focal Adhesion Kinase ◽  
Focal Adhesion ◽  
Cho Cells ◽  
Critical Role ◽  
Cellular Interactions ◽  
Cellular Functions ◽  
And Migration ◽  
Biochemical Signals

Cellular interactions with the extracellular matrix proteins play important roles in a variety of biological processes. Recent studies suggest that integrin-mediated cell-matrix interaction can transduce biochemical signals across the plasma membrane to regulate cellular functions such as proliferation, differentiation and migration. These studies have implicated a critical role of focal adhesion kinase (FAK) in integrin-mediated signal transduction pathways. We report here that overexpression of FAK in CHO cells increased their migration on fibronectin. A mutation of the major autophosphorylation site Y397 in FAK abolished its ability to stimulate cell migration, while phosphorylation of Y397 in a kinase-defective FAK by endogenous FAK led to increased migration. We also find that the wild-type and the kinase-defective FAK were associated with Src and Fyn in CHO cells whereas the F397 mutant was not. These results directly demonstrate a functional role for FAK in integrin signaling leading to cell migration. They also provide evidence for the functional significance of FAK/Src complex formation in vivo.

scholarly journals Focal Adhesion Kinase Controls Cellular Levels of p27/Kip1 and p21/Cip1 through Skp2-Dependent and -Independent Mechanisms

2006 ◽  
Vol 26 (11) ◽  
pp. 4201-4213 ◽  
Author(s):  
Patrick Bryant ◽  
Qingxia Zheng ◽  
Kevin Pumiglia
Keyword(s):  
Cell Proliferation ◽  
Endothelial Cell ◽  
Focal Adhesion Kinase ◽  
Focal Adhesion ◽  
Kinase Inhibitors ◽  
Critical Role ◽  
Dominant Negative ◽  
Regulatory Control ◽  
Endothelial Cell Proliferation ◽  
F Box Protein

ABSTRACT Endothelial cell proliferation is a critical step in angiogenesis and requires a coordinated response to soluble growth factors and the extracellular matrix. As focal adhesion kinase (FAK) integrates signals from both adhesion events and growth factor stimulation, we investigated its role in endothelial cell proliferation. Expression of a dominant-negative FAK protein, FAK-related nonkinase (FRNK), impaired phosphorylation of FAK and blocked DNA synthesis in response to multiple angiogenic stimuli. These results coincided with elevated cyclin-dependent kinase inhibitors (CDKIs) p21/Cip and p27/Kip, as a consequence of impaired degradation. FRNK inhibited the expression of Skp2, an F-box protein that targets CDKIs, by inhibiting mitogen-induced mRNA. The FAK-regulated degradation of p27/Kip was Skp2 dependent, while levels of p21/Cip were regulated independent of Skp2. Skp2 is required for endothelial cell proliferation as a consequence of degrading p27. Finally, knockdown of both p21 and p27 in FRNK-expressing cells completely restored mitogen-induced endothelial cell proliferation. These data demonstrate a critical role for FAK in the regulation of CDKIs through two independent mechanisms: Skp2 dependent and Skp2 independent. They also provide important insights into the requirement of focal adhesion kinase for normal vascular development and reveal novel regulatory control points for angiogenesis.

Activation of integrin β1-focal adhesion kinase-RasGTP pathway plays a critical role in TGF beta1-induced podocyte injury

2013 ◽  
Vol 25 (12) ◽  
pp. 2769-2779 ◽  
Author(s):  
Yu-Jing Zhang ◽  
Zhi-Liang Tian ◽  
Xin-Yan Yu ◽  
Xia-Xia Zhao ◽  
Li Yao
Keyword(s):  
Focal Adhesion Kinase ◽  
Focal Adhesion ◽  
Critical Role ◽  
Integrin Β1 ◽  
Podocyte Injury
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