Role of Formation of an ERK-FAK-Paxillin Complex in Migration of Human Corneal Epithelial Cells during Wound Closure In Vitro

Teranishi, Shinichiro; Kimura, Kazuhiro; Nishida, Teruo

doi:10.1167/iovs.08-2534

Cited by 57 publications

(55 citation statements)

References 27 publications

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“…5), followed by a second broader activation peak 3-5 hours later. Previous wound healing experiments involved activated ERK in regulating cell migration: Matsubayashi and co-workers demonstrated that inhibition of ERK activation results in markedly reduced movement of the epithelial sheet in wound healing experiments [23] and in corneal endothelial cells ERK has also been shown to affect migration speed [40,41]. Our results confirm the overall ERK involvement in regulating migration speed, and highlight its unique role in determining directional migration.…”

Section: Discussionsupporting

confidence: 83%

Ras Activated ERK and PI3K Pathways Differentially Affect Directional Movement of Cultured Fibroblasts

Sepe

Ferrari

Cantarella

et al. 2013

Cell Physiol Biochem

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Background: Cell migration is essential in physiological and pathological processes, such as wound healing and metastasis formation. Ras involvement in these processes has been extensively demonstrated. This work attempts to characterize Ras regulation of the phenomena determining directional cell migration by separately analyzing the role of its principal effector pathways, MAPK and PI3K. Methods: NIH3T3 and NIHRasV12 fibroblasts were followed in wound healing assays to study, in time and under a directional stimulus, cell migration both under standard conditions and in presence of MAPK and PI3K inhibitors. Several parameters, descriptive of specific aspects of cell motion, were evaluated by coupling dynamic microscopy with quantitative and statistical methods. Quantitative Western Blots coupled with immunofluorescence stainings, were used to evaluate ERK activation. Results: Constitutive RasV12 activation confers to NIH3T3 the ability to close the wound faster. Neither increased cell proliferation nor higher speed explains the accelerated healing, but the increased directional migration drives the wound closure. Inhibition of ERK activation, which occurs immediately after wound, greatly blocks the directional migration, while inhibition of PI3K pathway reduces cell speed but does not prevent wound closure. Conclusion: Ras is greatly involved in determining and regulating directionality, ERK is its key effector for starting, driving and regulating directional movement.

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Section: Discussionsupporting

confidence: 83%

Ras Activated ERK and PI3K Pathways Differentially Affect Directional Movement of Cultured Fibroblasts

Sepe

Ferrari

Cantarella

et al. 2013

Cell Physiol Biochem

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“…Paxillin was originally identified as a major substrate for the v-Src tyrosine kinase (14), and c-Src and FAK have been identified as key kinases responsible for paxillin tyrosine phosphorylation in response to a variety of stimuli (1,5,24,27,37,47). Although LPS stimulated c-Src and FAK phosphorylation, knockdown of Src with siRNA or overexpression FRANK, a dominant-negative form of FAK, did not block LPS-induced tyrosine phosphorylation of paxillin in HLMVECs.…”

Section: Discussionmentioning

confidence: 98%

c-Abl mediated tyrosine phosphorylation of paxillin regulates LPS-induced endothelial dysfunction and lung injury

Usatyuk

Lele

et al. 2015

American Journal of Physiology-Lung Cellular and Molecular Phys

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Fu P, Usatyuk PV, Lele A, Harijith A, Gregorio CC, Garcia JG, Salgia R, Natarajan V. c-Abl mediated tyrosine phosphorylation of paxillin regulates LPS-induced endothelial dysfunction and lung injury. Am J Physiol Lung Cell Mol Physiol 308: L1025-L1038, 2015. First published March 20, 2015 doi:10.1152/ajplung.00306.2014Paxillin is phosphorylated at multiple residues; however, the role of tyrosine phosphorylation of paxillin in endothelial barrier dysfunction and acute lung injury (ALI) remains unclear. We used siRNA and site-specific nonphosphorylable mutants of paxillin to abrogate the function of paxillin to determine its role in lung endothelial permeability and ALI. In vitro, lipopolysaccharide (LPS) challenge of human lung microvascular endothelial cells (HLMVECs) resulted in enhanced tyrosine phosphorylation of paxillin at Y31 and Y118 with no significant change in Y181 and significant barrier dysfunction. Knockdown of paxillin with siRNA attenuated LPS-induced endothelial barrier dysfunction and destabilization of VE-cadherin. LPS-induced paxillin phosphorylation at Y31 and Y118 was mediated by c-Abl tyrosine kinase, but not by Src and focal adhesion kinase. c-Abl siRNA significantly reduced LPS-induced endothelial barrier dysfunction. Transfection of HLMVECs with paxillin Y31F, Y118F, and Y31/118F double mutants mitigated LPS-induced barrier dysfunction and VE-cadherin destabilization. In vivo, the c-Abl inhibitor AG957 attenuated LPS-induced pulmonary permeability in mice. Together, these results suggest that c-Abl mediated tyrosine phosphorylation of paxillin at Y31 and Y118 regulates LPS-mediated pulmonary vascular permeability and injury.paxillin; c-Abl; lipopolysaccharide; tyrosine kinase; permeability; endothelial dysfunction; cytoskeleton GRAM-NEGATIVE BACTERIA-INDUCED sepsis affects more than 750,000 patients annually in the United States (23). Acute respiratory distress syndrome (ARDS), the most severe form of acute lung injury (ALI), is characterized by endothelial hyperpermeability leading to lung inflammation and injury. Lipopolysaccharide (LPS), the bacterial endotoxin and an outer membrane component, induces proinflammatory cytokines, adhesion molecules, endothelial permeability changes, and apoptosis regulated by TLR-4 signaling via activation of Rho GTPase-dependent signaling cascade, NF-B activation, and rearrangement of cytoskeletal proteins (15, 31). Despite recent advances in understanding the pathogenesis of ALI/ARDS, effective pharmacological therapies are not currently available and the molecular mechanisms regulating endothelial barrier dysfunction in ALI/ARDS are not completely defined.Interactions between focal adhesions, extracellular matrix, and cytoskeletal proteins have been implicated in endothelial cell (EC) remodeling and barrier dysfunction associated with LPS-and ventilator-induced lung injury (15,31,48). Specific posttranslational modifications such as phosphorylation of focal adhesion proteins are also known to play a role in endothelial barrier regulation to agonists...

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“…As described in the Introduction, activation of the ERK signaling pathway requires the participation and mediation of PXN in prostate cancer and non-small cell lung cancer (10,17 (11). Several studies have demonstrated that Erk always gives a positive feedback on PXN and plays an important part in the activitation of PXN and interaction between FAK and PXN (11,22,23). Based on these facts, we decided to examine the expression of p-Erk during this experiment.…”

Section: Discussionmentioning

confidence: 99%

Paxillin is positively correlated with the clinicopathological factors of colorectal cancer, and knockdown of Paxillin improves sensitivity to cetuximab in colorectal cancer cells

Wang

Zhang

et al. 2015

Oncology Reports

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Abstract. Paxillin (PXN) encodes a 68-kDa focal adhesionassociated protein and plays an important role in signal transduction, regulation of cell morphology, migration, proliferation and apoptosis. The aim of the present study was to evaluate the relationship between PXN and clinicopathological factors in colorectal cancer, the role of PXN in cetuximab resistance, and whether knockdown of PXN expression could improve the sensitivity to cetuximab in colorectal cancer cells. In the present study, immunohistochemical staining in 148 colorectal carcinoma and 126 normal adjacent tissues was performed, which showed that the positive rate of PXN was significantly higher in the colorectal adenocarcinoma samples than that in the normal colorectal mucosa samples (P<0.001). Moreover, PXN presence was also positively correlated with TNM stage (P=0.023), distant metastasis (P=0.014), recurrence (P=0.032) and reduced survival (P=0.004). In vitro, PXN expression was positively correlated with the proliferation rate in colorectal cells insensitive to cetuximab. Inhibition of PXN expression by PXN-siRNA clearly increased apoptosis by downregulating the phosphorylation of extracellular signal regulated kinase (p-Erk) level, and overexpression of PXN by PXN-cDNA decreased apoptosis by upregulating the p-Erk level. This suggests that overexpression of PXN could be one of the reasons for cetuximab resistance, and downregulation of PXN plays an important role in improving sensitivity to cetuximab by suppressing the activitation of p-Erk in colorectal cancer cells. Above all, knockdown of PXN could represent a rational therapeutic strategy for increasing the sensitivity or overcoming cetuximab-resistance in patients with colorectal cancer.

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Role of Formation of an ERK-FAK-Paxillin Complex in Migration of Human Corneal Epithelial Cells during Wound Closure In Vitro

Abstract: ERK regulates cell migration during wound healing in vitro by modulating the phosphorylation of FAK and paxillin and the consequent formation of focal adhesions. An ERK-FAK-paxillin signaling pathway may play an important role in corneal epithelial wound healing in vivo.

Cited by 57 publications

References 27 publications

Ras Activated ERK and PI3K Pathways Differentially Affect Directional Movement of Cultured Fibroblasts

Ras Activated ERK and PI3K Pathways Differentially Affect Directional Movement of Cultured Fibroblasts

c-Abl mediated tyrosine phosphorylation of paxillin regulates LPS-induced endothelial dysfunction and lung injury

Paxillin is positively correlated with the clinicopathological factors of colorectal cancer, and knockdown of Paxillin improves sensitivity to cetuximab in colorectal cancer cells

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