scholarly journals Mechanical Strain on Osteoblasts Activates Autophosphorylation of Focal Adhesion Kinase and Proline-rich Tyrosine Kinase 2 Tyrosine Sites Involved in ERK Activation

2004 ◽  
Vol 279 (29) ◽  
pp. 30588-30599 ◽  
Author(s):  
Nadia Boutahar ◽  
Alain Guignandon ◽  
Laurence Vico ◽  
Marie-Hélène Lafage-Proust
Keyword(s):  
Tyrosine Kinase ◽  
Focal Adhesion Kinase ◽  
Focal Adhesion ◽  
Mechanical Strain ◽  
Erk Activation ◽  
Tyrosine Kinase 2

Src and focal adhesion kinase mediate mechanical strain-induced proliferation and ERK1/2 phosphorylation in human H441 pulmonary epithelial cells

2007 ◽  
Vol 292 (5) ◽  
pp. C1701-C1713 ◽  
Author(s):  
Lakshmi S. Chaturvedi ◽  
Harold M. Marsh ◽  
Marc D. Basson
Keyword(s):  
Epithelial Cells ◽  
Focal Adhesion Kinase ◽  
Focal Adhesion ◽  
Mechanical Strain ◽  
A549 Cells ◽  
Cyclic Strain ◽  
Short Interfering Rna ◽  
Erk Activation ◽  
Pulmonary Epithelial Cells ◽  
Interfering Rna

Pulmonary epithelial cells are exposed to repetitive deformation during physiological breathing and mechanical ventilation. Such deformation may influence pulmonary growth, development, and barotrauma. Although deformation stimulates proliferation and activates extracellular signal-regulated kinases (ERK1/2) in human pulmonary epithelial H441 cells, the upstream mechanosensors that induce ERK activation are poorly understood. We investigated whether c-Src or focal adhesion kinase (FAK) mediates cyclic mechanical strain-induced ERK1/2 activation and proliferation in human pulmonary epithelial (NCI-H441) cells. The H441 and A549 cells were grown on collagen I-precoated membranes and were subjected to an average 10% cyclic mechanical strain at 20 cycles/min. Cyclic strain activated Src within 2 min by increasing phosphorylation at Tyr418, followed by rapid phosphorylation of FAK at Tyr397 and Tyr576 and ERK1/2 at Thr202/Tyr204 ( n = 5, P < 0.05). Twenty-four (A549 cells) and 24–72 h (H441 cells) of cyclic mechanical strain increased cell numbers compared with static culture. Twenty-four hours of cyclic strain also increased H441 FAK, Src, and ERK phosphorylation without affecting total FAK, Src, or ERK protein. The mitogenic effect was blocked by Src (10 μmol/l PP2 or short interfering RNA targeted to Src) or MEK (50 μmol/l PD-98059) inhibition. PP2 also blocked strain-induced phosphorylation of FAK-Tyr576 and ERK-Thr202/Tyr204 but not FAK-Tyr397. Reducing FAK by FAK-targeted short interfering RNA blocked mechanical strain-induced mitogenicity and significantly attenuated strain-induced ERK activation but not strain-induced Src phosphorylation. Together, these results suggest that repetitive mechanical deformation induced by ventilation supports pulmonary epithelial proliferation by a pathway involving Src, FAK, and then ERK signaling.

Proline-rich Tyrosine Kinase 2 and Focal Adhesion Kinase Are Involved in Different Phases of Platelet Activation by vWF

2002 ◽  
Vol 87 (03) ◽  
pp. 509-517 ◽  
Author(s):  
Ilaria Canobbio ◽  
Paolo Lova ◽  
Fabiola Sinigaglia ◽  
Cesare Balduini ◽  
Mauro Torti
Keyword(s):  
Tyrosine Kinase ◽  
Focal Adhesion Kinase ◽  
Tyrosine Phosphorylation ◽  
Focal Adhesion ◽  
Tyrosine Kinases ◽  
Human Platelets ◽  
Glycoprotein Ib ◽  
Tyrosine Kinase 2 ◽  
Calcium Chelation ◽  
Stimulation Of

SummaryStimulation of human platelets with von Willebrand factor (vWF) induces the rapid tyrosine phosphorylation of several proteins, but very little is known on the tyrosine kinases involved in this process. In the present work, we investigated and compared the activation of two related tyrosine kinases expressed in platelets: the proline-rich tyrosine kinase 2 (Pyk2) and the focal adhesion kinase (FAK). Both kinases were tyrosine phosphorylated upon vWF interaction with glycoprotein Ib-IX-V complex, but with different mechanisms. Tyrosine phosphorylation of FAK was totally dependent on thromboxane A2 production, and was inhibited by the integrin αIIbβ3 antagonist RGDS peptide. Moreover, chelation of intracellular calcium or inhibition of protein kinase C (PKC) totally blocked vWF-induced tyrosine phosphorylation of FAK, indicating that this event is downstream phospholipase A2 and phospholipase C activation. By contrast, tyrosine phosphorylation of Pyk2 was only partially reduced by aspirin and RGDS, and was not affected by either calcium chelation or PKC inhibition, suggesting that activation of this kinase does not require phospholipase-mediated signalling. Both FAK and Pyk2 translocated to the cytoskeleton upon vWF stimulation of human platelets by a mechanism depending on agonist-induced actin polymerisation. Prevention of cytoskeletal relocation of Pyk2 and FAK by cytochalasin D totally blocked vWFinduced tyrosine phosphorylation of both kinases. Finally, phosphorylation of Pyk2 induced by vWF, but not by thrombin, was inhibited by piceatannol, suggesting that this kinase lies downstream Syk. These results demonstrate that both Pyk2 and FAK are involved in platelet stimulation by vWF, but indicate that only Pyk2 may play a role in the early signal transduction events activated by ligand binding to glycoprotein Ib-IX-V.

scholarly journals Induction of Apoptosis after Expression of PYK2, a Tyrosine Kinase Structurally Related to Focal Adhesion Kinase

1997 ◽  
Vol 139 (2) ◽  
pp. 529-539 ◽  
Author(s):  
Wen-cheng Xiong ◽  
J. Thomas Parsons
Keyword(s):  
Tyrosine Kinase ◽  
Cell Survival ◽  
Focal Adhesion Kinase ◽  
Focal Adhesion ◽  
Cell Attachment ◽  
Uv Light ◽  
Apoptotic Cell ◽  
Cellular Adhesion ◽  
Tyrosine Kinase 2 ◽  
Induction Of Apoptosis

Many cells (e.g., epithelial cells) require attachment to the extracellular matrix (ECM) to survive, a phenomenon known as anchorage-dependent cell survival. Disruption of the cell–ECM interactions mediated by the integrin receptors results in apoptosis. Focal adhesion kinase (FAK), a 125-kD protein tyrosine kinase activated by integrin engagement, appears to be involved in mediating cell attachment and survival. Proline-rich tyrosine kinase 2 (PYK2), also known as cellular adhesion kinase β (CAKβ) and related adhesion focal tyrosine kinase, is a second member of the FAK subfamily and is activated by an increase in intracellular calcium levels, or treatment with TNFα and UV light. However, the function of PYK2 remains largely unknown. In this study, we show that over-expression of PYK2, but not FAK, in rat and mouse fibroblasts leads to apoptotic cell death. Using a series of deletion mutants and chimeric fusion proteins of PYK2/FAK, we determined that the NH2-terminal domain and tyrosine kinase activity of PYK2 were required for the efficient induction of apoptosis. Furthermore, the apoptosis mediated by PYK2 could be suppressed by over-expressing catalytically active v-Src, c-Src, phosphatidylinositol-3-kinase, or Akt/protein kinase B. In addition, it could also be suppressed by overexpressing an ICE or ICE-like proteinase inhibitor, crmA, but not Bcl2. Collectively, our results suggest that PYK2 and FAK, albeit highly homologous in primary structure, appear to have different functions; FAK is required for cell survival, whereas PYK2 induces apoptosis in fibroblasts.

scholarly journals Phosphorylation of Focal Adhesion Kinase (FAK) on Ser732 Is Induced by Rho-dependent Kinase and Is Essential for Proline-rich Tyrosine Kinase-2–mediated Phosphorylation of FAK on Tyr407 in Response to Vascular Endothelial Growth Factor

2006 ◽  
Vol 17 (8) ◽  
pp. 3508-3520 ◽  
Author(s):  
Fabrice Le Boeuf ◽  
François Houle ◽  
Mark Sussman ◽  
Jacques Huot
Keyword(s):  
Vascular Endothelial Growth Factor ◽  
Cell Migration ◽  
Growth Factor ◽  
Tyrosine Kinase ◽  
Focal Adhesion Kinase ◽  
Focal Adhesion ◽  
Endothelial Cell Migration ◽  
Vascular Endothelial ◽  
Tyrosine Kinase 2 ◽  
Endothelial Growth Factor

Focal adhesion kinase (FAK) is phosphorylated on tyrosine and serine residues after cell activation. In the present work, we investigated the relationship between tyrosine and serine phosphorylation of FAK in promoting endothelial cell migration in response to vascular endothelial growth factor (VEGF). We found that VEGF induces the activation of the Rho-dependent kinase (ROCK) downstream from vascular endothelial growth factor receptor (VEGFR) 2. In turn, activated ROCK directly phosphorylates FAK on Ser732. Proline-rich tyrosine kinase-2 (Pyk2) is also activated in response to VEGF. Its activation requires the clustering of integrin αvβ3 and triggers directly the phosphorylation of Tyr407 within FAK, an event necessary for cell migration. Interestingly, ROCK-mediated phosphorylation of Ser732 is essential for Pyk2-dependent phosphorylation of Tyr407, because the latter is abrogated in cells expressing a FAK mutant that is nonphosphorylatable on Ser732. We suggest that VEGF elicits the activation of the VEGFR2–ROCK pathway, leading to phosphorylation of Ser732 within FAK. In turn, phosphorylation of Ser732 would change the conformation of FAK, making it accessible to Pyk2 activated in response to its association with integrin β3. Then, activated Pyk2 triggers the phosphorylation of FAK on Tyr407, promoting cell migration.

Expression and characterization of splice variants of PYK2, a focal adhesion kinase-related protein

1998 ◽  
Vol 111 (14) ◽  
pp. 1981-1991 ◽  
Author(s):  
W.C. Xiong ◽  
M. Macklem ◽  
J.T. Parsons
Keyword(s):  
Tyrosine Kinase ◽  
Focal Adhesion Kinase ◽  
Rat Brain ◽  
Functional Diversity ◽  
Focal Adhesion ◽  
Focal Adhesions ◽  
Receptor Protein ◽  
Protein Tyrosine ◽  
Terminal Domain ◽  
Tyrosine Kinase 2

Focal adhesion kinase and the recently identified proline-rich tyrosine kinase 2 (PYK2), also known as cell adhesion kinase β, related adhesion focal tyrosine kinase or calcium-dependent protein tyrosine kinase, define a new family of non-receptor protein tyrosine kinases. Activation of PYK2 has been implicated in multiple signaling events, including modulation of ion channels, T- and B-cell receptor signaling and cell death. Mechanisms underlying the functional diversity of PYK2 are unclear. Here, we provide evidence for two novel alternatively expressed isoforms of PYK2. One isoform, designated PYK2s (PYK2 splice form), appears to be a splice variant of PYK2 lacking 42 amino acids within the C-terminal domain. A second isoform, referred to as PRNK (PYK2-related non-kinase), appears to be specified by mRNAs that encode only part of the C-terminal domain of PYK2. Northern blot analysis indicates that the unspliced PYK2 is expressed at high levels in the brain and poorly expressed in the spleen, whereas PYK2s and PRNK are expressed in the spleen. In situ hybridization studies of rat brain demonstrate that the unspliced PYK2 is selectively expressed at high levels in hippocampus, cerebral cortex and olfactory bulb, whereas PYK2s and PRNK are expressed at low levels in all regions of rat brain examined. Immunofluorescence analysis of ectopically expressed PRNK protein shows that PRNK, in contrast to full-length PYK2, is localized to focal adhesions by sequences within the focal adhesion targeting domain. In addition, PYK2, but not PRNK, interacts with p130(cas)and Graf. These results imply that PRNK may selectively regulate PYK2 function in certain cells by binding to some but not all PYK2 binding partners, and the functional diversity mediated by PYK2 may be due in part to complex alternative splicing.

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