Differential effect of shear stress on extracellular signal-regulated kinase and N-terminal Jun kinase in endothelial cells. Gi2- and Gbeta/gamma-dependent signaling pathways.
Shear stress differentially regulates production of many vasoactive factors at the level of gene expression in endothelial cells that may be mediated by mitogen-activated protein kinases, including extracellular signal-regulated kinase (ERK) and N-terminal Jun kinase (JNK). Here we show, using bovine aortic endothelial cells (BAEC), that shear stress differentially regulates ERK and JNK by mechanisms involving Gi2 and pertussis toxin (PTx)-insensitive G-protein-dependent pathways, respectively. Shear activated ERK with a rapid, biphasic time course (maximum by 5 min and basal by 30-min shear exposure) and force dependence (minimum and maximum at 1 and 10 dyn/cm2 shear stress, respectively). PTx treatment prevented shear-dependent activation of ERK1/2, consistent with a Gi-dependent mechanism. In contrast, JNK activity was maximally turned on by a threshold level of shear force (0.5 dyn/cm2 or higher) with a much slower and prolonged time course (requiring at least 30 min to 4 h) than that of ERK. Also, PTx had no effect on shear-dependent activation of JNK. To further define the shear-sensitive ERK and JNK pathways, vectors expressing hemagglutinin epitope-tagged ERK (HA-ERK) or HA-JNK were co-transfected with other vectors by using adenovirus-polylysine in BAEC. Expression of the mutant (alpha)i2(G203), antisense G(alpha)i2 and a dominant negative Ras (N17Ras) prevented shear-dependent activation of HA-ERK, while that of (alpha)i2(G204) and antisense (alpha)i3 did not. Expression of a Gbeta/gamma scavenger, the carboxyl terminus of beta-adrenergic receptor kinase (betaARK-ct), and N17Ras inhibited shear-dependent activation of HA-JNK. Treatment of BAEC with genistein prevented shear-dependent activation of ERK and JNK, indicating the essential role of tyrosine kinase(s) in both ERK and JNK pathways. These results provide evidence that 1) Gi2-protein, Ras, and tyrosine kinase(s) are upstream regulators of shear-dependent activation of ERK and 2) that shear-dependent activation of JNK is regulated by mechanisms involving Gbeta/gamma, Ras, and tyrosine kinase(s).
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Department of Pathology, University of Alabama at Birmingham, 35294, USA. Jo@path.vh.uab.edu
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Rong
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McDonald
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D000818:Animals; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000378:metabolism*; D002417:Cattle; D004730:Endothelium, Vascular / Q000201:enzymology*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D019204:GTP-Binding Proteins / Q000378:metabolism*; D019833:Genistein; D007529:Isoflavones / Q000494:pharmacology; D048031:JNK Mitogen-Activated Protein Kinases; D020928:Mitogen-Activated Protein Kinases; D037342:Pertussis Toxin; D011505:Protein-Tyrosine Kinases / Q000378:metabolism; D015398:Signal Transduction; D010566:Virulence Factors, Bordetella / Q000494:pharmacology; D018631:ras Proteins / Q000494:pharmacology
D004791:Enzyme Inhibitors; D007529:Isoflavones; D010566:Virulence Factors, Bordetella; D019833:Genistein; D037342:Pertussis Toxin; D011505:Protein-Tyrosine Kinases; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D048031:JNK Mitogen-Activated Protein Kinases; D020928:Mitogen-Activated Protein Kinases; D019204:GTP-Binding Proteins; D018631:ras Proteins