Role of cAMP-dependent protein kinase A activity in endothelial cell cytoskeleton rearrangement.
To examine signaling mechanisms relevant to cAMP/protein kinase A (PKA)-dependent endothelial cell barrier regulation, we investigated the impact of the cAMP/PKA inhibitors Rp diastereomer of adenosine 3',5'-cyclic monophosphorothioate (Rp-cAMPS) and PKA inhibitor (PKI) on bovine pulmonary artery and bovine lung microvascular endothelial cell cytoskeleton reorganization. Rp-cAMPS as well as PKI significantly increased the formation of actin stress fibers and intercellular gaps but did not alter myosin light chain (MLC) phosphorylation, suggesting that the Rp-cAMPS-induced contractile phenotype evolves in an MLC-independent fashion. We next examined the role of extracellular signal-regulated kinases (ERKs) in Rp-cAMPS- and PKI-induced actin rearrangement. The activities of both ERK1/2 and its upstream activator Raf-1 were transiently enhanced by Rp-cAMPS and linked to the phosphorylation of the well-known ERK cytoskeletal target caldesmon. Inhibition of the Raf-1 target ERK kinase (MEK) either attenuated or abolished Rp-cAMPS- and PKI-induced ERK activation, caldesmon phosphorylation, and stress fiber formation. In summary, our data elucidate the involvement of the p42/44 ERK pathway in cytoskeletal rearrangement evoked by reductions in PKA activity and suggest the involvement of significant cross talk between cAMP- and ERK-dependent signaling pathways in endothelial cell cytoskeletal organization and barrier regulation.
Liu
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F
Division of Pulmonary and Critical Care Medicine, Department of Medicine, Johns Hopkins University School of Medicine, Baltimore, MD 21224-6801, USA.
Verin
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AD
Borbiev
T
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Garcia
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JG
D015124:8-Bromo Cyclic Adenosine Monophosphate / Q000031:analogs & derivatives / Q000494:pharmacology; D000818:Animals; D002148:Calmodulin-Binding Proteins / Q000378:metabolism; D002199:Capillary Permeability / Q000187:drug effects; D002352:Carrier Proteins / Q000494:pharmacology; D002417:Cattle; D002478:Cells, Cultured; D000242:Cyclic AMP / Q000031:analogs & derivatives* / Q000378:metabolism* / Q000494:pharmacology; D017868:Cyclic AMP-Dependent Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D003599:Cytoskeleton / Q000378:metabolism*; D017097:Electric Impedance; D004730:Endothelium, Vascular / Q000166:cytology / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D047908:Intracellular Signaling Peptides and Proteins; D008168:Lung / Q000098:blood supply; D008833:Microcirculation / Q000378:metabolism; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D018994:Myosin Light Chains / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D011651:Pulmonary Artery; D015398:Signal Transduction / Q000187:drug effects; D013873:Thionucleotides / Q000494:pharmacology
C105593:8-bromoadenosine-3',5'-cyclic monophosphorothioate; D002148:Calmodulin-Binding Proteins; D002352:Carrier Proteins; D004791:Enzyme Inhibitors; D047908:Intracellular Signaling Peptides and Proteins; D018994:Myosin Light Chains; D013873:Thionucleotides; C015482:protein kinase modulator; D015124:8-Bromo Cyclic Adenosine Monophosphate; C016957:adenosine-3',5'-cyclic phosphorothioate; D000242:Cyclic AMP; D017868:Cyclic AMP-Dependent Protein Kinases; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases