14-3-3beta-Rac1-p21 activated kinase signaling regulates Akt1-mediated cytoskeletal organization, lamellipodia formation and fibronectin matrix assembly.
Akt1 belongs to the three-gene Akt family and functions as a serine-threonine kinase regulating phosphorylation of an array of substrates and mediating cellular processes such as cell migration, proliferation, survival, and cell cycle. Our previous studies have established the importance of Akt1 in angiogenesis and absence of Akt1 resulted in impaired integrin activation, adhesion, migration, and extracellular matrix assembly by endothelial cells and fibroblasts. In this study, we identify the downstream signaling pathways activated by Akt1 in the regulation of these cellular events. We demonstrate here that Akt1 is necessary for the growth factor stimulated activation of 14-3-3beta-Rac1-p21 activated kinase (Pak) pathway in endothelial cells and fibroblasts. While activation of Akt1 resulted in translocation of Rac1 to membrane ruffles, enhanced Rac1 activity, Pak1 phosphorylation, and lamellipodia formation, resulting in enhanced adhesion and assembly of fibronectin, inhibition of Akt1 resulted in inhibition of these processes due to impaired Rac1-Pak signaling. Formation of lamellipodia, adhesion, and fibronectin assembly by myristoylated Akt1 expression in NIH 3T3 fibroblasts was inhibited by co-expression with either dominant negative Rac1 or dominant negative Pak1. In contrast, impaired lamellipodia formation, adhesion, and fibronectin assembly by dominant negative-Akt1 expression was rescued by co-expression with either constitutively active-Rac1 or -Pak1. Moreover, previously reported defects in adhesion and extracellular matrix assembly by Akt1(-/-) fibroblasts could be rescued by expression with either active-Rac1 or -Pak1, implying the importance of Rac1-Pak signaling in growth factor stimulated cytoskeletal assembly, lamellipodia formation and cell migration in endothelial cells and fibroblasts downstream of Akt1 activation.
Joseph J. Jacobs Center for Thrombosis and Vascular Biology, Department of Molecular Cardiology, Lerner Research Institute, The Cleveland Clinic, Cleveland, Ohio, USA.
Payaningal R
PR
Somanath
Tatiana V
TV
Byzova
D048948:14-3-3 Proteins / Q000378:metabolism*; D000818:Animals; D001011:Aorta / Q000201:enzymology; D002448:Cell Adhesion / Q000187:drug effects; D002465:Cell Movement / Q000187:drug effects; D003599:Cytoskeleton / Q000187:drug effects / Q000378:metabolism*; D042783:Endothelial Cells / Q000166:cytology / Q000187:drug effects / Q000201:enzymology; D004789:Enzyme Activation / Q000187:drug effects; D005109:Extracellular Matrix / Q000187:drug effects / Q000378:metabolism; D005347:Fibroblasts / Q000166:cytology / Q000201:enzymology; D005353:Fibronectins / Q000378:metabolism*; D006801:Humans; D066298:In Vitro Techniques; D036341:Intercellular Signaling Peptides and Proteins / Q000494:pharmacology; D051379:Mice; D041681:NIH 3T3 Cells; D018919:Neovascularization, Physiologic / Q000187:drug effects; D021381:Protein Transport / Q000187:drug effects; D051057:Proto-Oncogene Proteins c-akt / Q000172:deficiency / Q000378:metabolism*; D011554:Pseudopodia / Q000187:drug effects / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects; D054462:p21-Activated Kinases / Q000378:metabolism*; D020830:rac1 GTP-Binding Protein / Q000378:metabolism*
D048948:14-3-3 Proteins; D005353:Fibronectins; D036341:Intercellular Signaling Peptides and Proteins; D051057:Proto-Oncogene Proteins c-akt; D054462:p21-Activated Kinases; D020830:rac1 GTP-Binding Protein