title: Inhibition of Rac activation as a mechanism for negative regulation of actin cytoskeletal reorganization and cell motility by cAMP.
abstract: cAMP has been found to play a role in mediating the negative regulation of cell motility, although its underlying molecular mechanism remains poorly understood. By using CHO (Chinese-hamster ovary) cells that express the EP2 subtype of PGE2 (prostaglandin E2) receptors, we provide evidence that an increase in cellular cAMP content leads to inhibition of cellular Rac activity, which serves as a mechanism for this negative regulation. In CHO cells expressing EP2, but not in vector control cells, PGE2 dose-dependently inhibited chemotaxis towards IGF-I (insulin-like growth factor-I), which is a Rac-dependent process, with the maximal 75% inhibition observed at 10(-8) M PGE2. EP2 stimulation failed to inhibit tyrosine phosphorylation either of IGF-I receptor or IRS-1 (insulin receptor substrate-1), or activation of phosphoinositide 3-kinase or Akt in response to IGF-I, but potently and dose-dependently inhibited IGF-I-induced activation of cellular Rac activity and membrane ruffling. However, PGE2 failed to inhibit Val12-Rac-induced membrane ruffling. Similar to the case of CHO cells, PGE2 inhibited PDGF (platelet-derived growth factor)-induced Rac activation and chemotaxis in vascular smooth muscle cells endogenously expressing EP2. The inhibitory effects of PGE2 on IGF-I-induced chemotaxis, membrane ruffling and Rac activation were faithfully reproduced by a low concentration of forskolin, which induced a comparable extent of cAMP elevation as with 10(-8) M PGE2, and were potentiated by isobutylmethylxanthine. The protein kinase A inhibitor Rp isomer of adenosine 3',5'-cyclic monophosphorothioate reduced PGE2 inhibition of Rac activation and chemotaxis. These results indicate that EP2 mediates Rac inhibition through a mechanism involving cAMP and protein kinase A, thereby inhibiting membrane ruffling and chemotaxis.
authors.affiliation: Department of Physiology, Kanazawa University Graduate School of Medicine, 13-1 Takara-machi, Kanazawa, Ishikawa 920-8640, Japan.
authors.forename: Shin-ya
authors.initials: SY
authors.lastname: Nagasawa
authors.forename: Noriko
authors.initials: N
authors.lastname: Takuwa
authors.forename: Naotoshi
authors.initials: N
authors.lastname: Sugimoto
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Mabuchi
authors.forename: Yoh
authors.initials: Y
authors.lastname: Takuwa
mesh_terms: D000199:Actins / Q000378:metabolism*; D000818:Animals; D016466:CHO Cells; D002465:Cell Movement / Q000187:drug effects; D022081:Cell Surface Extensions / Q000187:drug effects; D002633:Chemotaxis / Q000187:drug effects; D006224:Cricetinae; D000242:Cyclic AMP / Q000378:metabolism*; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D003599:Cytoskeleton / Q000187:drug effects / Q000378:metabolism*; D015232:Dinoprostone / Q000494:pharmacology; D004789:Enzyme Activation / Q000187:drug effects; D020962:Heterotrimeric GTP-Binding Proteins / Q000378:metabolism; D006801:Humans; D055504:Insulin Receptor Substrate Proteins; D007334:Insulin-Like Growth Factor I / Q000037:antagonists & inhibitors / Q000494:pharmacology; D051379:Mice; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D010750:Phosphoproteins / Q000378:metabolism; D010766:Phosphorylation; D010982:Platelet-Derived Growth Factor / Q000494:pharmacology; D017526:Receptor, IGF Type 1 / Q000378:metabolism; D018342:Receptors, Adrenergic, beta-1 / Q000235:genetics / Q000378:metabolism; D018078:Receptors, Prostaglandin E / Q000235:genetics / Q000378:metabolism; D020744:rac GTP-Binding Proteins / Q000037:antagonists & inhibitors* / Q000378:metabolism
chemical_list: D000199:Actins; C526292:IRS1 protein, human; D055504:Insulin Receptor Substrate Proteins; C526295:Irs1 protein, mouse; D010750:Phosphoproteins; D010982:Platelet-Derived Growth Factor; D018342:Receptors, Adrenergic, beta-1; D018078:Receptors, Prostaglandin E; D007334:Insulin-Like Growth Factor I; D000242:Cyclic AMP; D019869:Phosphatidylinositol 3-Kinases; D017526:Receptor, IGF Type 1; D017868:Cyclic AMP-Dependent Protein Kinases; D020962:Heterotrimeric GTP-Binding Proteins; D020744:rac GTP-Binding Proteins; D015232:Dinoprostone