Thyroid-stimulating hormone and cyclic AMP activate p38 mitogen-activated protein kinase cascade. Involvement of protein kinase A, rac1, and reactive oxygen species.
p38 mitogen-activated protein kinases (p38-MAPKs) are activated by cytokines, cellular stresses, growth factors, and hormones. We show here that p38-MAPKs are activated upon stimulation by thyroid-stimulating hormone (TSH) or cAMP. TSH caused the phosphorylation of p38-MAPK in Chinese hamster ovary cells stably transfected with the human TSH receptor but not in wild-type Chinese hamster ovary cells. The effect of TSH was fully mimicked by the adenylyl cyclase activator, forskolin, and by a permeant analog of cAMP. The effect of forskolin was reproduced in FRTL5 rat thyroid cells. TSH also stimulated the phosphorylation of MAPK kinase 3 or 6, over the same time scale as that of p38-MAPKs. TSH and forskolin stimulated the activity of the alpha-isoform of p38-MAPK assayed by phosphorylation of the transcription factor ATF2. The activity of MAPK-activated protein kinase-2 was stimulated by TSH and forskolin. This stimulation was abolished by SB203580, a specific inhibitor of p38-MAPKs. The protein kinase A inhibitor H89 inhibited the stimulation of phosphorylation of p38-MAPKs by forskolin, whereas inhibitors of protein kinase C, p70(S6k), and phosphatidylinositol 3-kinase were ineffective. Expression of the dominant negative form of Rac1, but not that of Ras, blocked forskolin-induced p38-MAPK activation. Diphenylene iodonium, a potent inhibitor of NADPH oxidase(s), and ascorbic acid, an effective free radical scavenger, suppressed TSH- or forskolin-stimulated p38-MAPK phosphorylation, indicating that the generation of reactive oxygen species plays a key role in signaling from cAMP to p38-MAPKs. Inhibition of the p38-MAPK pathway with SB203580 partially but significantly, attenuates cAMP- and TSH-induced expression of the sodium iodide symporter in FRTL-5 cells. These results point to a new signaling pathway for the G(s)-coupled TSH receptor, involving cAMP, protein kinase A, Rac1, and reactive oxygen species and resulting in the activation of a signaling kinase cascade that includes MAPK kinase 3 or 6, p38-MAPK, and MAPK-activated protein kinase-2.
Pomerance
M
M
Unité 486 INSERM, Transduction Hormonale et Régulation Cellulaire, Faculté de Pharmacie, 92296 Châtenay-Malabry, France. martine.pomerance@cep.u-psud.fr
Abdullah
H B
HB
Kamerji
S
S
Correze
C
C
Blondeau
J P
JP
D000818:Animals; D016466:CHO Cells; D002460:Cell Line; D006224:Cricetinae; D000242:Cyclic AMP / Q000502:physiology*; D017868:Cyclic AMP-Dependent Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D007211:Indoles / Q000494:pharmacology; D007546:Isoquinolines / Q000494:pharmacology; D008301:Maleimides / Q000494:pharmacology; D020928:Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D010766:Phosphorylation; D051381:Rats; D017382:Reactive Oxygen Species / Q000378:metabolism*; D013449:Sulfonamides; D013972:Thyrotropin / Q000502:physiology*; D048051:p38 Mitogen-Activated Protein Kinases; D020830:rac1 GTP-Binding Protein / Q000378:metabolism*
D004791:Enzyme Inhibitors; D007211:Indoles; D007546:Isoquinolines; D008301:Maleimides; D017382:Reactive Oxygen Species; D013449:Sulfonamides; D013972:Thyrotropin; D000242:Cyclic AMP; D017868:Cyclic AMP-Dependent Protein Kinases; D020928:Mitogen-Activated Protein Kinases; D048051:p38 Mitogen-Activated Protein Kinases; D020830:rac1 GTP-Binding Protein; C070515:bisindolylmaleimide I; C063509:N-(2-(4-bromocinnamylamino)ethyl)-5-isoquinolinesulfonamide