A dominant role for the Raf-MEK pathway in forskolin, 12-O-tetradecanoyl-phorbol acetate, and platelet-derived growth factor-induced CREB (cAMP-responsive element-binding protein) activation, uncoupled from serine 133 phosphorylation in NIH 3T3 cells.
In this study we describe that platelet-derived growth factor (PDGF), 12-O-tetradecanoyl-phorbol-acetate (TPA), and forskolin induced CREB (cAMP-responsive element-binding protein) Ser-133 phosphorylation with comparable magnitude and kinetics in NIH 3T3 cells. While forskolin was the most potent activator of CREB, TPA or PDGF modestly increased CREB activity. The role of protein kinase C, protein kinase A, and the Raf-MEK kinase pathway in the activation and Ser-133 phosphorylation of CREB by these three stimuli was investigated. We found that inhibition of the Raf-MEK kinase pathway efficiently blocks transcriptional activation of CREB by all three stimuli. This dominant involvement of Raf-MEK in CREB transcriptional activation seems to be uncoupled from CREB Ser-133 phosphorylation. We further demonstrate that although inhibition of Raf-MEK represses forskolin-induced CREB activation, forskolin by itself failed to activate ERK1/2 and Elk-1 mediated transcription. These results suggest that a basal level of Raf-MEK activity is necessary for both PDGF- and forskolin-induced CREB activation, independent of CREB Ser-133 phosphorylation.
Department of Gene Biology, Institute of Medical Biology, University of Tromsø, Norway.
O M
OM
Seternes
B
B
Johansen
U
U
Moens
D016475:3T3 Cells / Q000187:drug effects / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000187:drug effects / Q000378:metabolism; D018797:Cell Cycle Proteins; D005576:Colforsin / Q000378:metabolism / Q000494:pharmacology; D017362:Cyclic AMP Response Element-Binding Protein / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D017868:Cyclic AMP-Dependent Protein Kinases / Q000187:drug effects / Q000378:metabolism; D004268:DNA-Binding Proteins; D054638:Dual Specificity Phosphatase 1; D054642:Dual Specificity Phosphatase 6; D004789:Enzyme Activation / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005419:Flavonoids / Q000494:pharmacology; D017874:Immediate-Early Proteins / Q000187:drug effects / Q000235:genetics / Q000378:metabolism; D048728:MAP Kinase Kinase Kinase 1; D051379:Mice; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases; D008969:Molecular Sequence Data; D010749:Phosphoprotein Phosphatases; D010766:Phosphorylation; D010982:Platelet-Derived Growth Factor / Q000378:metabolism / Q000494:pharmacology; D011493:Protein Kinase C / Q000187:drug effects / Q000378:metabolism; D054645:Protein Phosphatase 1; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D017027:Protein Tyrosine Phosphatases / Q000187:drug effects / Q000235:genetics / Q000378:metabolism; D011518:Proto-Oncogene Proteins / Q000187:drug effects / Q000235:genetics / Q000378:metabolism; D019908:Proto-Oncogene Proteins c-raf / Q000378:metabolism; D012694:Serine / Q000378:metabolism; D013755:Tetradecanoylphorbol Acetate / Q000378:metabolism / Q000494:pharmacology; D014157:Transcription Factors; D015533:Transcriptional Activation; D050807:ets-Domain Protein Elk-1
D018797:Cell Cycle Proteins; D017362:Cyclic AMP Response Element-Binding Protein; D004268:DNA-Binding Proteins; C494113:Elk1 protein, mouse; D004791:Enzyme Inhibitors; D005419:Flavonoids; D017874:Immediate-Early Proteins; D010982:Platelet-Derived Growth Factor; D011518:Proto-Oncogene Proteins; D014157:Transcription Factors; D050807:ets-Domain Protein Elk-1; D005576:Colforsin; D012694:Serine; D017346:Protein Serine-Threonine Kinases; D019908:Proto-Oncogene Proteins c-raf; D017868:Cyclic AMP-Dependent Protein Kinases; D011493:Protein Kinase C; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases; D048728:MAP Kinase Kinase Kinase 1; C482578:Map3k1 protein, mouse; D010749:Phosphoprotein Phosphatases; D054645:Protein Phosphatase 1; D054638:Dual Specificity Phosphatase 1; D054642:Dual Specificity Phosphatase 6; C517020:Dusp1 protein, mouse; C517036:Dusp6 protein, mouse; D017027:Protein Tyrosine Phosphatases; D013755:Tetradecanoylphorbol Acetate; C093973:2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one