ERK1/2 control phosphorylation and protein level of cAMP-responsive element-binding protein: a key role in glucose-mediated pancreatic beta-cell survival.
cAMP-responsive element-binding protein (CREB) is required for beta-cell survival by regulating expression of crucial genes such as bcl-2 and IRS-2. Using MIN6 cells and isolated rat pancreatic islets, we investigated the signaling pathway that controls phosphorylation and protein level of CREB. We observed that 10 mmol/l glucose-induced CREB phosphorylation was totally inhibited by the protein kinase A (PKA) inhibitor H89 (2 micromol/l) and reduced by 50% with the extracellular signal-regulated kinase (ERK)1/2 inhibitor PD98059 (20 micromol/l). This indicates that ERK1/2, reported to be located downstream of PKA, participates in the PKA-mediated CREB phosphorylation elicited by glucose. In ERK1/2-downregulated MIN6 cells by siRNA, glucose-stimulated CREB phosphorylation was highly reduced and CREB protein content was decreased by 60%. In MIN6 cells and islets cultured for 24-48 h in optimal glucose concentration (10 mmol/l), which promotes survival, blockade of ERK1/2 activity with PD98059 caused a significant decrease in CREB protein level, whereas CREB mRNA remained unaffected (measured by real-time quantitative PCR). This was associated with loss of bcl-2 mRNA and protein contents, caspase-3 activation, and emergence of ultrastructural apoptotic features detected by electron microscopy. Our results indicate that ERK1 and -2 control the phosphorylation and protein level of CREB and play a key role in glucose-mediated pancreatic beta-cell survival.
Costes
Safia
S
INSERM U661, Equipe Avenir, Institut de Génomique Fonctionnelle, 141, rue de la cardonille, 34094 Montpellier Cedex 5, France.
Broca
Christophe
C
Bertrand
Gyslaine
G
Lajoix
Anne-Dominique
AD
Bataille
Dominique
D
Bockaert
Joel
J
Dalle
Stéphane
S
D000818:Animals; D017209:Apoptosis; D050882:CREB-Binding Protein / Q000032:analysis* / Q000378:metabolism*; D002460:Cell Line; D002470:Cell Survival; D017868:Cyclic AMP-Dependent Protein Kinases / Q000037:antagonists & inhibitors / Q000502:physiology; D005419:Flavonoids / Q000494:pharmacology; D015870:Gene Expression; D019254:Genes, bcl-2 / Q000235:genetics; D005947:Glucose / Q000494:pharmacology; D007515:Islets of Langerhans / Q000502:physiology* / Q000648:ultrastructure; D007546:Isoquinolines / Q000494:pharmacology; D008297:Male; D046529:Microscopy, Electron, Transmission; D019950:Mitogen-Activated Protein Kinase 1 / Q000037:antagonists & inhibitors / Q000235:genetics / Q000502:physiology*; D048052:Mitogen-Activated Protein Kinase 3 / Q000037:antagonists & inhibitors / Q000235:genetics / Q000502:physiology*; D010766:Phosphorylation; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000032:analysis; D012333:RNA, Messenger / Q000032:analysis; D034741:RNA, Small Interfering / Q000235:genetics; D051381:Rats; D017208:Rats, Wistar; D013449:Sulfonamides / Q000494:pharmacology; D014162:Transfection
D005419:Flavonoids; D007546:Isoquinolines; D047428:Protein Kinase Inhibitors; D019253:Proto-Oncogene Proteins c-bcl-2; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D013449:Sulfonamides; D050882:CREB-Binding Protein; C494365:Crebbp protein, rat; D017868:Cyclic AMP-Dependent Protein Kinases; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D005947:Glucose; C063509:N-(2-(4-bromocinnamylamino)ethyl)-5-isoquinolinesulfonamide; C093973:2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one