Alleviating the suppression of glycogen synthase kinase-3beta by Akt leads to the phosphorylation of cAMP-response element-binding protein and its transactivation in intact cell nuclei.
Glycogen synthase kinase-3beta (GSK-3beta) activity is suppressed when it becomes phosphorylated on serine 9 by protein kinase B (Akt). To determine how GSK-3beta activity opposes Akt function we used various methods to alleviate GSK-3beta suppression in prostate carcinoma cells. In some experiments, LY294002, a specific inhibitor of phosphatidylinositol 3-kinase (a kinase involved in activating Akt) and tumor necrosis factor-alpha (TNF-alpha) were used to activate GSK-3beta. In other experiments mutant forms of GSK-3beta, GSK-3betadelta9 (a constitutively active deletion mutant of GSK-3beta) and GSK-3betaY216F (an inactive point mutant of GSK-3beta) were used to alter GSK-3beta activity. LY294002, TNF-alpha, and overexpression of wild-type GSK-3beta or of GSK-3betadelta9, but not GSK-3betaY216F, alleviated the suppression of GSK-3beta activity in prostate carcinoma cells and enhanced the turnover of beta-catenin. Forced expression of wild-type GSK-3beta or of GSK-3betadelta9, but not GSK-3betaY216F, suppressed cell growth and showed that the phosphorylation status of GSK-3beta can affect its intracellular distribution. When transcription factors activator protein-1 and cyclic AMP-response element (CRE)-binding protein were analyzed as targets of GSK-3beta activity, overexpression of wild-type GSK-3beta suppressed AP1-mediated transcription and activated CRE-mediated transcription. Overexpression of GSK-3betadelta9 caused an (80-fold) increase in CRE-mediated transcription, which was further amplified (up to 130-fold) by combining GSK-3betadelta9 overexpression with the suppression of Jun activity. This study also demonstrated for the first time that expression of constitutively active GSK-3betadelta9 results in the phosphorylation of CRE-binding protein on serine 129 and enhancement of CRE-mediated transcription in intact cell nuclei.
Salas
Thomas R
TR
Departments of Clinical Cancer Prevention and Gastrointestinal Medical Oncology, The University of Texas M D Anderson Cancer Center, Houston, Texas 77030, USA.
Reddy
Shrikanth A
SA
Clifford
John L
JL
Davis
Roger J
RJ
Kikuchi
Akira
A
Lippman
Scott M
SM
Menter
David G
DG
D015153:Blotting, Western; D002455:Cell Division; D045744:Cell Line, Tumor; D002467:Cell Nucleus / Q000378:metabolism*; D002867:Chromones / Q000494:pharmacology; D017362:Cyclic AMP Response Element-Binding Protein / Q000378:metabolism; D018076:DNA, Complementary / Q000378:metabolism; D003720:Densitometry; D004791:Enzyme Inhibitors / Q000494:pharmacology; D038362:Glycogen Synthase Kinase 3 / Q000037:antagonists & inhibitors* / Q000378:metabolism*; D000071679:Glycogen Synthase Kinase 3 beta; D006801:Humans; D008156:Luciferases / Q000378:metabolism; D008856:Microscopy, Fluorescence; D008958:Models, Molecular; D009025:Morpholines / Q000494:pharmacology; D009154:Mutation; D010455:Peptides / Q000737:chemistry; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases; D017434:Protein Structure, Tertiary; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D051057:Proto-Oncogene Proteins c-akt; D012694:Serine / Q000737:chemistry; D013997:Time Factors; D018808:Transcription Factor AP-1 / Q000378:metabolism; D014158:Transcription, Genetic; D015533:Transcriptional Activation; D014162:Transfection; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism
D002867:Chromones; D017362:Cyclic AMP Response Element-Binding Protein; D018076:DNA, Complementary; D004791:Enzyme Inhibitors; D038362:Glycogen Synthase Kinase 3; D000071679:Glycogen Synthase Kinase 3 beta; D008156:Luciferases; D009025:Morpholines; D010455:Peptides; D019869:Phosphatidylinositol 3-Kinases; D017346:Protein Serine-Threonine Kinases; D011518:Proto-Oncogene Proteins; D051057:Proto-Oncogene Proteins c-akt; D012694:Serine; D018808:Transcription Factor AP-1; D014409:Tumor Necrosis Factor-alpha; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; C494918:AKT1 protein, human; C000605506:GSK3B protein, human