Alleviating the Suppression of Glycogen Synthase Kinase-3β by Akt Leads to the Phosphorylation of cAMP-response Element-binding Protein and Its Transactivation in Intact Cell Nuclei.

Glycogen synthase kinase-3β (GSK-3β) activity is suppressed when it becomes phosphorylated on serine 9 by protein kinase B (Akt). To determine how GSK-3β activity opposes Akt function we used various methods to alleviate GSK-3β 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-α (TNF-α) were used to activate GSK-3β. In other experiments mutant forms of GSK-3β, GSK-3β[sup Δ9] (a constitutively active deletion mutant of GSK-3β) and GSK-3β[sup Y216F] (an inactive point mutant of GSK-3β) were used to alter GSK-3β activity. LY294002, TNF-α, and overexpression of wild-type GSK-3β or of GSK-3β[sup Δ9], but not GSK-3β[sup Y216F], alleviated the suppression of GSK-3β activity in prostate carcinoma cells and enhanced the turnover of β-catenin. Forced expression of wild-type GSK-3β or of GSK-3β[sup Δ9], but not GSK3β[sup Y216F], suppressed cell growth and showed that the phosphorylation status of GSK-3β 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-3β activity, overexpression of wild-type GSK-3β suppressed AP1-mediated transcription and activated CRE-mediated transcription. Overexpression of GSK-3β[sup Δ9] caused an (80-fold) increase in CRE-mediated transcription, which was further amplifled (up to 130-fold) by combining GSK-3β[sup Δ9] overexpression with the suppression of Jun activity. This study also demonstrated for the first time that expression of constitutively active GSK-3β[sup Δ9] results in the phosphorylation of CRE-binding protein on serine 129 and enhancement of CRE-mediated transcription in intact cell nuclei. [ABSTRACT FROM AUTHOR]