A Cellular Assay for Measuring the Inhibition of Glycogen Synthase Kinase-3 via the Accumulation of β-Catenin in Chinese Hamster Ovary Clone K1 Cells

2006 ◽  
Vol 4 (4) ◽  
pp. 451-460 ◽  
Author(s):  
Karen Yeow ◽  
Laurence Novo-Perez ◽  
Pascale Gaillard ◽  
Patrick Page ◽  
Jean-Pierre Gotteland ◽  
...  
Keyword(s):  
Chinese Hamster Ovary ◽  
Glycogen Synthase ◽  
Chinese Hamster ◽  
Glycogen Synthase Kinase ◽  
Glycogen Synthase Kinase 3 ◽  
Cellular Assay

scholarly journals Glycogen synthase kinase-3 is rapidly inactivated in response to insulin and phosphorylates eukaryotic initiation factor eIF-2B

1993 ◽  
Vol 294 (3) ◽  
pp. 625-629 ◽  
Author(s):  
G I Welsh ◽  
C G Proud
Keyword(s):  
Kinase Activity ◽  
Glycogen Synthase ◽  
Chinese Hamster Ovary Cells ◽  
Chinese Hamster ◽  
Glycogen Synthase Kinase ◽  
Initiation Factor ◽  
Glycogen Synthase Kinase 3 ◽  
Eukaryotic Initiation Factor ◽  
Nucleotide Exchange ◽  
Cell Extracts

We have studied the control of insulin-regulated protein kinases in Chinese hamster ovary cells transfected with the human insulin receptor (CHO.T cells). Among these enzymes is one that is obtained after chromatography of cell extracts on Mono-S, whose activity is decreased (7.3 +/- 1.9-fold) within 10 min of insulin treatment. This enzyme phosphorylates glycogen synthase and the largest subunit of protein synthesis eukaryotic initiation factor (eIF)-2B (the guanine nucleotide exchange factor). The kinase appears to be glycogen synthase kinase-3 (GSK-3), on the basis of: (1) its ability to phosphorylate a peptide based on the phosphorylation sites for GSK-3 in glycogen synthase, and (2) the finding that the fractions possessing this activity contain immunoreactive GSK-3, whose peak is coincident with that of kinase activity, as judged by immunoblotting using antibodies specific for the alpha- and beta-isoforms of GSK-3. The decrease in kinase activity induced by insulin was reversed by treatment of the column fractions with protein phosphatase-2A. These data indicate that insulin rapidly causes inactivation of GSK-3 and that this is due to phosphorylation of GSK-3. The implications of these findings for the control of glycogen and protein metabolism are discussed.

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