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

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.

[1]  R. Rhoads.  Regulation of eukaryotic protein synthesis by initiation factors. , 1993, The Journal of biological chemistry.

[2]  J. Woodgett,et al.  Modulation of the glycogen synthase kinase‐3 family by tyrosine phosphorylation. , 1993, The EMBO journal.

[3]  C. Proud,et al.  Evidence for a role for protein kinase C in the stimulation of protein synthesis by insulin in swiss 3T3 fibroblasts , 1993, FEBS letters.

[4]  J. Woodgett,et al.  Identification of multifunctional ATP-citrate lyase kinase as the alpha-isoform of glycogen synthase kinase-3. , 1992, The Biochemical journal.

[5]  J. Tavaré,et al.  Characterization of insulin-stimulated protein serine/threonine kinases in CHO cells expressing human insulin receptors with point and deletion mutations. , 1992, The Biochemical journal.

[6]  J. Woodgett,et al.  Differential regulation of glycogen synthase kinase-3 beta by protein kinase C isotypes. , 1992, The Journal of biological chemistry.

[7]  S. Oldfield,et al.  Purification, phosphorylation and control of the guanine-nucleotide-exchange factor from rabbit reticulocyte lysates. , 1992, European journal of biochemistry.

[8]  C. Proud,et al.  Regulation of protein synthesis in Swiss 3T3 fibroblasts. Rapid activation of the guanine-nucleotide-exchange factor by insulin and growth factors. , 1992, The Biochemical journal.

[9]  J. Woodgett,et al.  Baculovirus-mediated expression and characterisation of rat glycogen synthase kinase-3 beta, the mammalian homologue of the Drosophila melanogaster zeste-white 3sgg homeotic gene product. , 1992, European journal of biochemistry.

[10]  C. Proud,et al.  Protein phosphorylation in translational control. , 1992, Current topics in cellular regulation.

[11]  P. Cohen,et al.  The molecular mechanism by which adrenalin inhibits glycogen synthesis. , 1991, European journal of biochemistry.

[12]  D. Stumpo,et al.  Insulin activation of protein kinase C: a reassessment. , 1991, The Journal of biological chemistry.

[13]  Tony Hunter,et al.  Activation of protein kinase C decreases phosphorylation of c-Jun at sites that negatively regulate its DNA-binding activity , 1991, Cell.

[14]  P. Dent,et al.  The molecular mechanism by which insulin stimulates glycogen synthesis in mammalian skeletal muscle , 1990, Nature.

[15]  P. Simpson,et al.  An early embryonic product of the gene shaggy encodes a serine/threonine protein kinase related to the CDC28/cdc2+ subfamily. , 1990, The EMBO journal.

[16]  J. Woodgett,et al.  Molecular cloning and expression of glycogen synthase kinase‐3/factor A. , 1990, The EMBO journal.

[17]  Norbert Perrimon,et al.  Putative protein kinase product of the Drosophila segment-polarity gene zeste-white3 , 1990, Nature.

[18]  D. Buckley,et al.  Effects of Insulin on Diacylglycerol–Protein Kinase C Signaling in Rat Diaphragm and Soleus Muscles and Relationship to Glucose Transport , 1990, Diabetes.

[19]  E. Villa-Moruzzi.  Stimulation of FA and phosphatase‐1 activities by insulin in 3T3‐L1 cells , 1989, FEBS letters.

[20]  R. Farese,et al.  Potential role of phospholipid-signaling systems in insulin action and states of clinical insulin resistance. , 1989, Diabetes/metabolism reviews.

[21]  S. Yang,et al.  Insulin induces activation of kinase FA in membranes and thereby promotes activation of ATP.Mg-dependent phosphatase in adipocytes. , 1989, Biochemical and biophysical research communications.

[22]  K. S. Murthy,et al.  Effect of insulin on ATP-citrate lyase phosphorylation: regulation of peptide A and peptide B phosphorylations. , 1989, Biochemistry.

[23]  Blackshear Pj.  Insulin-stimulated protein biosynthesis as a paradigm of protein kinase C-independent growth factor action. , 1989 .

[24]  W. Benjamin,et al.  Insulin action rapidly decreases multifunctional protein kinase activity in rat adipose tissue. , 1988, The Journal of biological chemistry.

[25]  B. Gibson,et al.  Analysis of the in vivo phosphorylation state of rabbit skeletal muscle glycogen synthase by fast-atom-bombardment mass spectrometry. , 1988, European journal of biochemistry.

[26]  P. Roach,et al.  Formation of protein kinase recognition sites by covalent modification of the substrate. Molecular mechanism for the synergistic action of casein kinase II and glycogen synthase kinase 3. , 1987, The Journal of biological chemistry.

[27]  R. Farese,et al.  Insulin increases membrane and cytosolic protein kinase C activity in BC3H-1 myocytes. , 1987, The Journal of biological chemistry.

[28]  W. Benjamin,et al.  Cyclic nucleotide-independent protein kinase from rat liver. Purification and characterization of a multifunctional protein kinase. , 1985, The Journal of biological chemistry.

[29]  Philip R. Cohen,et al.  Multisite phosphorylation of glycogen synthase. Molecular basis for the substrate specificity of glycogen synthase kinase-3 and casein kinase-II (glycogen synthase kinase-5). , 1984, Biochimica et biophysica acta.

[30]  P. Cohen,et al.  Glycogen synthase kinase-3 from rabbit skeletal muscle. , 2005, Methods in enzymology.

[31]  Philip R. Cohen,et al.  Phosphorylation of the type-II regulatory subunit of cyclic-AMP-dependent protein kinase by glycogen synthase kinase 3 and glycogen synthase kinase 5. , 2005, European journal of biochemistry.

[32]  P. Cohen,et al.  Purification of glycogen synthase kinase 3 from rabbit skeletal muscle. Copurification with the activating factor (FA) of the (Mg-ATP) dependent protein phosphatase. , 1981, European journal of biochemistry.

[33]  W. Merlevede,et al.  ATP x Mg-dependent protein phosphatase from rabbit skeletal muscle. II. Purification of the activating factor and its characterization as a bifunctional protein also displaying synthase kinase activity. , 1980, The Journal of biological chemistry.