Phosphorylation of Ser640 in muscle glycogen synthase by DYRK family protein kinases.
Glycogen synthase, a key enzyme in the regulation of glycogen synthesis by insulin, is controlled by multisite phosphorylation. Glycogen synthase kinase-3 (GSK-3) phosphorylates four serine residues in the COOH terminus of glycogen synthase. Phosphorylation of one of these residues, Ser(640) (site 3a), causes strong inactivation of glycogen synthase. In previous work, we demonstrated in cell models that site 3a can be phosphorylated by an as yet unidentified protein kinase (3a-kinase) distinct from GSK-3. In the present study, we purified the 3a-kinase from rabbit skeletal muscle and identified one constituent polypeptide as HAN11, a WD40 domain protein with unknown function. Another polypeptide was identified as DYRK1A, a member of the dual-specificity tyrosine phosphorylated and regulated protein kinase (DYRK) family. Two isoforms of DYRK, DYRK1A and DYRK1B, co-immunoprecipitate with HAN11 when coexpressed in COS cells indicating that the proteins interact in mammalian cells. Co-expression of DYRK1A, DYRK1B, or DYRK2 with a series of glycogen synthase mutants with Ser/Ala substitutions at the phosphorylation sites in COS cells revealed that protein kinases cause phosphorylation of site 3a in glycogen synthase. To confirm that DYRKs directly phosphorylate glycogen synthase, recombinant DYRK1A, DYRK2, and glycogen synthase were produced in bacterial cells. In the presence of Mg-ATP, both DYRKs inactivated glycogen synthase by more than 10-fold. The inactivation correlated with phosphorylation of site 3a in glycogen synthase. These results indicate that protein kinase(s) from the DYRK family may be involved in a new mechanism for the regulation of glycogen synthesis.
Department of Biochemistry and Molecular Biology, Indiana University School of Medicine, Indianapolis, Indiana 46202, USA. askurat@iupui.edu
Alexander V
AV
Skurat
Amy D
AD
Dietrich
D000818:Animals; D005347:Fibroblasts / Q000378:metabolism; D006003:Glycogen / Q000096:biosynthesis*; D006006:Glycogen Synthase / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D018482:Muscle, Skeletal / Q000201:enzymology; D009154:Mutation; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D011817:Rabbits; D051381:Rats; D012694:Serine / Q000378:metabolism; D000097603:Dyrk Kinases
D006003:Glycogen; D006006:Glycogen Synthase; D017346:Protein Serine-Threonine Kinases; D011505:Protein-Tyrosine Kinases; D012694:Serine; D000097603:Dyrk Kinases