title: Importance of autophosphorylation at Ser186 in the A-loop of salt inducible kinase 1 for its sustained kinase activity.
abstract: Autophosphorylation is an important mechanism by which protein kinases regulate their own biological activities. Salt inducible kinase 1 (SIK1) is a regulator in the feedback cascades of cAMP-mediated gene expression, while its kinase domain also features autophosphorylation activity. We provide evidence that Ser186 in the activation loop is the site of autophosphorylation and essential for the kinase activity. Ser186 is located at the +4 position of the critical Thr residue Thr182, which is phosphorylated by upstream kinases such as LKB1. The relationship between phosphorylation at Ser186 and at Thr182 in COS-7 cells indicates that the former is a prerequisite for the latter. Glycogen synthase kinase-3beta (GSK-3beta) phosphorylates Ser/Thr residues located at the fourth position ahead of the pre-phosphorylated Ser/Thr residues, and inhibitors of GSK-3beta reduce the phosphorylation at Thr182. The results of an in vitro reconstitution assay also indicate that GSK-3beta could be the SIK1 kinase. However, overexpression and knockdown of GSK-3beta in LKB1-defective HeLa cells suggests that GSK-3beta alone may not be able to phosphorylate or activate SIK1, indicating that LKB1 may play a crucial role by phosphorylating SIK1 at Thr182, possibly as an initiator of the autophosphorylation cascade, and GSK-3beta may phosphorylate SIK1 at Thr182 by recognizing the priming-autophosphorylation at Ser186 in cultured cells. This may also be the case for the other isoform SIK2, but not for SIK3.
authors.affiliation: Laboratory of Cell Signaling and Metabolism, National Institute of Biomedical Innovation, Osaka 567-0085, Japan.
authors.forename: Yoshiko Katoh
authors.initials: YK
authors.lastname: Hashimoto
authors.forename: Tomohiro
authors.initials: T
authors.lastname: Satoh
authors.forename: Mitsuhiro
authors.initials: M
authors.lastname: Okamoto
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Takemori
mesh_terms: D048948:14-3-3 Proteins / Q000378:metabolism; D055372:AMP-Activated Protein Kinases; D000818:Animals; D019556:COS Cells; D002522:Chlorocebus aethiops; D038362:Glycogen Synthase Kinase 3 / Q000378:metabolism; D000071679:Glycogen Synthase Kinase 3 beta; D006367:HeLa Cells; D006801:Humans; D007211:Indoles / Q000494:pharmacology; D051379:Mice; D010091:Oximes / Q000494:pharmacology; D010766:Phosphorylation / Q000187:drug effects; D010768:Phosphoserine / Q000378:metabolism*; D010769:Phosphothreonine / Q000378:metabolism; D011485:Protein Binding / Q000187:drug effects; D017346:Protein Serine-Threonine Kinases / Q000737:chemistry / Q000378:metabolism*; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D051381:Rats
chemical_list: D048948:14-3-3 Proteins; D007211:Indoles; D010091:Oximes; C474021:indirubin-3'-monoxime; D010769:Phosphothreonine; D010768:Phosphoserine; C474808:salt-inducible kinase-2, mouse; C000605506:GSK3B protein, human; D000071679:Glycogen Synthase Kinase 3 beta; C000605507:Gsk3b protein, mouse; C000605508:Gsk3b protein, rat; D017346:Protein Serine-Threonine Kinases; C120299:Sik1 protein, rat; C491654:Stk11 protein, mouse; D038362:Glycogen Synthase Kinase 3; D055372:AMP-Activated Protein Kinases; C064758:Ro 31-8220