title: Regulation of brain-type creatine kinase by AMP-activated protein kinase: interaction, phosphorylation and ER localization.
abstract: AMP-activated protein kinase (AMPK) and cytosolic brain-type creatine kinase (BCK) cooperate under energy stress to compensate for loss of adenosine triphosphate (ATP) by either stimulating ATP-generating and inhibiting ATP-consuming pathways, or by direct ATP regeneration from phosphocreatine, respectively. Here we report on AMPK-dependent phosphorylation of BCK from different species identified by in vitro screening for AMPK substrates in mouse brain. Mass spectrometry, protein sequencing, and site-directed mutagenesis identified Ser6 as a relevant residue with one site phosphorylated per BCK dimer. Yeast two-hybrid analysis revealed interaction of active AMPK specifically with non-phosphorylated BCK. Pharmacological activation of AMPK mimicking energy stress led to BCK phosphorylation in astrocytes and fibroblasts, as evidenced with a highly specific phospho-Ser6 antibody. BCK phosphorylation at Ser6 did not affect its enzymatic activity, but led to the appearance of the phosphorylated enzyme at the endoplasmic reticulum (ER), close to the ER calcium pump, a location known for muscle-type cytosolic creatine kinase (CK) to support Ca²⁺-pumping.
authors.affiliation: Univ. Grenoble Alpes, Laboratory of Fundamental and Applied Bioenergetics, Grenoble, France; Inserm, U1055, Grenoble, France.
authors.forename: Sacnicte
authors.initials: S
authors.lastname: Ramírez Ríos
authors.affiliation: Univ. Grenoble Alpes, Laboratory of Fundamental and Applied Bioenergetics, Grenoble, France; Inserm, U1055, Grenoble, France.
authors.forename: Frédéric
authors.initials: F
authors.lastname: Lamarche
authors.affiliation: Univ. Grenoble Alpes, Laboratory of Fundamental and Applied Bioenergetics, Grenoble, France; Inserm, U1055, Grenoble, France.
authors.forename: Cécile
authors.initials: C
authors.lastname: Cottet-Rousselle
authors.affiliation: Univ. Grenoble Alpes, Laboratory of Fundamental and Applied Bioenergetics, Grenoble, France; Inserm, U1055, Grenoble, France.
authors.forename: Anna
authors.initials: A
authors.lastname: Klaus
authors.affiliation: Institute of Cell Biology, ETH Zurich, Switzerland.
authors.forename: Roland
authors.initials: R
authors.lastname: Tuerk
authors.affiliation: Institute of Cell Biology, ETH Zurich, Switzerland.
authors.forename: Ramon
authors.initials: R
authors.lastname: Thali
authors.affiliation: Functional Genomics Center Zurich, ETH Zurich/University of Zurich, Switzerland.
authors.forename: Yolanda
authors.initials: Y
authors.lastname: Auchli
authors.affiliation: Functional Genomics Center Zurich, ETH Zurich/University of Zurich, Switzerland.
authors.forename: René
authors.initials: R
authors.lastname: Brunisholz
authors.affiliation: Institute of Cell Biology, ETH Zurich, Switzerland.
authors.forename: Dietbert
authors.initials: D
authors.lastname: Neumann
authors.affiliation: Univ. Grenoble Alpes, Laboratory of Fundamental and Applied Bioenergetics, Grenoble, France; Inserm, U1055, Grenoble, France.
authors.forename: Luc
authors.initials: L
authors.lastname: Barret
authors.affiliation: Univ. Grenoble Alpes, Laboratory of Fundamental and Applied Bioenergetics, Grenoble, France; Inserm, U1055, Grenoble, France.
authors.forename: Malgorzata
authors.initials: M
authors.lastname: Tokarska-Schlattner
authors.affiliation: Univ. Grenoble Alpes, Laboratory of Fundamental and Applied Bioenergetics, Grenoble, France; Inserm, U1055, Grenoble, France. Electronic address: uwe.schlattner@ujf-grenoble.fr.
authors.forename: Uwe
authors.initials: U
authors.lastname: Schlattner
mesh_terms: D055372:AMP-Activated Protein Kinases / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D001253:Astrocytes / Q000378:metabolism / Q000648:ultrastructure; D001921:Brain / Q000201:enzymology* / Q000648:ultrastructure; D003402:Creatine Kinase / Q000235:genetics / Q000378:metabolism*; D003600:Cytosol / Q000378:metabolism; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D051379:Mice; D009097:Multienzyme Complexes / Q000378:metabolism; D016297:Mutagenesis, Site-Directed; D010766:Phosphorylation; D012694:Serine / Q000378:metabolism
chemical_list: D009097:Multienzyme Complexes; D012694:Serine; D055372:AMP-Activated Protein Kinases; D003402:Creatine Kinase
keywords: Creatine kinase; Endoplasmic reticulum; Energy homeostasis; Protein kinase; Protein phosphorylation; Subcellular localization