AMP-activated protein kinase (AMPK) is a metabolic stress-sensing kinase. We previously showed that glucose deprivation induces autophosphorylation of AMPKβ at Thr-148, which prevents the binding of AMPK to glycogen. Furthermore, in MIN6 cells, AMPKβ1 binds to R6 (PPP1R3D), a glycogen-targeting subunit of protein phosphatase type 1 (PP1), thereby regulating the glucose-induced inactivation of AMPK. In the present study, we further investigated the interaction of R6 with AMPKβ and the possible dependency on Thr-148 phosphorylation status. Yeast two-hybrid (Y2H) analyses and co-immunoprecipitation (IP) of the overexpressed proteins in human embryonic kidney (HEK) 293T) cells revealed that both AMPKβ1 and AMPK-β2 wild-type (WT) isoforms bind to R6. The AMPKβ–R6 interaction was stronger with the muscle-specific AMPKβ2-WT and required association with the substrate-binding motif of R6. When HEK293T cells or C2C12 myotubes were cultured in high-glucose medium, AMPKβ2-WT and R6 weakly interacted. In contrast, glycogen depletion significantly enhanced this protein interaction. Mutation of AMPKβ2 Thr-148 prevented the interaction with R6 irrespective of the intracellular glycogen content. Treatment with the AMPK activator oligomycin enhanced the AMPKβ2–R6 interaction in conjunction with increased Thr-148 phosphorylation in cells grown in low-glucose medium. These data are in accordance with R6 binding directly to AMPKβ2 when both proteins detach from the diminishing glycogen particle, which is simultaneous with increased AMPKβ2 Thr-148 autophosphorylation. Such a model points to a possible control of AMPK by PP1-R6 upon glycogen depletion in muscle.
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Research Article|
March 29 2016
The interaction between AMPKβ2 and the PP1-targeting subunit R6 is dynamically regulated by intracellular glycogen content
Yvonne Oligschlaeger;
Yvonne Oligschlaeger
*Department of Molecular Genetics, CARIM School for Cardiovascular Diseases, Maastricht University, 6200 MD Maastricht, The Netherlands
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Marie Miglianico;
Marie Miglianico
*Department of Molecular Genetics, CARIM School for Cardiovascular Diseases, Maastricht University, 6200 MD Maastricht, The Netherlands
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Vivian Dahlmans;
Vivian Dahlmans
†Department of Molecular Genetics, GROW School for Oncology & Developmental Biology, Maastricht University, 6200 MD Maastricht, The Netherlands
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Carla Rubio-Villena;
Carla Rubio-Villena
‡Instituto de Biomedicina de Valencia, Consejo Superior de Investigaciones Cientificas (CSIC) and Centro de Investigación Biomédica en Red de Enfermedades Raras (CIBERER), 46010 Valencia, Spain
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Dipanjan Chanda;
Dipanjan Chanda
*Department of Molecular Genetics, CARIM School for Cardiovascular Diseases, Maastricht University, 6200 MD Maastricht, The Netherlands
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Maria Adelaida Garcia-Gimeno;
Maria Adelaida Garcia-Gimeno
‡Instituto de Biomedicina de Valencia, Consejo Superior de Investigaciones Cientificas (CSIC) and Centro de Investigación Biomédica en Red de Enfermedades Raras (CIBERER), 46010 Valencia, Spain
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Will A. Coumans;
Will A. Coumans
*Department of Molecular Genetics, CARIM School for Cardiovascular Diseases, Maastricht University, 6200 MD Maastricht, The Netherlands
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Yilin Liu;
Yilin Liu
*Department of Molecular Genetics, CARIM School for Cardiovascular Diseases, Maastricht University, 6200 MD Maastricht, The Netherlands
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J. Willem Voncken;
J. Willem Voncken
†Department of Molecular Genetics, GROW School for Oncology & Developmental Biology, Maastricht University, 6200 MD Maastricht, The Netherlands
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Joost J.F.P. Luiken;
Joost J.F.P. Luiken
*Department of Molecular Genetics, CARIM School for Cardiovascular Diseases, Maastricht University, 6200 MD Maastricht, The Netherlands
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Jan F.C. Glatz;
Jan F.C. Glatz
*Department of Molecular Genetics, CARIM School for Cardiovascular Diseases, Maastricht University, 6200 MD Maastricht, The Netherlands
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Pascual Sanz;
Pascual Sanz
‡Instituto de Biomedicina de Valencia, Consejo Superior de Investigaciones Cientificas (CSIC) and Centro de Investigación Biomédica en Red de Enfermedades Raras (CIBERER), 46010 Valencia, Spain
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Dietbert Neumann
Dietbert Neumann
1
*Department of Molecular Genetics, CARIM School for Cardiovascular Diseases, Maastricht University, 6200 MD Maastricht, The Netherlands
1To whom correspondence should be addressed (email d.neumann@maastrichtuniversity.nl).
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Publisher: Portland Press Ltd
Received:
September 30 2015
Revision Received:
January 22 2016
Accepted:
February 01 2016
Accepted Manuscript online:
February 01 2016
Online ISSN: 1470-8728
Print ISSN: 0264-6021
© 2016 Authors; published by Portland Press Limited
2016
Biochem J (2016) 473 (7): 937–947.
Article history
Received:
September 30 2015
Revision Received:
January 22 2016
Accepted:
February 01 2016
Accepted Manuscript online:
February 01 2016
Citation
Yvonne Oligschlaeger, Marie Miglianico, Vivian Dahlmans, Carla Rubio-Villena, Dipanjan Chanda, Maria Adelaida Garcia-Gimeno, Will A. Coumans, Yilin Liu, J. Willem Voncken, Joost J.F.P. Luiken, Jan F.C. Glatz, Pascual Sanz, Dietbert Neumann; The interaction between AMPKβ2 and the PP1-targeting subunit R6 is dynamically regulated by intracellular glycogen content. Biochem J 1 April 2016; 473 (7): 937–947. doi: https://doi.org/10.1042/BJ20151035
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