Genetic disruption of AMPK signaling abolishes both contraction- and insulin-stimulated TBC1D1 phosphorylation and 14-3-3 binding in mouse skeletal muscle

Christian Pehmoller, Jonas T. Treebak, Jesper B. Birk, Shuai Chen, Carol MacKintosh, D. Grahame Hardie, Erik A. Richter, Jorgen F. P. Wojtaszewski

    Research output: Contribution to journalArticle

    99 Citations (Scopus)

    Abstract

    Pehmoller C, Treebak JT, Birk JB, Chen S, MacKintosh C, Hardie DG, Richter EA, Wojtaszewski JF. Genetic disruption of AMPK signaling abolishes both contraction- and insulin-stimulated TBC1D1 phosphorylation and 14-3-3 binding in mouse skeletal muscle. Am J Physiol Endocrinol Metab 297: E665-E675, 2009. First published June 16, 2009; doi: 10.1152/ajpendo.00115.2009.-TBC1D1 is a Rab-GTPase-activating protein (GAP) known to be phosphorylated in response to insulin, growth factors, pharmacological agonists that activate 5'-AMP-activated protein kinase (AMPK), and muscle contraction. Silencing TBC1D1 in L6 muscle cells by siRNA increases insulin-stimulated GLUT4 translocation, and overexpression of TBC1D1 in 3T3-L1 adipocytes with low endogenous TBC1D1 expression inhibits insulin-stimulated GLUT4 translocation, suggesting a role of TBC1D1 in regulating GLUT4 translocation. Aiming to unravel the regulation of TBC1D1 during contraction and the potential role of AMPK in intact skeletal muscle, we used EDL muscles from wild-type (WT) and AMPK kinase dead (KD) mice. We explored the site-specific phosphorylation of TBC1D1 Ser(237) and Thr(596) and their relation to 14-3-3 binding, a proposed mechanism for regulation of GAP function of TBC1D1. We show that muscle contraction increases 14-3-3 binding to TBC1D1 as well as phosphorylation of Ser(237) and Thr(596) in an AMPK-dependent manner. AMPK activation by AICAR induced similar Ser(237) and Thr(596) phosphorylation of, and 14-3-3 binding to, TBC1D1 as muscle contraction. Insulin did not increase Ser(237) phosphorylation or 14-3-3 binding to TBC1D1. However, insulin increased Thr(596) phosphorylation, and intriguingly this response was fully abolished in the AMPK KD mice. Thus, TBC1D1 is differentially regulated in response to insulin and contraction. This study provides genetic evidence to support an important role for AMPK in regulating TBC1D1 in response to both of these physiological stimuli.

    Original languageEnglish
    Pages (from-to)E665-E675
    Number of pages11
    JournalAmerican Journal of Physiology, Endocrinology and Metabolism
    Volume297
    Issue number3
    DOIs
    Publication statusPublished - Sep 2009

    Keywords

    • 5 '-AMP-activated protein kinase
    • wortmannin
    • 5-aminoimidazole-4-carboxamide-1-beta-D-ribofuranoside
    • AMPK kinase dead
    • extensor digitorum longus muscle
    • ACTIVATED PROTEIN-KINASE
    • ACETYL-COA CARBOXYLASE
    • GLUT4 TRANSLOCATION
    • GLUCOSE-TRANSPORT
    • SUBSTRATE-SPECIFICITY
    • IN-VITRO
    • AS160
    • INHIBITION
    • INCREASES
    • EXERCISE

    Cite this

    @article{b1194632a94241f4b1fe74ba4cb28c2e,
    title = "Genetic disruption of AMPK signaling abolishes both contraction- and insulin-stimulated TBC1D1 phosphorylation and 14-3-3 binding in mouse skeletal muscle",
    abstract = "Pehmoller C, Treebak JT, Birk JB, Chen S, MacKintosh C, Hardie DG, Richter EA, Wojtaszewski JF. Genetic disruption of AMPK signaling abolishes both contraction- and insulin-stimulated TBC1D1 phosphorylation and 14-3-3 binding in mouse skeletal muscle. Am J Physiol Endocrinol Metab 297: E665-E675, 2009. First published June 16, 2009; doi: 10.1152/ajpendo.00115.2009.-TBC1D1 is a Rab-GTPase-activating protein (GAP) known to be phosphorylated in response to insulin, growth factors, pharmacological agonists that activate 5'-AMP-activated protein kinase (AMPK), and muscle contraction. Silencing TBC1D1 in L6 muscle cells by siRNA increases insulin-stimulated GLUT4 translocation, and overexpression of TBC1D1 in 3T3-L1 adipocytes with low endogenous TBC1D1 expression inhibits insulin-stimulated GLUT4 translocation, suggesting a role of TBC1D1 in regulating GLUT4 translocation. Aiming to unravel the regulation of TBC1D1 during contraction and the potential role of AMPK in intact skeletal muscle, we used EDL muscles from wild-type (WT) and AMPK kinase dead (KD) mice. We explored the site-specific phosphorylation of TBC1D1 Ser(237) and Thr(596) and their relation to 14-3-3 binding, a proposed mechanism for regulation of GAP function of TBC1D1. We show that muscle contraction increases 14-3-3 binding to TBC1D1 as well as phosphorylation of Ser(237) and Thr(596) in an AMPK-dependent manner. AMPK activation by AICAR induced similar Ser(237) and Thr(596) phosphorylation of, and 14-3-3 binding to, TBC1D1 as muscle contraction. Insulin did not increase Ser(237) phosphorylation or 14-3-3 binding to TBC1D1. However, insulin increased Thr(596) phosphorylation, and intriguingly this response was fully abolished in the AMPK KD mice. Thus, TBC1D1 is differentially regulated in response to insulin and contraction. This study provides genetic evidence to support an important role for AMPK in regulating TBC1D1 in response to both of these physiological stimuli.",
    keywords = "5 '-AMP-activated protein kinase, wortmannin, 5-aminoimidazole-4-carboxamide-1-beta-D-ribofuranoside, AMPK kinase dead, extensor digitorum longus muscle, ACTIVATED PROTEIN-KINASE, ACETYL-COA CARBOXYLASE, GLUT4 TRANSLOCATION, GLUCOSE-TRANSPORT, SUBSTRATE-SPECIFICITY, IN-VITRO, AS160, INHIBITION, INCREASES, EXERCISE",
    author = "Christian Pehmoller and Treebak, {Jonas T.} and Birk, {Jesper B.} and Shuai Chen and Carol MacKintosh and Hardie, {D. Grahame} and Richter, {Erik A.} and Wojtaszewski, {Jorgen F. P.}",
    year = "2009",
    month = "9",
    doi = "10.1152/ajpendo.00115.2009",
    language = "English",
    volume = "297",
    pages = "E665--E675",
    journal = "American Journal of Physiology, Endocrinology and Metabolism",
    issn = "0193-1849",
    publisher = "American Physiological Society",
    number = "3",

    }

    Genetic disruption of AMPK signaling abolishes both contraction- and insulin-stimulated TBC1D1 phosphorylation and 14-3-3 binding in mouse skeletal muscle. / Pehmoller, Christian; Treebak, Jonas T.; Birk, Jesper B.; Chen, Shuai; MacKintosh, Carol; Hardie, D. Grahame; Richter, Erik A.; Wojtaszewski, Jorgen F. P.

    In: American Journal of Physiology, Endocrinology and Metabolism, Vol. 297, No. 3, 09.2009, p. E665-E675.

    Research output: Contribution to journalArticle

    TY - JOUR

    T1 - Genetic disruption of AMPK signaling abolishes both contraction- and insulin-stimulated TBC1D1 phosphorylation and 14-3-3 binding in mouse skeletal muscle

    AU - Pehmoller, Christian

    AU - Treebak, Jonas T.

    AU - Birk, Jesper B.

    AU - Chen, Shuai

    AU - MacKintosh, Carol

    AU - Hardie, D. Grahame

    AU - Richter, Erik A.

    AU - Wojtaszewski, Jorgen F. P.

    PY - 2009/9

    Y1 - 2009/9

    N2 - Pehmoller C, Treebak JT, Birk JB, Chen S, MacKintosh C, Hardie DG, Richter EA, Wojtaszewski JF. Genetic disruption of AMPK signaling abolishes both contraction- and insulin-stimulated TBC1D1 phosphorylation and 14-3-3 binding in mouse skeletal muscle. Am J Physiol Endocrinol Metab 297: E665-E675, 2009. First published June 16, 2009; doi: 10.1152/ajpendo.00115.2009.-TBC1D1 is a Rab-GTPase-activating protein (GAP) known to be phosphorylated in response to insulin, growth factors, pharmacological agonists that activate 5'-AMP-activated protein kinase (AMPK), and muscle contraction. Silencing TBC1D1 in L6 muscle cells by siRNA increases insulin-stimulated GLUT4 translocation, and overexpression of TBC1D1 in 3T3-L1 adipocytes with low endogenous TBC1D1 expression inhibits insulin-stimulated GLUT4 translocation, suggesting a role of TBC1D1 in regulating GLUT4 translocation. Aiming to unravel the regulation of TBC1D1 during contraction and the potential role of AMPK in intact skeletal muscle, we used EDL muscles from wild-type (WT) and AMPK kinase dead (KD) mice. We explored the site-specific phosphorylation of TBC1D1 Ser(237) and Thr(596) and their relation to 14-3-3 binding, a proposed mechanism for regulation of GAP function of TBC1D1. We show that muscle contraction increases 14-3-3 binding to TBC1D1 as well as phosphorylation of Ser(237) and Thr(596) in an AMPK-dependent manner. AMPK activation by AICAR induced similar Ser(237) and Thr(596) phosphorylation of, and 14-3-3 binding to, TBC1D1 as muscle contraction. Insulin did not increase Ser(237) phosphorylation or 14-3-3 binding to TBC1D1. However, insulin increased Thr(596) phosphorylation, and intriguingly this response was fully abolished in the AMPK KD mice. Thus, TBC1D1 is differentially regulated in response to insulin and contraction. This study provides genetic evidence to support an important role for AMPK in regulating TBC1D1 in response to both of these physiological stimuli.

    AB - Pehmoller C, Treebak JT, Birk JB, Chen S, MacKintosh C, Hardie DG, Richter EA, Wojtaszewski JF. Genetic disruption of AMPK signaling abolishes both contraction- and insulin-stimulated TBC1D1 phosphorylation and 14-3-3 binding in mouse skeletal muscle. Am J Physiol Endocrinol Metab 297: E665-E675, 2009. First published June 16, 2009; doi: 10.1152/ajpendo.00115.2009.-TBC1D1 is a Rab-GTPase-activating protein (GAP) known to be phosphorylated in response to insulin, growth factors, pharmacological agonists that activate 5'-AMP-activated protein kinase (AMPK), and muscle contraction. Silencing TBC1D1 in L6 muscle cells by siRNA increases insulin-stimulated GLUT4 translocation, and overexpression of TBC1D1 in 3T3-L1 adipocytes with low endogenous TBC1D1 expression inhibits insulin-stimulated GLUT4 translocation, suggesting a role of TBC1D1 in regulating GLUT4 translocation. Aiming to unravel the regulation of TBC1D1 during contraction and the potential role of AMPK in intact skeletal muscle, we used EDL muscles from wild-type (WT) and AMPK kinase dead (KD) mice. We explored the site-specific phosphorylation of TBC1D1 Ser(237) and Thr(596) and their relation to 14-3-3 binding, a proposed mechanism for regulation of GAP function of TBC1D1. We show that muscle contraction increases 14-3-3 binding to TBC1D1 as well as phosphorylation of Ser(237) and Thr(596) in an AMPK-dependent manner. AMPK activation by AICAR induced similar Ser(237) and Thr(596) phosphorylation of, and 14-3-3 binding to, TBC1D1 as muscle contraction. Insulin did not increase Ser(237) phosphorylation or 14-3-3 binding to TBC1D1. However, insulin increased Thr(596) phosphorylation, and intriguingly this response was fully abolished in the AMPK KD mice. Thus, TBC1D1 is differentially regulated in response to insulin and contraction. This study provides genetic evidence to support an important role for AMPK in regulating TBC1D1 in response to both of these physiological stimuli.

    KW - 5 '-AMP-activated protein kinase

    KW - wortmannin

    KW - 5-aminoimidazole-4-carboxamide-1-beta-D-ribofuranoside

    KW - AMPK kinase dead

    KW - extensor digitorum longus muscle

    KW - ACTIVATED PROTEIN-KINASE

    KW - ACETYL-COA CARBOXYLASE

    KW - GLUT4 TRANSLOCATION

    KW - GLUCOSE-TRANSPORT

    KW - SUBSTRATE-SPECIFICITY

    KW - IN-VITRO

    KW - AS160

    KW - INHIBITION

    KW - INCREASES

    KW - EXERCISE

    U2 - 10.1152/ajpendo.00115.2009

    DO - 10.1152/ajpendo.00115.2009

    M3 - Article

    C2 - 19531644

    VL - 297

    SP - E665-E675

    JO - American Journal of Physiology, Endocrinology and Metabolism

    JF - American Journal of Physiology, Endocrinology and Metabolism

    SN - 0193-1849

    IS - 3

    ER -