LKB1, an upstream AMPK kinase, regulates glucose and lipid metabolism in cultured liver and muscle cells.
LKB1 is a 50 kDa serine/threonine kinase that phosphorylates and activates the catalytic subunit of AMPK at its T-loop residue Thr 172. We prepared adenoviruses expressing the constitutive active (wild-type) form (CA) or dominant negative (kinase inactive, D194A mutant) form (DN) of LKB1 and overexpressed these proteins in cultured myotubes (C2C12 cells) and rat hepatoma cells (FAO cells). When analyzed by immunoblotting with the antibody against Thr172-phosphorylated AMPK, the phosphorylation of AMPK was increased (2.5-fold) and decreased (0.4-fold) in cells expressing CA and DN LKB1, respectively, as compared with Lac-Z expressing control cells. Immunoprecipitation experiments, using isoform-specific antibody, revealed these alterations of AMPK phosphorylation to be attributable to altered phosphorylation of AMPK alpha2, but not alpha1 catalytic subunits, strongly suggesting the alpha2 catalytic subunit to be the major substrate for LKB1 in mammalian cells. In addition, adiponectin or AICAR-stimulated AMPK phosphorylation was inhibited by overexpression of DN LKB1, while phenformin-stimulated phosphorylation was unaffected. These results may explain the difference in AMPK activation mechanisms between AMP and phenformin, and also indicate that AMPK phosphorylation by LKB1 is involved in AMP-stimulated AMPK activation. As a downstream target for AMPK, AICAR-induced glucose uptake and ACCbeta phosphorylation were found to be significantly reduced in DN LKB1 expressing C2C12 cells. The expression of key enzymes for gluconeogenesis, glucose-6-phosphatase and phosphoenolpyruvate carboxykinase, was also dependent on LKB1 activities in FAO cells. These results demonstrate that LKB1 is a crucial regulator of AMPK activation in muscle and liver cells and, therefore, that LKB1 activity is potentially of importance to our understanding of glucose and lipid metabolism.
Imai
Kenta
K
Division of Endocrinology and Diabetes, Department of Medicine, Saitama Medical School, Iruma-gun, Saitama 350-0495, Japan.
Inukai
Kouichi
K
Ikegami
Yuichi
Y
Awata
Takuya
T
Katayama
Shigehiro
S
D000091162:AMP-Activated Protein Kinase Kinases; D055372:AMP-Activated Protein Kinases; D000818:Animals; D002460:Cell Line; D005246:Feedback / Q000502:physiology; D005947:Glucose / Q000378:metabolism*; D022781:Hepatocytes / Q000378:metabolism*; D050356:Lipid Metabolism / Q000502:physiology*; D051379:Mice; D018485:Muscle Fibers, Skeletal / Q000378:metabolism*; D011494:Protein Kinases / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*
D011494:Protein Kinases; D017346:Protein Serine-Threonine Kinases; C491654:Stk11 protein, mouse; D000091162:AMP-Activated Protein Kinase Kinases; D055372:AMP-Activated Protein Kinases; D005947:Glucose