title: Disruption of AMPKalpha1 signaling prevents AICAR-induced inhibition of AS160/TBC1D4 phosphorylation and glucose uptake in primary rat adipocytes.
abstract: The aim of this study was to investigate the molecular mechanisms by which AMP-kinase (AMPK) activation inhibits basal and insulin-stimulated glucose uptake in primary adipocytes. Rat epididymal adipocytes were exposed to 5-aminoimidazole-4-carboxamide-1-beta-d-ribofuranoside (AICAR) for 1 h. Subsequently, basal and insulin-stimulated glucose uptake and the phosphorylation of AMPK, acetyl-CoA carboxylase, Akt, and the Akt substrate of 160 kDa (AS160/TBC1D4) were determined. In order to investigate whether these effects of AICAR were mediated by AMPK activation, these parameters were also assessed in adipocytes either expressing LacZ (control) or a kinase-dead AMPKalpha1 mutant. AICAR increased AMPK activation without affecting basal and insulin-stimulated Akt1/2 phosphorylation on Thr(308) and Ser(473) residues. However, AMPK activation suppressed the phosphorylation of AS160/TBC1D4 and its interaction with the 14-3-3 signal transduction-regulatory protein, which was accompanied by significant reductions in plasma membrane glucose transporter 4 content and glucose uptake under basal and insulin-stimulated conditions. Phosphorylation of Akt substrates glycogen synthase kinase 3alpha and -beta were unaltered by AICAR, indicating that the AMPK-regulatory effects were specific to the AS160/TBC1D4 signaling pathway. Expression of the kinase-dead AMPKalpha1 mutant fully prevented the suppression of AS160/TBC1D4 phosphorylation, plasma membrane glucose transporter 4 content, and the inhibitory effect of AICAR-induced AMPK activation on basal and insulin-stimulated glucose uptake. This study is the first to provide evidence that disruption of AMPKalpha1 signaling prevents the suppressive effects of AMPK activation on AS160/TBC1D4 phosphorylation and glucose uptake, indicating that insulin-signaling steps that are common to white adipose tissue and skeletal muscle regulation of glucose uptake are distinctly affected by AMPK activation.
authors.affiliation: School of Kinesiology and Health Science, York University, Toronto, Ontario, Canada M3J1P3.
authors.forename: Mandeep P
authors.initials: MP
authors.lastname: Gaidhu
authors.forename: Robert L S
authors.initials: RL
authors.lastname: Perry
authors.forename: Fawad
authors.initials: F
authors.lastname: Noor
authors.forename: Rolando B
authors.initials: RB
authors.lastname: Ceddia
mesh_terms: D055372:AMP-Activated Protein Kinases / Q000235:genetics / Q000378:metabolism*; D017667:Adipocytes / Q000187:drug effects* / Q000378:metabolism*; D000620:Aminoimidazole Carboxamide / Q000031:analogs & derivatives* / Q000494:pharmacology; D000818:Animals; D001692:Biological Transport / Q000187:drug effects / Q000235:genetics; D020690:GTPase-Activating Proteins / Q000235:genetics / Q000378:metabolism*; D005947:Glucose / Q000378:metabolism*; D051275:Glucose Transporter Type 4 / Q000235:genetics / Q000378:metabolism; D015151:Immunoblotting; D047468:Immunoprecipitation; D008297:Male; D010766:Phosphorylation / Q000187:drug effects; D021381:Protein Transport / Q000187:drug effects / Q000235:genetics; D051381:Rats; D017208:Rats, Wistar; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012265:Ribonucleotides / Q000494:pharmacology*; D015398:Signal Transduction / Q000187:drug effects
chemical_list: D020690:GTPase-Activating Proteins; D051275:Glucose Transporter Type 4; D012265:Ribonucleotides; C495694:Slc2a4 protein, rat; C533101:TBC1D4 protein, rat; D000620:Aminoimidazole Carboxamide; D055372:AMP-Activated Protein Kinases; C031143:AICA ribonucleotide; D005947:Glucose