title: Activation of protein phosphatase 2A by palmitate inhibits AMP-activated protein kinase.
abstract: Elevated levels of free fatty acids contribute to cardiovascular diseases, but the mechanisms remain poorly understood. The present study was aimed to determine if free fatty acid inhibits the AMP-activated kinase (AMPK). Exposure of cultured bovine aortic endothelial cells (BAECs) to palmitate (0.4 mM) but not to palmitoleic or oleic acid (0.4 mM) for 40 h significantly reduced the Thr(172) phosphorylation of AMPK-alpha without altering its protein expression or the phosphorylation of LKB1-Ser(428), a major AMPK kinase in BAECs. Further, in LKB1-deficient cells, palmitate suppressed AMPK-Thr(172) implying that the inhibitory effects of palmitate on AMPK might be independent of LKB1. In contrast, 2-bromopalmitate, a non-metabolizable analog of palmitate, did not alter the phosphorylation of AMPK and acetyl-CoA carboxylase. Further, palmitate significantly increased the activity of protein phosphatase (PP)2A. Inhibition of PP2A with either okadaic acid, a selective PP2A inhibitor, or PP2A small interference RNA abolished palmitate-induced inhibition on AMPK-Thr(172) phosphorylation. Exposure of BAECs to C(2)-ceramide, a cell-permeable analog of ceramide, mimicked the effects of palmitate. Conversely, fumonisin B1, which selectively inhibits ceramide synthase and decreases de novo formation of ceramide, abolished the effects of palmitate on both PP2A and AMPK. Inhibition of AMPK in parallel with increased PP2A activity was founded in C57BL/6J mice fed with high fat diet (HFD) rich in palmitate but not in mice fed with HFD rich in oleate. Moreover, inhibition of PP2A with PP2A-specific siRNA but not scrambled siRNA reversed HFD-induced inhibition on the phosphorylation of AMPK-Thr(172) and endothelial nitric-oxide synthase (eNOS)-Ser(1177) in mice fed with high fat diets. Taken together, we conclude that palmitate inhibits the phosphorylation of both AMPK and endothelial nitric-oxide synthase in endothelial cells via ceramide-dependent PP2A activation.
authors.affiliation: Division of Endocrinology and Diabetes, Department of Medicine, University of Oklahoma Health Science Center, Oklahoma City, Oklahoma 73104.
authors.forename: Yong
authors.initials: Y
authors.lastname: Wu
authors.affiliation: Division of Endocrinology and Diabetes, Department of Medicine, University of Oklahoma Health Science Center, Oklahoma City, Oklahoma 73104.
authors.forename: Ping
authors.initials: P
authors.lastname: Song
authors.affiliation: Division of Endocrinology and Diabetes, Department of Medicine, University of Oklahoma Health Science Center, Oklahoma City, Oklahoma 73104.
authors.forename: Jian
authors.initials: J
authors.lastname: Xu
authors.affiliation: Division of Endocrinology and Diabetes, Department of Medicine, University of Oklahoma Health Science Center, Oklahoma City, Oklahoma 73104.
authors.forename: Miao
authors.initials: M
authors.lastname: Zhang
authors.affiliation: Division of Endocrinology and Diabetes, Department of Medicine, University of Oklahoma Health Science Center, Oklahoma City, Oklahoma 73104. Electronic address: ming-hui-zou@ouhsc.edu.
authors.forename: Ming-Hui
authors.initials: MH
authors.lastname: Zou
mesh_terms: D055372:AMP-Activated Protein Kinases; D000818:Animals; D002417:Cattle; D002460:Cell Line; D004789:Enzyme Activation / Q000502:physiology; D020536:Enzyme Activators / Q000494:pharmacology*; D006367:HeLa Cells; D006801:Humans; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D009097:Multienzyme Complexes / Q000037:antagonists & inhibitors*; D052250:Nitric Oxide Synthase Type III / Q000037:antagonists & inhibitors; D010168:Palmitates / Q000494:pharmacology*; D010749:Phosphoprotein Phosphatases / Q000378:metabolism* / Q000502:physiology; D010766:Phosphorylation; D054648:Protein Phosphatase 2; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors*
chemical_list: D020536:Enzyme Activators; D009097:Multienzyme Complexes; D010168:Palmitates; D052250:Nitric Oxide Synthase Type III; D017346:Protein Serine-Threonine Kinases; D055372:AMP-Activated Protein Kinases; D010749:Phosphoprotein Phosphatases; D054648:Protein Phosphatase 2