title: Pre-B cell colony enhancing factor induces Nampt-dependent translocation of the insulin receptor out of lipid microdomains in A549 lung epithelial cells.
abstract: Pre-B cell colony-enhancing factor (PBEF) is a highly conserved pleiotropic protein reported to be an alternate ligand for the insulin receptor (IR). We sought to clarify the relationship between PBEF and insulin signaling by evaluating the effects of PBEF on the localization of the IRβ chain to lipid rafts in A549 epithelial cells. We isolated lipid rafts from A549 cells and detected the IR by immunoprecipitation from raft fractions or whole cell lysates. Cells were treated with rPBEF, its enzymatic product nicotinamide adenine dinucleotide (NAD), or the Nampt inhibitor daporinad to study the effect of PBEF on IRβ movement. We used coimmunoprecipitation studies in cells transfected with PBEF and IRβ constructs to detect interactions between PBEF, the IRβ, and caveolin-1 (Cav-1). PBEF was present in both lipid raft and nonraft fractions, whereas the IR was found only in lipid raft fractions of resting A549 cells. The IR-, PBEF-, and Cav-1-coimmunoprecipitated rPBEF treatment resulted in the movement of IRβ- and tyrosine-phosphorylated Cav-1 from lipid rafts to nonrafts, an effect that could be blocked by daporinad, suggesting that this effect was facilitated by the Nampt activity of PBEF. The addition of PBEF to insulin-treated cells resulted in reduced Akt phosphorylation of both Ser⁴⁷³ and Thr³⁰⁸. We conclude that PBEF can inhibit insulin signaling through the IR by Nampt-dependent promotion of IR translocation into the nonraft domains of A549 epithelial cells. PBEF-induced alterations in the spatial geometry of the IR provide a mechanistic explanation for insulin resistance in inflammatory states associated with upregulation of PBEF.
authors.affiliation: Keenan Research Centre for Biomedical Science, St. Michael's Hospital, and the Interdepartmental Division of Critical Care Medicine, University of Toronto, Toronto, Ontario, Canada; and Department of Critical Care Medicine, Xiangya Hospital, Central South University, Changsha, China.
authors.forename: Qianyi
authors.initials: Q
authors.lastname: Peng
authors.affiliation: Department of Surgery, Keenan Research Centre for Biomedical Science, St. Michael's Hospital, and the Interdepartmental Division of Critical Care Medicine, University of Toronto, Toronto, Ontario, Canada; and.
authors.forename: Song Hui
authors.initials: SH
authors.lastname: Jia
authors.affiliation: Department of Surgery, Keenan Research Centre for Biomedical Science, St. Michael's Hospital, and the Interdepartmental Division of Critical Care Medicine, University of Toronto, Toronto, Ontario, Canada; and.
authors.forename: Jean
authors.initials: J
authors.lastname: Parodo
authors.affiliation: Department of Critical Care Medicine, Xiangya Hospital, Central South University, Changsha, China.
authors.forename: Yuhang
authors.initials: Y
authors.lastname: Ai
authors.affiliation: Department of Surgery, Department of Critical Care Medicine, and Keenan Research Centre for Biomedical Science, St. Michael's Hospital, and the Interdepartmental Division of Critical Care Medicine, University of Toronto, Toronto, Ontario, Canada; and marshallj@smh.ca.
authors.forename: John C
authors.initials: JC
authors.lastname: Marshall
mesh_terms: D000178:Acrylamides / Q000494:pharmacology; D015703:Antigens, CD / Q000235:genetics / Q000378:metabolism*; D051242:Caveolin 1 / Q000235:genetics / Q000378:metabolism; D002460:Cell Line; D016207:Cytokines / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D007328:Insulin / Q000378:metabolism*; D007333:Insulin Resistance; D008168:Lung / Q000187:drug effects / Q000378:metabolism*; D021962:Membrane Microdomains / Q000187:drug effects / Q000378:metabolism*; D009243:NAD / Q000378:metabolism; D054409:Nicotinamide Phosphoribosyltransferase / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation / Q000187:drug effects; D010880:Piperidines / Q000494:pharmacology; D011499:Protein Processing, Post-Translational / Q000187:drug effects; D021381:Protein Transport / Q000187:drug effects; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D011972:Receptor, Insulin / Q000819:agonists / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D020545:Respiratory Mucosa / Q000187:drug effects / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects
chemical_list: D000178:Acrylamides; D015703:Antigens, CD; C495540:CAV1 protein, human; D051242:Caveolin 1; D016207:Cytokines; D004791:Enzyme Inhibitors; D007328:Insulin; C480543:N-(4-(1-benzoylpiperidin-4-yl)butyl)-3-(pyridin-3-yl)acrylamide; D010880:Piperidines; D011994:Recombinant Proteins; D009243:NAD; D054409:Nicotinamide Phosphoribosyltransferase; C085455:nicotinamide phosphoribosyltransferase, human; C435941:INSR protein, human; D011972:Receptor, Insulin; C494918:AKT1 protein, human; D051057:Proto-Oncogene Proteins c-akt
keywords: caveolin-1; insulin; insulin receptor; lipid rafts; nicotinamide phosphoribosyl transferase; phosphorylation; pre-B cell colony-enhancing factor; signaling