title: Protein kinase Cδ regulates nuclear export of FOXO1 through phosphorylation of the chaperone 14-3-3ζ.
abstract: OBJECTIVE
  Forkhead box protein O1 (FOXO1) is a transcription factor essential for beta cell fate. Protein kinase B-dependent phosphorylation of FOXO1 at S256 (P-FOXO1) enables its binding to 14-3-3 dimers and nuclear export. Dephosphorylated FOXO1 enters nuclei and activates pro-apoptotic genes. Since our previous observations suggest that protein kinase C delta (PKCδ) induces nuclear accumulation of FOXO1, the underlying mechanism was examined.
  
  
  METHODS
  In human islets, genetically modified mice and INS-1E cells apoptosis was assessed by TUNEL staining. Subcellular translocation of proteins was examined by confocal microscopy and signalling pathways were analysed by western blotting and overlay assay.
  
  
  RESULTS
  In PKCδ-overexpressing (PKCδ-tg) mouse islet cells and INS-1E cells FOXO1 accumulated in nuclei, surprisingly, as P-FOXO1. PKCδ-tg decelerated IGF-1-dependent stimulation of nuclear export, indicating that changes in export caused nuclear retention of P-FOXO1. Nuclear accumulation of P-FOXO1 was accompanied by increased phosphorylation of 14-3-3ζ at S58 and reduced dimerisation of 14-3-3ζ. Palmitic acid further augmented phosphorylation of 14-3-3ζ and triggered nuclear accumulation of FOXO1 in both INS-1E and human islet cells. Furthermore, the overexpression of a phosphomimicking mutant of 14-3-3ζ (S58D) enhanced nuclear FOXO1. In accordance with the nuclear accumulation of P-FOXO1, PKCδ overexpression alone did not increase apoptotic cell death. Additionally, insulin secretion and glucose homeostasis in PKCδ-overexpressing mice remained unaffected.
  
  
  CONCLUSIONS
  These results suggest that PKCδ-mediated phosphorylation of 14-3-3ζ contributes to the nuclear retention of FOXO1, even when FOXO1 is phosphorylated as under non-stress conditions. P-FOXO1 does not induce pro-apoptotic genes, but may rather exert beneficial effects on beta cells.
authors.affiliation: Division of Endocrinology, Diabetology, Angiology, Nephrology and Clinical Chemistry, Department of Internal Medicine IV, University Hospital Tübingen, D-72076, Tübingen, Germany.|Institute for Diabetes Research and Metabolic Diseases of the Helmholtz Center Munich at the University of Tübingen (IDM), Partner in the German Center for Diabetes Research (DZD), Tübingen, Germany.
authors.forename: Felicia
authors.initials: F
authors.lastname: Gerst
authors.affiliation: Division of Endocrinology, Diabetology, Angiology, Nephrology and Clinical Chemistry, Department of Internal Medicine IV, University Hospital Tübingen, D-72076, Tübingen, Germany.|Institute for Diabetes Research and Metabolic Diseases of the Helmholtz Center Munich at the University of Tübingen (IDM), Partner in the German Center for Diabetes Research (DZD), Tübingen, Germany.
authors.forename: Gabriele
authors.initials: G
authors.lastname: Kaiser
authors.affiliation: Division of Endocrinology, Diabetology, Angiology, Nephrology and Clinical Chemistry, Department of Internal Medicine IV, University Hospital Tübingen, D-72076, Tübingen, Germany.
authors.forename: Madhura
authors.initials: M
authors.lastname: Panse
authors.affiliation: Division of Endocrinology, Diabetology, Angiology, Nephrology and Clinical Chemistry, Department of Internal Medicine IV, University Hospital Tübingen, D-72076, Tübingen, Germany.|Institute for Diabetes Research and Metabolic Diseases of the Helmholtz Center Munich at the University of Tübingen (IDM), Partner in the German Center for Diabetes Research (DZD), Tübingen, Germany.
authors.forename: Tina
authors.initials: T
authors.lastname: Sartorius
authors.affiliation: Center of Animal Biotechnology and Gene Therapy, Universitat Autònoma de Barcelona, Bellaterra, Barcelona, Spain.|CIBER de Diabetes y Enfermedades Metabólicas Asociadas (CIBERDEM), Spain.
authors.forename: Anna
authors.initials: A
authors.lastname: Pujol
authors.affiliation: Division of Endocrinology, Diabetology, Angiology, Nephrology and Clinical Chemistry, Department of Internal Medicine IV, University Hospital Tübingen, D-72076, Tübingen, Germany.
authors.forename: Anita M
authors.initials: AM
authors.lastname: Hennige
authors.affiliation: Division of Endocrinology, Diabetology, Angiology, Nephrology and Clinical Chemistry, Department of Internal Medicine IV, University Hospital Tübingen, D-72076, Tübingen, Germany.|Institute for Diabetes Research and Metabolic Diseases of the Helmholtz Center Munich at the University of Tübingen (IDM), Partner in the German Center for Diabetes Research (DZD), Tübingen, Germany.
authors.forename: Fausto
authors.initials: F
authors.lastname: Machicao
authors.affiliation: Division of Endocrinology, Diabetology, Angiology, Nephrology and Clinical Chemistry, Department of Internal Medicine IV, University Hospital Tübingen, D-72076, Tübingen, Germany.
authors.forename: Reiner
authors.initials: R
authors.lastname: Lammers
authors.affiliation: Center of Animal Biotechnology and Gene Therapy, Universitat Autònoma de Barcelona, Bellaterra, Barcelona, Spain.|CIBER de Diabetes y Enfermedades Metabólicas Asociadas (CIBERDEM), Spain.
authors.forename: Fatima
authors.initials: F
authors.lastname: Bosch
authors.affiliation: Division of Endocrinology, Diabetology, Angiology, Nephrology and Clinical Chemistry, Department of Internal Medicine IV, University Hospital Tübingen, D-72076, Tübingen, Germany.|Institute for Diabetes Research and Metabolic Diseases of the Helmholtz Center Munich at the University of Tübingen (IDM), Partner in the German Center for Diabetes Research (DZD), Tübingen, Germany.
authors.forename: Hans-Ulrich
authors.initials: HU
authors.lastname: Häring
authors.affiliation: Division of Endocrinology, Diabetology, Angiology, Nephrology and Clinical Chemistry, Department of Internal Medicine IV, University Hospital Tübingen, D-72076, Tübingen, Germany. susanne.ullrich@med.uni-tuebingen.de.|Institute for Diabetes Research and Metabolic Diseases of the Helmholtz Center Munich at the University of Tübingen (IDM), Partner in the German Center for Diabetes Research (DZD), Tübingen, Germany. susanne.ullrich@med.uni-tuebingen.de.
authors.forename: Susanne
authors.initials: S
authors.lastname: Ullrich
mesh_terms: D048948:14-3-3 Proteins / Q000235:genetics*; D021581:Active Transport, Cell Nucleus / Q000235:genetics; D000818:Animals; D002467:Cell Nucleus / Q000378:metabolism; D000071161:Forkhead Box Protein O1; D051858:Forkhead Transcription Factors / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D050417:Insulin-Secreting Cells / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D008822:Mice, Transgenic; D010766:Phosphorylation / Q000235:genetics; D061251:Primary Cell Culture; D051745:Protein Kinase C-delta / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction / Q000235:genetics
chemical_list: D048948:14-3-3 Proteins; D000071161:Forkhead Box Protein O1; D051858:Forkhead Transcription Factors; D051745:Protein Kinase C-delta; C000590975:Ywhaz protein, mouse; C467200:Foxo1 protein, mouse; C497259:Prkcd protein, mouse
keywords: 14-3-3ζ; Apoptosis; FOXO1; Insulin-secreting beta cells; JNK; PKCδ; Palmitate