title: Resveratrol-mediated autophagy requires WIPI-1-regulated LC3 lipidation in the absence of induced phagophore formation.
abstract: Canonical autophagy is positively regulated by the Beclin 1/phosphatidylinositol 3-kinase class III (PtdIns3KC3) complex that generates an essential phospholipid, phosphatidylinositol 3-phosphate (PtdIns(3)P), for the formation of autophagosomes. Previously, we identified the human WIPI protein family and found that WIPI-1 specifically binds PtdIns(3)P, accumulates at the phagophore and becomes a membrane protein of generated autophagosomes. Combining siRNA-mediated protein downregulation with automated high through-put analysis of PtdIns(3)P-dependent autophagosomal membrane localization of WIPI-1, we found that WIPI-1 functions upstream of both Atg7 and Atg5, and stimulates an increase of LC3-II upon nutrient starvation. Resveratrol-mediated autophagy was shown to enter autophagic degradation in a noncanonical manner, independent of Beclin 1 but dependent on Atg7 and Atg5. By using electron microscopy, LC3 lipidation and GFP-LC3 puncta-formation assays we confirmed these results and found that this effect is partially wortmannin-insensitive. In line with this, resveratrol did not promote phagophore localization of WIPI-1, WIPI-2 or the Atg16L complex above basal level. In fact, the presence of resveratrol in nutrient-free conditions inhibited phagophore localization of WIPI-1. Nevertheless, we found that resveratrol-mediated autophagy functionally depends on canonical-driven LC3-II production, as shown by siRNA-mediated downregulation of WIPI-1 or WIPI-2. From this it is tempting to speculate that resveratrol promotes noncanonical autophagic degradation downstream of the PtdIns(3)P-WIPI-Atg7-Atg5 pathway, by engaging a distinct subset of LC3-II that might be generated at membrane origins apart from canonical phagophore structures.
authors.affiliation: Autophagy Laboratory, Department of Molecular Biology, Interfaculty Institute for Cell Biology, University of Tuebingen, Tuebingen, Germany.
authors.forename: Mario
authors.initials: M
authors.lastname: Mauthe
authors.forename: Anke
authors.initials: A
authors.lastname: Jacob
authors.forename: Sandra
authors.initials: S
authors.lastname: Freiberger
authors.forename: Katharina
authors.initials: K
authors.lastname: Hentschel
authors.forename: York-Dieter
authors.initials: YD
authors.lastname: Stierhof
authors.forename: Patrice
authors.initials: P
authors.lastname: Codogno
authors.forename: Tassula
authors.initials: T
authors.lastname: Proikas-Cezanne
mesh_terms: D000730:Androstadienes / Q000494:pharmacology; D000818:Animals; D001343:Autophagy / Q000187:drug effects*; D000071192:Autophagy-Related Protein 12; D000071187:Autophagy-Related Protein 5; D000071193:Autophagy-Related Protein 7; D000071183:Autophagy-Related Proteins; D002352:Carrier Proteins / Q000187:drug effects / Q000378:metabolism*; D002460:Cell Line; D049452:Green Fluorescent Proteins / Q000378:metabolism; D006801:Humans; D008055:Lipids / Q000737:chemistry*; D008565:Membrane Proteins / Q000378:metabolism*; D051379:Mice; D008869:Microtubule-Associated Proteins / Q000378:metabolism*; D010588:Phagosomes / Q000187:drug effects / Q000378:metabolism* / Q000648:ultrastructure; D018129:Phosphatidylinositol Phosphates / Q000378:metabolism; D021381:Protein Transport / Q000187:drug effects; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D000077185:Resveratrol; D025841:Small Ubiquitin-Related Modifier Proteins / Q000378:metabolism; D013267:Stilbenes / Q000494:pharmacology*; D044764:Ubiquitin-Activating Enzymes / Q000378:metabolism; D000077191:Wortmannin
chemical_list: C466624:ATG12 protein, human; C502778:ATG5 protein, human; D000730:Androstadienes; C495773:Atg5 protein, mouse; C502661:Atg7 protein, mouse; D000071192:Autophagy-Related Protein 12; D000071187:Autophagy-Related Protein 5; D000071183:Autophagy-Related Proteins; D002352:Carrier Proteins; D008055:Lipids; C477012:MAP1LC3A protein, human; D008565:Membrane Proteins; D008869:Microtubule-Associated Proteins; D018129:Phosphatidylinositol Phosphates; D011993:Recombinant Fusion Proteins; D025841:Small Ubiquitin-Related Modifier Proteins; D013267:Stilbenes; C522812:WIPI1 protein, human; C426003:ZFYVE1 protein, human; C055525:phosphatidylinositol 3-phosphate; D049452:Green Fluorescent Proteins; C509349:ATG7 protein, human; D000071193:Autophagy-Related Protein 7; D044764:Ubiquitin-Activating Enzymes; D000077185:Resveratrol; D000077191:Wortmannin
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