title: Hypertonic stress promotes autophagy and microtubule-dependent autophagosomal clusters.
abstract: Osmotic homeostasis is fundamental for most cells, which face recurrent alterations of environmental osmolality that challenge cell viability. Protein damage is a consequence of hypertonic stress, but whether autophagy contributes to the osmoprotective response is unknown. Here, we investigated the possible implications of autophagy and microtubule organization on the response to hypertonic stress. We show that hypertonicity rapidly induced long-lived protein degradation, LC3-II generation and Ptdlns3K-dependent formation of LC3- and ATG12-positive puncta. Lysosomotropic agents chloroquine and bafilomycin A 1, but not nutrient deprivation or rapamycin treatment, further increased LC3-II generation, as well as ATG12-positive puncta, indicating that hypertonic stress increases autophagic flux. Autophagy induction upon hypertonic stress enhanced cell survival since cell death was increased by ATG12 siRNA-mediated knockdown and reduced by rapamycin. We additionally showed that hypertonicity induces fast reorganization of microtubule networks, which is associated with strong reorganization of microtubules at centrosomes and fragmentation of Golgi ribbons. Microtubule remodeling was associated with pericentrosomal clustering of ATG12-positive autolysosomes that colocalized with SQSTM1/p62 and ubiquitin, indicating that autophagy induced by hypertonic stress is at least partly selective. Efficient autophagy by hypertonic stress required microtubule remodeling and was DYNC/dynein-dependent as autophagosome clustering was enhanced by paclitaxel-induced microtubule stabilization and was reduced by nocodazole-induced tubulin depolymerization as well as chemical (EHNA) or genetic [DCTN2/dynactin 2 (p50) overexpression] interference of DYNC activity. The data document a general and hitherto overlooked mechanism, where autophagy and microtubule remodeling play prominent roles in the osmoprotective response.
authors: Department of Cellular Physiology and Metabolism, University of Geneva, Geneva, Switzerland.
authors: Paula
authors: P
authors: Nunes
authors: Thomas
authors: T
authors: Ernandez
authors: Isabelle
authors: I
authors: Roth
authors: Xiaomu
authors: X
authors: Qiao
authors: Déborah
authors: D
authors: Strebel
authors: Richard
authors: R
authors: Bouley
authors: Anne
authors: A
authors: Charollais
authors: Pierluigi
authors: P
authors: Ramadori
authors: Michelangelo
authors: M
authors: Foti
authors: Paolo
authors: P
authors: Meda
authors: Eric
authors: E
authors: Féraille
authors: Dennis
authors: D
authors: Brown
authors: Udo
authors: U
authors: Hasler
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism; D000818:Animals; D001343:Autophagy / Q000187:drug effects*; D002467:Cell Nucleus / Q000187:drug effects / Q000378:metabolism; D002470:Cell Survival / Q000187:drug effects; D000072159:Dynactin Complex; D004398:Dyneins / Q000378:metabolism; D006801:Humans; D006982:Hypertonic Solutions / Q000494:pharmacology*; D018374:LLC-PK1 Cells; D008247:Lysosomes / Q000187:drug effects / Q000378:metabolism; D008869:Microtubule-Associated Proteins / Q000378:metabolism; D008870:Microtubules / Q000187:drug effects / Q000378:metabolism*; D010588:Phagosomes / Q000187:drug effects / Q000378:metabolism*; D013312:Stress, Physiological / Q000187:drug effects*; D013552:Swine
chemical_list: D048868:Adaptor Proteins, Signal Transducing; C000606361:DCTN2 protein, human; D000072159:Dynactin Complex; D006982:Hypertonic Solutions; D008869:Microtubule-Associated Proteins; D004398:Dyneins
keywords: ATG12; Golgi apparatus; MAP1LC3/LC3; MAPRE1/EB1; MTOC; SQSTM1/p62; apoptosis; autophagy; dynein; lysosome; microtubule; osmotic stress; proteasome; rapamycin; tubulin; ubiquitin
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