title: Secreted autotransporter toxin (Sat) triggers autophagy in epithelial cells that relies on cell detachment.
abstract: The secreted autotransporter toxin, Sat, which belongs to the subfamily of serine protease autotransporters of Enterobacteriaceae, acts as a virulence factor in extraintestinal and intestinal pathogenic strains of Escherichia coli. We observed that HeLa cells exposed to the cell-free culture supernatant of recombinant strain AAEC185p(Sat-IH11128) producing the Sat toxin (CFCS(Sat) ), displayed dramatic disorganization of the F-actin cytoskeleton before loosening cell-to-cell junctions and detachment. Examination of the effect of Sat on GFP-microtubule-associated protein light chain 3 (LC3) HeLa cells revealed that CFCS(Sat) -induced autophagy follows CFCS(Sat) -induced F-actin cytoskeleton rearrangement. The induced autophagy shows an acceleration of the autophagy flux soon after Sat treatment, followed later by a blockade of the flux leading to the accumulation of large GFP-LC3-positive vacuoles in the cell cytoplasm. CFCS(Sat) did not induce cell detachment in autophagy-deficient mouse embryonic fibroblasts in contrast with wild-type mouse embryonic fibroblasts. The CFCS(Sat) -induced large GFP-LC3 dots do not display the characteristics of autophagolysosomes including expression of cathepsin D and Lamp-1 and 2 proteins, and Lysotracker Red- and DQ-BSA-positive labelling. We provide evidences that CFCS(Sat) -induced autophagy is not a cell response intended to get rid of the intracellular toxin. By a pharmacological blockers approach, we found that the blockade of Erk1/2 and p38 MAPKs, but not JNK, inhibited the CFCS(Sat) -induced autophagy and cell detachment whereas phosphatidylinositol-3 kinase blockers inhibiting canonical autophagy were inactive. When attached CFCS(Sat) -treated cells start to detach they showed caspase-independent cell death and rearrangements of the focal adhesion-associated vinculin and paxillin. Collectively, our results support that Sat triggers autophagy in epithelial cells that relies on its cell-detachment effect.
authors.affiliation: INSERM, UMR 756 Signalisation and Physiopathology of Epithelial cells, IFR 141 Plateforme Imagerie cellulaire Université Paris-Sud, Faculté de Pharmacie, Châtenay-Malabry, France.
authors.forename: Vanessa
authors.initials: V
authors.lastname: Liévin-Le Moal
authors.forename: Yannick
authors.initials: Y
authors.lastname: Comenge
authors.forename: Vincent
authors.initials: V
authors.lastname: Ruby
authors.forename: Raymonde
authors.initials: R
authors.lastname: Amsellem
authors.forename: Valérie
authors.initials: V
authors.lastname: Nicolas
authors.forename: Alain L
authors.initials: AL
authors.lastname: Servin
mesh_terms: D000818:Animals; D001343:Autophagy; D001427:Bacterial Toxins; D002448:Cell Adhesion; D003599:Cytoskeleton / Q000378:metabolism; D004847:Epithelial Cells / Q000382:microbiology* / Q000502:physiology*; D004926:Escherichia coli / Q000472:pathogenicity*; D029968:Escherichia coli Proteins / Q000378:metabolism*; D005347:Fibroblasts / Q000382:microbiology; D006367:HeLa Cells; D006801:Humans; D051379:Mice; D015398:Signal Transduction; D037521:Virulence Factors / Q000378:metabolism*
chemical_list: D001427:Bacterial Toxins; D029968:Escherichia coli Proteins; D037521:Virulence Factors; C506813:secreted autotransporter toxin, E coli