title: Class III phosphoinositide 3-kinase/VPS34 and dynamin are critical for apical endocytic recycling.
abstract: Recycling is a limiting step for receptor-mediated endocytosis. We first report three in vitro or in vivo evidences that class III PI3K/VPS34 is the key PI3K isoform regulating apical recycling. A substractive approach, comparing in Opossum Kidney (OK) cells a pan-class I/II/III PI3K inhibitor (LY294002) with a class I/II PI3K inhibitor (ZSTK474), suggested that class III PI3K/VPS34 inhibition induced selective apical endosome swelling and sequestration of the endocytic receptor, megalin/LRP-2, causing surface down-regulation. GFP-(FYVE)x2 overexpression to sequester PI(3)P caused undistinguishable apical endosome swelling. In mouse kidney proximal tubular cells, conditional Vps34 inactivation also led to vacuolation and intracellular megalin redistribution. We next report that removal of LY294002 from LY294002-treated OK cells induced a spectacular burst of recycling tubules and restoration of megalin surface pool. Acute triggering of recycling tubules revealed recruitment of dynamin-GFP and dependence of dynamin-GTPase, guidance directionality by microtubules, and suggested that a microfilamentous net constrained endosomal swelling. We conclude that (i) besides its role in endosome fusion, PI3K-III is essential for endosome fission/recycling; and (ii) besides its role in endocytic entry, dynamin also supports tubulation of recycling endosomes. The unleashing of recycling upon acute reversal of PI3K inhibition may help study its dynamics and associated machineries.
authors.affiliation: CELL Unit, Université catholique de Louvain & de Duve Institute, Brussels, Belgium.
authors.forename: Sarah
authors.initials: S
authors.lastname: Carpentier
authors.forename: Francisca
authors.initials: F
authors.lastname: N'Kuli
authors.forename: Giuseppina
authors.initials: G
authors.lastname: Grieco
authors.forename: Patrick
authors.initials: P
authors.lastname: Van Der Smissen
authors.forename: Virginie
authors.initials: V
authors.lastname: Janssens
authors.forename: Hervé
authors.initials: H
authors.lastname: Emonard
authors.forename: Benoît
authors.initials: B
authors.lastname: Bilanges
authors.forename: Bart
authors.initials: B
authors.lastname: Vanhaesebroeck
authors.forename: Héloïse P
authors.initials: HP
authors.lastname: Gaide Chevronnay
authors.forename: Christophe E
authors.initials: CE
authors.lastname: Pierreux
authors.forename: Donatienne
authors.initials: D
authors.lastname: Tyteca
authors.forename: Pierre J
authors.initials: PJ
authors.lastname: Courtoy
mesh_terms: D000818:Animals; D018929:Cell Culture Techniques; D002867:Chromones / Q000494:pharmacology; D058546:Class III Phosphatidylinositol 3-Kinases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D034281:Dynamins / Q000378:metabolism*; D004705:Endocytosis; D011992:Endosomes / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D007294:Inositol / Q000031:analogs & derivatives / Q000494:pharmacology; D026561:Low Density Lipoprotein Receptor-Related Protein-2 / Q000235:genetics / Q000378:metabolism; D051379:Mice; D009025:Morpholines / Q000494:pharmacology; D009893:Opossums
chemical_list: C477931:1L-6-hydroxymethyl-chiro-inositol 2(R)-2-O-methyl-3-O-octadecylcarbonate; D002867:Chromones; D004791:Enzyme Inhibitors; D026561:Low Density Lipoprotein Receptor-Related Protein-2; D009025:Morpholines; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; D007294:Inositol; D058546:Class III Phosphatidylinositol 3-Kinases; D034281:Dynamins
keywords: LY294002; PI3K class III; VPS34; ZSTK474; dynamin; kidney; megalin; recycling; tubulation; wortmannin