title: Decoupling of activation and effector binding underlies ARF6 priming of fast endocytic recycling.
abstract: The small GTP-binding protein ADP-ribosylation factor 6 (ARF6) controls the endocytic recycling pathway of several plasma membrane receptors. We analyzed the localization and GDP/GTP cycle of GFP-tagged ARF6 by total internal reflection fluorescent microscopy. We found that ARF6-GFP associates with clathrin-coated pits (CCPs) at the plasma membrane in a GTP-dependent manner in a mechanism requiring the adaptor protein complex AP-2. In CCP, GTP-ARF6 mediates the recruitment of the ARF-binding domain of downstream effectors including JNK-interacting proteins 3 and 4 (JIP3 and JIP4) after the burst recruitment of the clathrin uncoating component auxilin. ARF6 does not contribute to receptor-mediated clathrin-dependent endocytosis. In contrast, we found that interaction of ARF6 and JIPs on endocytic vesicles is required for trafficking of the transferrin receptor in the fast, microtubule-dependent endocytic recycling pathway. Our findings unravel a novel mechanism of separation of ARF6 activation and effector function, ensuring that fast recycling may be determined at the level of receptor incorporation into CCPs.
authors: Centre de Recherche, Institut Curie, CNRS, UMR 144, 26 rue d'Ulm, 75248 Paris Cedex 05, France. guillaume.montagnac@curie.fr
authors: Guillaume
authors: G
authors: Montagnac
authors: Hélène
authors: H
authors: de Forges
authors: Elizabeth
authors: E
authors: Smythe
authors: Charles
authors: C
authors: Gueudry
authors: Maryse
authors: M
authors: Romao
authors: Jean
authors: J
authors: Salamero
authors: Philippe
authors: P
authors: Chavrier
mesh_terms: D000089982:ADP-Ribosylation Factor 6; D020727:ADP-Ribosylation Factors / Q000378:metabolism*; D033962:Adaptor Protein Complex 2 / Q000378:metabolism; D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism*; D034021:Auxilins; D001692:Biological Transport; D022163:Clathrin-Coated Vesicles / Q000502:physiology*; D004705:Endocytosis; D055785:Gene Knockdown Techniques; D006367:HeLa Cells; D006801:Humans; D008856:Microscopy, Fluorescence; D009419:Nerve Tissue Proteins / Q000378:metabolism*; D011485:Protein Binding; D018160:Receptors, Cytoplasmic and Nuclear / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D022161:Transport Vesicles / Q000378:metabolism*
chemical_list: D000089982:ADP-Ribosylation Factor 6; D033962:Adaptor Protein Complex 2; D048868:Adaptor Proteins, Signal Transducing; D034021:Auxilins; C485750:MAPK8IP3 protein, human; D009419:Nerve Tissue Proteins; D018160:Receptors, Cytoplasmic and Nuclear; D011993:Recombinant Fusion Proteins; D020727:ADP-Ribosylation Factors; C000715348:ARF6 protein, human