Protein networks supporting AP-3 function in targeting lysosomal membrane proteins.
The AP-3 adaptor complex targets selected transmembrane proteins to lysosomes and lysosome-related organelles. We reconstituted its preferred interaction with liposomes containing the ADP ribosylation factor (ARF)-1 guanosine triphosphatase (GTPase), specific cargo tails, and phosphatidylinositol-3 phosphate, and then we performed a proteomic screen to identify new proteins supporting its sorting function. We identified approximately 30 proteins belonging to three networks regulating either AP-3 coat assembly or septin polymerization or Rab7-dependent lysosomal transport. RNA interference shows that, among these proteins, the ARF-1 exchange factor brefeldin A-inhibited exchange factor 1, the ARF-1 GTPase-activating protein 1, the Cdc42-interacting Cdc42 effector protein 4, an effector of septin-polymerizing GTPases, and the phosphatidylinositol-3 kinase IIIC3 are key components regulating the targeting of lysosomal membrane proteins to lysosomes in vivo. This analysis reveals that these proteins, together with AP-3, play an essential role in protein sorting at early endosomes, thereby regulating the integrity of these organelles.
Baust
Thorsten
T
Biotechnological Center, Dresden University of Technology, 01307 Dresden, Germany.
Anitei
Mihaela
M
Czupalla
Cornelia
C
Parshyna
Iryna
I
Bourel
Line
L
Thiele
Christoph
C
Krause
Eberhard
E
Hoflack
Bernard
B
D020823:ADP-Ribosylation Factor 1 / Q000378:metabolism; D033963:Adaptor Protein Complex 3 / Q000378:metabolism*; D003598:Cytoskeletal Proteins / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D008081:Liposomes / Q000737:chemistry / Q000378:metabolism; D052118:Lysosomal-Associated Membrane Protein 1 / Q000378:metabolism; D051907:Lysosomal Membrane Proteins / Q000378:metabolism*; D018129:Phosphatidylinositol Phosphates / Q000378:metabolism; D011485:Protein Binding; D021382:Protein Sorting Signals; D021381:Protein Transport; D040901:Proteomics; D034741:RNA, Small Interfering / Q000378:metabolism; D013816:Thermodynamics; D020691:rab GTP-Binding Proteins / Q000378:metabolism; D000090802:rab7 GTP-Binding Proteins
D033963:Adaptor Protein Complex 3; D003598:Cytoskeletal Proteins; D008081:Liposomes; D052118:Lysosomal-Associated Membrane Protein 1; D051907:Lysosomal Membrane Proteins; D018129:Phosphatidylinositol Phosphates; D021382:Protein Sorting Signals; D034741:RNA, Small Interfering; C055525:phosphatidylinositol 3-phosphate; D000090802:rab7 GTP-Binding Proteins; C000716463:rab7 GTP-binding proteins, human; D020823:ADP-Ribosylation Factor 1; D020691:rab GTP-Binding Proteins