Cytosol-derived proteins are sufficient for Arp2/3 recruitment and ARF/coatomer-dependent actin polymerization on Golgi membranes.
The actin cytoskeleton has been implicated in protein trafficking at the Golgi apparatus and in Golgi orientation and morphology. Actin dynamics at the Golgi are regulated in part by recruiting Cdc42 or Rac to the membrane through a binding interaction with the coatomer-coated (COPI)-vesicle coat protein, coatomer. This leads to actin polymerization through the effector, N-WASP and the Arp2/3 complex. Here, we have used reconstitution of vesicle budding to test whether Arp2/3 is recruited to membranes during the formation of COPI vesicles. Our results revealed that ARF1 activation leads to greatly increased Arp3 levels on the membranes. Coatomer-bound Cdc42 and pre-existing F-actin are important for Arp2/3 binding. ARF1-dependent Arp2/3 recruitment and actin polymerization can be reconstituted on liposomal membranes, indicating that no membrane proteins are necessary. These results show that activated ARF1 can stimulate Arp2/3 recruitment to Golgi membranes through coatomer, Cdc42 or Rac, and N-WASP.
Chen
Ji-Long
JL
Department of Physiology and Biophysics, Roy J. and Lucille A. Carver College of Medicine, The University of Iowa, Iowa City, IA 52242, USA.
Lacomis
Lynne
L
Erdjument-Bromage
Hediye
H
Tempst
Paul
P
Stamnes
Mark
M
D020823:ADP-Ribosylation Factor 1 / Q000235:genetics / Q000378:metabolism*; D051377:Actin-Related Protein 2; D051378:Actin-Related Protein 3; D000199:Actins / Q000737:chemistry* / Q000378:metabolism*; D000818:Animals; D001426:Bacterial Proteins; D001427:Bacterial Toxins / Q000378:metabolism; D001921:Brain / Q000378:metabolism; D022181:COP-Coated Vesicles / Q000378:metabolism*; D002417:Cattle; D020756:Coatomer Protein / Q000737:chemistry / Q000378:metabolism; D003598:Cytoskeletal Proteins / Q000378:metabolism*; D003600:Cytosol / Q000378:metabolism; D003603:Cytotoxins / Q000378:metabolism; D006056:Golgi Apparatus / Q000378:metabolism*; D007425:Intracellular Membranes / Q000378:metabolism*; D008081:Liposomes / Q000378:metabolism; D008099:Liver / Q000648:ultrastructure; D009419:Nerve Tissue Proteins / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding; D011817:Rabbits; D051381:Rats; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D051317:Wiskott-Aldrich Syndrome Protein, Neuronal; D020764:cdc42 GTP-Binding Protein / Q000737:chemistry / Q000378:metabolism
D051377:Actin-Related Protein 2; D051378:Actin-Related Protein 3; D000199:Actins; D001426:Bacterial Proteins; D001427:Bacterial Toxins; D020756:Coatomer Protein; D003598:Cytoskeletal Proteins; D003603:Cytotoxins; D008081:Liposomes; D009419:Nerve Tissue Proteins; D011994:Recombinant Proteins; C495770:Wasl protein, rat; D051317:Wiskott-Aldrich Syndrome Protein, Neuronal; C031815:toxB protein, Clostridium difficile; D020823:ADP-Ribosylation Factor 1; D020764:cdc42 GTP-Binding Protein