Binding of coatomer to Golgi membranes requires ADP-ribosylation factor.
Coatomer, a complex of seven proteins, appears to be the precursor of the coat structure of non-clathrin-coated Golgi-derived vesicles. Another component of this vesicle coat is the cytosolic protein ADP-ribosylation factor (ARF). Like coatomer, ARF appears to reversibly associate with Golgi membranes. We now report that ARF is required for coatomer binding to Golgi membranes and that myristoylated, but not non-myristoylated, ARF is the required species. We utilize an antibody directed against the beta-subunit of coatomer (beta-COP) to follow coatomer binding. ARF and beta-COP bind stoichiometrically to Golgi membranes. ARF-dependent beta-COP binding requires a membrane-associated protein, is saturable, and is enhanced in the presence of stable GTP analogues like guanosine 5'-O-(3-thiotriphosphate) (GTP gamma S). ARF and beta-COP bind sequentially to Golgi membranes, since beta-COP can be bound to reisolated membranes that had been previously incubated with ARF and GTP gamma S. We conclude that membrane-bound ARF confers to Golgi membranes all of the requirements for specific beta-COP binding.
Program in Cellular Biochemistry and Biophysics, Rockefeller Research Laboratory, Sloan-Kettering Institute, New York, New York 10021.
D J
DJ
Palmer
J B
JB
Helms
C J
CJ
Beckers
L
L
Orci
J E
JE
Rothman
D020727:ADP-Ribosylation Factors; D000818:Animals; D001921:Brain / Q000378:metabolism*; D002417:Cattle; D020756:Coatomer Protein; D003600:Cytosol / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D019204:GTP-Binding Proteins / Q000302:isolation & purification / Q000378:metabolism*; D006056:Golgi Apparatus / Q000378:metabolism*; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000494:pharmacology; D015151:Immunoblotting; D007425:Intracellular Membranes / Q000378:metabolism; D046911:Macromolecular Substances; D008565:Membrane Proteins / Q000302:isolation & purification / Q000378:metabolism*; D008869:Microtubule-Associated Proteins / Q000302:isolation & purification / Q000378:metabolism*; D008970:Molecular Weight; D019814:Myristic Acid; D009227:Myristic Acids / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism
D020756:Coatomer Protein; D046911:Macromolecular Substances; D008565:Membrane Proteins; D008869:Microtubule-Associated Proteins; D009227:Myristic Acids; D011994:Recombinant Proteins; D019814:Myristic Acid; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D019204:GTP-Binding Proteins; D020727:ADP-Ribosylation Factors