In vivo assembly of coatomer, the COP-I coat precursor.
Coatomer, a seven-subunit hetero-oligomeric complex, is the major component of the COP-I membrane coat of transport vesicles of the early secretory pathway. We have followed the assembly of this complex in vivo by pulse-chase experiments and immunoprecipitation of native coatomer subunits and found that it is an ordered process that takes 1-2 h to complete. During assembly, direct interactions between alpha-, beta'- and delta-COP, beta- and delta-COP, and gamma-, zeta-, and delta-COP occur. Coatomer, once it has assembled, is stable with a half-life of approximately 28 h. No significant amounts of partial coatomer complexes have been detected. The only subunit to exist at steady state out of the complex is zeta-COP, which has a similar half-life to coatomer subunits within the complex. Assembly is inhibited by brefeldin A, suggesting that it may be a regulated process. These results describe for the first time in vivo assembly of a coat protein complex involved in membrane traffic and extend our knowledge of how coatomer is structured.
Lowe
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Département de Biologie Cellulaire, Université de Genève, Sciences III, 30, quai Ernest-Ansermet, CH-1211 Geneva, Switzerland. kreis@sc2a.unige.ch
Kreis
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D000818:Animals; D001692:Biological Transport; D020126:Brefeldin A; D002522:Chlorocebus aethiops; D020756:Coatomer Protein; D003517:Cyclopentanes / Q000494:pharmacology; D007158:Immunologic Techniques; D007425:Intracellular Membranes / Q000378:metabolism; D046911:Macromolecular Substances; D008565:Membrane Proteins / Q000737:chemistry / Q000378:metabolism*; D008869:Microtubule-Associated Proteins / Q000378:metabolism; D011485:Protein Binding; D014709:Vero Cells
D020756:Coatomer Protein; D003517:Cyclopentanes; D046911:Macromolecular Substances; D008565:Membrane Proteins; D008869:Microtubule-Associated Proteins; D020126:Brefeldin A