Beta-COP is essential for transport of protein from the endoplasmic reticulum to the Golgi in vitro.
Using a novel in vitro assay which allows us to distinguish vesicle budding from subsequent targeting and fusion steps, we provide the first biological evidence that beta-COP, a component of non-clathrin-coated vesicles believed to mediate intraGolgi transport, is essential for transport of protein from the ER to the cis-Golgi compartment. Incubation in the presence of beta-COP specific antibodies and F(ab) fragments prevents the exit of vesicular stomatitis virus glycoprotein (VSV-G) from the ER. These results demonstrate that beta-COP is required for the assembly of coat complexes mediating vesicle budding. Fractionation of rat liver cytosol revealed that a major biologically active form of beta-COP was found in a high molecular pool (> 1,000 kD) distinct from coatomer and which promoted efficient vesicle budding from the ER. Surprisingly, rab1B could be quantitatively coprecipitated with this beta-COP containing complex and was also essential for function. We suggest that beta-COP functions in an early step during vesicle formation and that rab1B may be recruited as a component of a precoat complex which participates in the export of protein from the ER via vesicular carriers.
Department of Cell Biology, Scripps Research Institute, La Jolla, California 92037.
F
F
Peter
H
H
Plutner
H
H
Zhu
T E
TE
Kreis
W E
WE
Balch
D000818:Animals; D000906:Antibodies / Q000276:immunology / Q000494:pharmacology; D000918:Antibody Specificity; D000936:Antigen-Antibody Complex / Q000032:analysis; D001692:Biological Transport / Q000187:drug effects; D002458:Cell Fractionation; D002460:Cell Line; D020756:Coatomer Protein; D003600:Cytosol / Q000737:chemistry / Q000648:ultrastructure; D004721:Endoplasmic Reticulum / Q000737:chemistry / Q000502:physiology* / Q000648:ultrastructure; D019204:GTP-Binding Proteins / Q000032:analysis / Q000276:immunology / Q000502:physiology*; D006056:Golgi Apparatus / Q000737:chemistry / Q000502:physiology* / Q000648:ultrastructure; D007140:Immunoglobulin Fab Fragments / Q000494:pharmacology; D008099:Liver / Q000737:chemistry / Q000648:ultrastructure; D008562:Membrane Glycoproteins; D008869:Microtubule-Associated Proteins / Q000032:analysis / Q000276:immunology / Q000502:physiology*; D011233:Precipitin Tests; D011499:Protein Processing, Post-Translational / Q000502:physiology; D051381:Rats; D014759:Viral Envelope Proteins / Q000378:metabolism; D020761:rab1 GTP-Binding Proteins
D000906:Antibodies; D000936:Antigen-Antibody Complex; D020756:Coatomer Protein; C028391:G protein, vesicular stomatitis virus; D007140:Immunoglobulin Fab Fragments; D008562:Membrane Glycoproteins; D008869:Microtubule-Associated Proteins; D014759:Viral Envelope Proteins; D019204:GTP-Binding Proteins; C061569:RAB1B protein, rat; D020761:rab1 GTP-Binding Proteins