A putative heterotrimeric G protein inhibits the fusion of COPI-coated vesicles. Segregation of heterotrimeric G proteins from COPI-coated vesicles.
Heterotrimeric G proteins have been implicated in the regulation of intracellular protein transport, but their mechanism of action remains unclear. In vivo, secretion of chromogranin B, tagged with the green fluorescent protein, was inhibited by the addition of a general activator of trimeric G proteins (AlF4-) to stably transfected Vero cells and resulted in an accumulation of the tagged protein in the Golgi apparatus. In an in vitro assay that reconstitutes intra-Golgi protein transport, we find that a membrane-bound and AlF4--sensitive factor is involved in the fusion reaction. To determine whether this effect is mediated by a heterotrimeric G protein localized to COPI-coated transport vesicles, we determined the presence of G proteins on these vesicles and found that they were segregated relative to the donor membranes. Because G proteins do not have an obvious sorting, retention, or retrieval signal, we considered the possibility that other interactions might be responsible for this segregation. In agreement with this, we found that trimeric G proteins from isolated Golgi membranes were partially insoluble in Triton X-100. Identification of the proteins that interact with the heterotrimeric G proteins in the Golgi-derived detergent-insoluble complex might help to reveal the regulation of protein secretion mediated by heterotrimeric G proteins.
Helms
J B
JB
Biochemie-Zentrum Heidelberg (BZH), University of Heidelberg, Im Neuenheimer Feld 328, 69120 Heidelberg, Germany. Helms@urz.uni-heidelberg.de
Helms-Brons
D
D
Brügger
B
B
Gkantiragas
I
I
Eberle
H
H
Nickel
W
W
Nürnberg
B
B
Gerdes
H H
HH
Wieland
F T
FT
D017607:Aluminum Compounds / Q000494:pharmacology; D000818:Animals; D002460:Cell Line; D002864:Chromogranins / Q000378:metabolism; D018699:Coated Vesicles / Q000378:metabolism*; D020756:Coatomer Protein; D003902:Detergents / Q000494:pharmacology; D005459:Fluorides / Q000494:pharmacology; D019204:GTP-Binding Proteins / Q000378:metabolism*; D006056:Golgi Apparatus / Q000502:physiology; D049452:Green Fluorescent Proteins; D006160:Guanosine Triphosphate / Q000031:analogs & derivatives / Q000494:pharmacology; D008164:Luminescent Proteins / Q000235:genetics; D008561:Membrane Fusion / Q000502:physiology*; D008565:Membrane Proteins / Q000378:metabolism*; D008856:Microscopy, Fluorescence; D011487:Protein Conformation; D012995:Solubility; D014162:Transfection / Q000235:genetics
D017607:Aluminum Compounds; D002864:Chromogranins; D020756:Coatomer Protein; D003902:Detergents; D008164:Luminescent Proteins; D008565:Membrane Proteins; D049452:Green Fluorescent Proteins; D006160:Guanosine Triphosphate; D019204:GTP-Binding Proteins; D005459:Fluorides; C032311:aluminum fluoride