KDEL-cargo regulates interactions between proteins involved in COPI vesicle traffic: measurements in living cells using FRET.
How the occupied KDEL receptor ERD2 is sorted into COPI vesicles for Golgi-to-ER transport is largely unknown. Here, interactions between proteins of the COPI transport machinery occurring during a "wave" of transport of a KDEL ligand were studied in living cells. FRET between CFP and YFP fusion proteins was measured by multifocal multiphoton microscopy and bulk-cell spectrofluorimetry. Ligand binding induces oligomerization of ERD2 and recruitment of ARFGAP to the Golgi, where the (ERD2)n/ARFGAP complex interacts with membrane-bound ARF1. During KDEL ligand transport, interactions of ERD2 with beta-COP and p23 decrease and the proteins segregate. Both p24a and p23 interact with ARF1, but only p24 interacts with ARFGAP. These findings suggest a model for how cargo-induced oligomerization of ERD2 regulates its sorting into COPI-coated buds.
Majoul
I
I
Department of Neurobiology, Max-Planck-Institute of Biophysical Chemistry, Göttingen, Germany.
Straub
M
M
Hell
S W
SW
Duden
R
R
Söling
H D
HD
D020823:ADP-Ribosylation Factor 1 / Q000378:metabolism; D020727:ADP-Ribosylation Factors / Q000378:metabolism; D000818:Animals; D002522:Chlorocebus aethiops; D020755:Coat Protein Complex I / Q000378:metabolism*; D020756:Coatomer Protein / Q000378:metabolism; D003593:Cytoplasm / Q000378:metabolism; D020690:GTPase-Activating Proteins / Q000378:metabolism; D006056:Golgi Apparatus / Q000378:metabolism; D008024:Ligands; D008565:Membrane Proteins / Q000378:metabolism; D010446:Peptide Fragments / Q000378:metabolism; D011485:Protein Binding / Q000502:physiology; D021381:Protein Transport / Q000502:physiology*; D018000:Receptors, Peptide / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D015203:Reproducibility of Results; D013050:Spectrometry, Fluorescence / Q000592:standards; D014709:Vero Cells
D020755:Coat Protein Complex I; D020756:Coatomer Protein; D020690:GTPase-Activating Proteins; C064459:KDEL receptor; D008024:Ligands; D008565:Membrane Proteins; D010446:Peptide Fragments; D018000:Receptors, Peptide; D011993:Recombinant Fusion Proteins; D020823:ADP-Ribosylation Factor 1; D020727:ADP-Ribosylation Factors