Dissecting the role of the ARF guanine nucleotide exchange factor GBF1 in Golgi biogenesis and protein trafficking.
COPI recruitment to membranes appears to be essential for the biogenesis of the Golgi and for secretory trafficking. Preventing COPI recruitment by expressing inactive forms of the ADP-ribosylation factor (ARF) or the ARF-activating guanine nucleotide exchange factor GBF1, or by treating cells with brefeldin A (BFA), causes the collapse of the Golgi into the endoplasmic reticulum (ER) and arrests trafficking of soluble and transmembrane proteins at the ER. Here, we assess COPI function in Golgi biogenesis and protein trafficking by preventing COPI recruitment to membranes by removing GBF1. We report that siRNA-mediated depletion of GBF1 causes COPI dispersal but does not lead to collapse of the Golgi. Instead, it causes extensive tubulation of the cis-Golgi. The Golgi-derived tubules target to peripheral ER-Golgi intermediate compartment (ERGIC) sites and create dynamic continuities between the ERGIC and the cis-Golgi compartment. COPI dispersal in GBF1-depleted cells causes dramatic inhibition of the trafficking of transmembrane proteins. Unexpectedly, soluble proteins continue to be secreted from GBF1-depleted cells. Our findings suggest that a secretory pathway capable of trafficking soluble proteins can be maintained in cells in which COPI recruitment is compromised by GBF1 depletion. However, the trafficking of transmembrane proteins through the existing pathway requires GBF1-mediated ARF activation and COPI recruitment.
Szul
Tomasz
T
Department of Cell Biology, University of Alabama at Birmingham, 1918 University Blvd, Birmingham, AL 35924, USA.
Grabski
Robert
R
Lyons
Susan
S
Morohashi
Yuichi
Y
Shestopal
Svetlana
S
Lowe
Martin
M
Sztul
Elizabeth
E
D020823:ADP-Ribosylation Factor 1 / Q000378:metabolism; D020727:ADP-Ribosylation Factors / Q000378:metabolism*; D002451:Cell Compartmentation; D002465:Cell Movement; D020755:Coat Protein Complex I / Q000378:metabolism; D004721:Endoplasmic Reticulum / Q000378:metabolism; D006056:Golgi Apparatus / Q000378:metabolism*; D020662:Guanine Nucleotide Exchange Factors / Q000172:deficiency / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D008565:Membrane Proteins / Q000378:metabolism; D008954:Models, Biological; D011485:Protein Binding; D021381:Protein Transport; D012995:Solubility
D020755:Coat Protein Complex I; C120445:GBF1 protein, human; D020662:Guanine Nucleotide Exchange Factors; D008565:Membrane Proteins; D020823:ADP-Ribosylation Factor 1; D020727:ADP-Ribosylation Factors; C116688:ARF4 protein, human