Membrane metabolism mediated by Sec14 family members influences Arf GTPase activating protein activity for transport from the trans-Golgi.
The budding yeast Saccharomyces cerevisiae contains a family of Arf (ADP-ribosylation factor) GTPase activating protein (GAP) proteins with the Gcs1 + Age2 ArfGAP pair providing essential overlapping function for the movement of transport vesicles from the trans-Golgi network. We have generated a temperature-sensitive but stable version of the Gcs1 protein that is impaired only for trans-Golgi transport and find that deleterious effects of this enfeebled Gcs1-4 mutant protein are relieved by increased gene dosage of the gcs1-4 mutant gene itself or by the SFH2 gene (also called CSR1), encoding a phosphatidylinositol transfer protein (PITP). This effect was not seen for the SEC14 gene, encoding the founding member of the yeast PITP protein family, even though the Gcs1 and Age2 ArfGAPs are known to be downstream effectors of Sec14-mediated activity for trans-Golgi transport. Sfh2-mediated suppression of inadequate Gcs1-4 function depended on phospholipase D, whereas inadequate Gcs1-4 activity was relieved by increasing levels of diacylglycerol (DAG). Recombinant Gcs1 protein was found to bind certain phospholipids but not DAG. Our findings favor a model of Gcs1 localization through binding to specific phospholipids and activation of ArfGAP activity by DAG-mediated membrane curvature as the transport vesicle is formed. Thus, ArfGAPs are subject to both temporal and spatial regulation that is facilitated by Sfh2-mediated modulation of the lipid environment.
Wong
Tania A
TA
Department of Microbiology and Immunology, Faculty of Medicine, Dalhousie University, Halifax, NS, Canada B3H 1X5.
Fairn
Gregory D
GD
Poon
Pak P
PP
Shmulevitz
Maya
M
McMaster
Christopher R
CR
Singer
Richard A
RA
Johnston
Gerald C
GC
D020727:ADP-Ribosylation Factors / Q000235:genetics / Q000378:metabolism*; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D004075:Diglycerides / Q000737:chemistry / Q000494:pharmacology; D004789:Enzyme Activation; D020690:GTPase-Activating Proteins / Q000235:genetics / Q000378:metabolism; D015966:Gene Expression Regulation, Fungal / Q000187:drug effects / Q000235:genetics; D006056:Golgi Apparatus / Q000187:drug effects / Q000378:metabolism*; D009154:Mutation / Q000235:genetics; D010739:Phospholipase D / Q000235:genetics / Q000378:metabolism; D049294:Phospholipid Transfer Proteins / Q000145:classification / Q000235:genetics / Q000378:metabolism*; D021381:Protein Transport / Q000187:drug effects; D011730:Pyridoxal / Q000031:analogs & derivatives / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000166:cytology* / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000145:classification / Q000235:genetics / Q000378:metabolism*; D013696:Temperature
C483430:CSR1 protein, S cerevisiae; D004268:DNA-Binding Proteins; D004075:Diglycerides; C089147:GCS1 protein, S cerevisiae; D020690:GTPase-Activating Proteins; D049294:Phospholipid Transfer Proteins; C483438:SEC14 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D011730:Pyridoxal; C041277:1-dodecylpyridoxal; D010739:Phospholipase D; C471177:SPO14 protein, S cerevisiae; D020727:ADP-Ribosylation Factors