Membrane metabolism mediated by Sec14 family members influences Arf GTPase activating protein activity for transport from the trans-Golgi

• Published in: Proceedings of the National Academy of Sciences - PNAS Vol. 102; no. 36; pp. 12777 - 12782
• Main Authors: Wong, T.A, Fairn, G.D, Poon, P.P, Shmulevitz, M, McMaster, C.R, Singer, R.A, Johnston, G.C
• Format: Journal Article
• Language: English
• Published: United States National Academy of Sciences 06.09.2005; National Acad Sciences
• Subjects: Adenosine diphosphate; ADP-ribosylation factor; ADP-Ribosylation Factors - genetics; ADP-Ribosylation Factors - metabolism; Biological Sciences; Cell growth; Cell Membrane - drug effects; Cell Membrane - metabolism; cell membranes; Cellular biology; Diglycerides - chemistry; Diglycerides - pharmacology; DNA-Binding Proteins - genetics; DNA-Binding Proteins - metabolism; Enzyme Activation; fungal proteins; G-proteins; Gene dosage; Gene Expression Regulation, Fungal - drug effects; Gene Expression Regulation, Fungal - genetics; Genetic mutation; Golgi apparatus; Golgi Apparatus - drug effects; Golgi Apparatus - metabolism; GTPase-Activating Proteins - genetics; GTPase-Activating Proteins - metabolism; guanosinetriphosphatase; guanosinetriphosphatase activating protein; Lipids; Membranes; Metabolism; Mutant proteins; Mutation - genetics; organelles; P branes; phosphatidylinositol transfer protein; phospholipase D; Phospholipase D - genetics; Phospholipase D - metabolism; phospholipid transfer protein; Phospholipid Transfer Proteins - classification; Phospholipid Transfer Proteins - genetics; Phospholipid Transfer Proteins - metabolism; Plasmids; protein transport; Protein Transport - drug effects; Proteins; Pyridoxal - analogs & derivatives; Pyridoxal - metabolism; Saccharomyces cerevisiae; Saccharomyces cerevisiae - cytology; Saccharomyces cerevisiae - drug effects; Saccharomyces cerevisiae - genetics; Saccharomyces cerevisiae - metabolism; Saccharomyces cerevisiae Proteins - classification; Saccharomyces cerevisiae Proteins - genetics; Saccharomyces cerevisiae Proteins - metabolism; Secretion; Temperature; Trans golgi network; transport vesicles; Yeasts
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.0506156102

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.