Positive roles for specific lipid-regulated ARFGAPs in Sec14p-dependent Golgi secretory function in yeast.

摘要

Sec14p is the major phosphatidylinositol-transfer protein (PITP) in yeast. To understand how PITPs contribute to cell function, we have undertaken a detailed study of this prototypic PITP. Sec14p is an essential component of the Saccharomyces cerevisiae secretory pathway. Cells with dysfunctional Sec14p exhibit a late Golgi secretory block and subsequently die. Our current hypothesis for Sec14p function is that Sec14p promotes secretory vesicle budding from the Golgi by regulating lipid metabolism at the trans-Golgi surface. Sec14p-regulated lipids might then serve as a docking site for proteins involved in vesicle bud formation.; In the past, much of our attention focused on how Sec14p regulates lipid metabolism and which lipid metabolic pathways are subject to Sec14p control. The requirement for Sec14p can be bypassed by loss-of-function mutations in other genes. Characterization of such ‘bypass Sec14p’ mutants has suggested that they function by altering lipids levels in ways that somewhat mimic Sec14p. All cases of ‘bypass Sec14p’ are accompanied by the essential activation of phospholipase D, an enzyme that converts phosphatidylcholine (PC) to phosphatidic acid (PA). Collectively, genetic analyses have identified diacylglycerol, PC, PA and inositol-containing phospholipids as potential Sec14p-regulated lipids that influence Golgi secretory function.; Our model predicts the existence of a lipid sensor that links Sec14p's regulation of lipid metabolism to downstream components of the secretory pathway. Herein, we describe the novel involvement of two adenosine diphosphate-ribosylation factor (ARF) guanosine triphosphatase-activating proteins (ARFGAPs), Gcs1p and Age2p, in the Sec14p pathway. Genetic and biochemical analyses indicate that Gcs1p and Age2p are an essential pair of downstream Sec14p effector proteins that are sensitive to Sec14p-regulated lipids and are required for proper Golgi secretory function. These data also indicate a positive role for ARFGAPs in promoting secretory vesicle biogenesis and provide the basis for a novel interpretation of ARF-mediated protein transport.