YeastARV1Is Required for Efficient Delivery of an Early GPI Intermediate to the First Mannosyltransferase during GPI Assembly and Controls Lipid Flow from the Endoplasmic Reticulum

Abstract

Glycosylphosphatidylinositol (GPI), covalently attached to many eukaryotic proteins, not only acts as a membrane anchor but is also thought to be a sorting signal for GPI-anchored proteins that are associated with sphingolipid and sterol-enriched domains. GPI anchors contain a core structure conserved among all species. The core structure is synthesized in two topologically distinct stages on the leaflets of the endoplasmic reticulum (ER). Early GPI intermediates are assembled on the cytoplasmic side of the ER and then are flipped into the ER lumen where a complete GPI precursor is synthesized and transferred to protein. The flipping process is predicted to be mediated by a protein referred as flippase; however, its existence has not been proven. Here we show that yeast Arv1p is an important protein required for the delivery of an early GPI intermediate, GlcN-acylPI, to the first mannosyltransferase of GPI synthesis in the ER lumen. We also provide evidence that ARV1 deletion and mutations in other proteins involved in GPI anchor synthesis affect inositol phosphorylceramide synthesis as well as the intracellular distribution and amounts of sterols, suggesting a role of GPI anchor synthesis in lipid flow from the ER.

The Putative Lipid Transporter, Arv1, Is Required for Activating Pheromone-Induced MAP Kinase Signaling in Saccharomyces cerevisiae

“…arv1 cells harbor defects in sphingolipid and GPI synthesis, and although their overall sterol content is normal, our data, as well as that of others, strongly suggest these lipids are trafficked improperly and thus are mislocalized (Tinkelenberg et al 2000;Swain et al 2002;Fei et al 2008;Kajiwara et al 2008). Membrane microdomains, also known as lipid rafts, are sphingolipidand sterol-rich regions of the plasma membrane, which are capable of mediating membrane sorting, cell adhesion, and signal transduction (Harder et al 1998).…”

“…Thus general vesicle-mediated trafficking is only slightly affected in arv1 cells and most likely does not contribute to the mating defects observed. In addition, the kinetics of a-factor internalization in mutant cells is similar to that seen in wild type, indicating that pheromone receptors are localized properly and are functional (Kajiwara et al 2008). Therefore, it is unlikely Ste5 mislocalization results from defects in general protein trafficking.…”

“…arv1 cells have a slight defect in the ER-to-Golgi transport of the GPI-anchored proteins Gas1 and Yps1, while there is no vacuolar trafficking defect in carboxypeptidase Y (CPY) (Kajiwara et al 2008). Thus general vesicle-mediated trafficking is only slightly affected in arv1 cells and most likely does not contribute to the mating defects observed.…”

“…It is known that Arv1 function is essential at high temperatures and in yeast mutants unable to esterify sterols (Tinkelenberg et al 2000). Moreover, arv1 cells harbor defects in sphingolipid metabolism (Swain et al 2002), glycosylphosphatidylinositol (GPI) biosynthesis (Kajiwara et al 2008), and may harbor defects in sterol trafficking (Tinkelenberg et al 2000;Fei et al 2008). It has been suggested that Arv1 plays a role in cholesterol trafficking from the ER to plasma membrane in mammalian cells (Tong et al 2010).…”

First Step of Glycosylphosphatidylinositol (GPI) Biosynthesis Cross-talks with Ergosterol Biosynthesis and Ras Signaling in Candida albicans

“…For example, in mammals the GPI-GnT complex has been shown to have regulatory contacts with Dpm2, a subunit that also regulates dolichol-phosphate mannose synthase (29). Likewise, in Saccharomyces cerevisiae, subunits of the GPI-GnT complex were shown to physically interact with and regulate Ras signaling (30,31), as well as control intracellular sterol distribution via genetic interactions with ARV1 (32). A large scale split ubiquitin-based protein-protein interaction study also indicated that ScGpi2 in S. cerevisiae interacted with ScGpi19 as well as with ScErg11 and a large number of other proteins involved in many different biochemical pathways of the cell (33).…”