The VRG4 Gene Is Required for GDP-mannose Transport into the Lumen of the Golgi in the Yeast, Saccharomyces cerevisiae

Abstract

In the yeast Saccharomyces cerevisiae, glycoproteins and sphingolipids are modified in the Golgi by the addition of mannose residues. The critical mannosyl donor for these reactions is the nucleotide sugar, GDP-mannose, whose transport into the Golgi from the cytoplasm is required for mannosylation. This transport reaction has been well characterized, but the nucleotide sugar transporter has yet to be identified in yeast. VRG4 is an essential gene whose product is required for a number of Golgi-specific functions, including glycosylation and the organization of the endomembrane system. Here, data are presented that demonstrate that the primary role of Vrg4p is in the transport of GDP-mannose into the Golgi. The vrg4 mutation causes a general impairment in mannosylation, affecting N-linked and O-linked glycoprotein modifications as well as the mannosylation of sphingolipids. By using an in vitro assay, vrg4 mutants were shown to be specifically defective in the transport of GDP-mannose into Golgi vesicles. The Vrg4 protein localizes to the Golgi complex in a pattern that suggests a wide distribution throughout the Golgi. Vrg4p displays homology to other putative nucleotide sugar transporters, suggesting that the VRG4 gene encodes a Golgi GDP-mannose transporter. As Vrg4p is essential, these results suggest that a complete lack of mannosylation of glycoproteins in the Golgi leads to inviability. Alternatively, the essential function of Vrg4p in yeast involves its effect on sphingolipids, which would imply a critical role for mannosylinositol phosphorylceramides or mannosyl diphosphoinositol ceramides on growth and viability.

Architecture and Biosynthesis of the Saccharomyces cerevisiae Cell Wall

“…Transport activities for UDP--Glc, UDP--GlcNAc, and UDP--Gal also occur in S. cerevisiae (Roy et al 1998;Castro et al 1999, and there are eight further candidate transporters (Dean et al 1997;Esther et al 2008), a couple of which have been associated with these transport activities. Some of the transporters may have specificity for more than one sugar nucleotide.…”

“…GMP, generated from GDP formed in Man-T reactions by GDPase activity, serves as antiporter. Vrg4/ Vig4 is essential, and vrg4 mutants are defective in mannosylation of N-and O-linked glycans and mannosyl inositolphosphoceramides (Dean et al 1997;Abe et al 1999).…”

“…Yea4 was characterized as an ER--localized UDP--GlcNAc transporter and its deletion impacts chitin synthesis ; Section V). Of the other transport defect (Dean et al 1997). …”

“…Other sugar nucleotide transport activities: Transport activities for UDP-Glc, UDP-GlcNAc, and UDP-Gal also occur in S. cerevisiae (Roy et al 1998Castro et al 1999), and there are eight more candidate transporters (Dean et al 1997;Esther et al 2008) whose functions are unclear. UDPGlc transport activity is present in the ER (Castro et al 1999), and one possible need for it might be for a glucosylation reaction at an early stage of b1,6-glucan assembly (see b1, 6-Glucan).…”

Inhibition of Golgi Apparatus Glycosylation Causes Endoplasmic Reticulum Stress and Decreased Protein Synthesis

“…Developmental phenotypes caused by mutations in nucleotide sugar transporters have been described in Drosophila (2,7), Caenorhabditis elegans (1, 2), Leishmania (1, 2, 8 -10), and yeast (1,2,11).…”