A pathway for cell wall anchorage of Saccharomyces cerevisiae alpha-agglutinin.

Abstract

Saccharomyces cerevisiae ot-agglutinin is a cell wall-anchored adhesion glycoprotein. The previously identified 140-kDa form, which contains a glycosyl-phosphatidylinositol (GPI) anchor (D. Wojciechowicz, C.-F. Lu, J. Kurjan, and P. N. Lipke, Mol. Cell. Biol. 13:2554-2563, 1993, and additional forms of 80, 150, 250 to 300, and >300 kDa had the properties of intermediates in a transport and cell wall anchorage pathway. N glycosylation and additional modifications resulted in successive increases in size during transport. The 150-and 250-to 300-kDa forms were membrane associated and are likely to be intermediates between the 140-kDa form and a cell surface GPI-anchored form of >300 kDa. A soluble form of >300 kDa that lacked the GPI anchor had properties of a periplasmic intermediate between the plasma membrane form and the >300-kDa cell wall-anchored form. These results constitute experimental support for the hypothesis that GPI anchors act to localize a-agglutinin to the plasma membrane and that cell wall anchorage involves release from the GPI anchor to produce a periplasmic intermediate followed by linkage to the cell wall.The sexual agglutinins of Saccharomyces cerevisiae are complementary cell surface glycoproteins expressed by a and (x haploid cells. Agglutinin levels are increased following treatment with the pheromone produced by cells of the opposite mating type (39,48,50), and binding between the ax-agglutinin on at cells and the a-agglutinin on a cells causes aggregation. The aggregation facilitates cell fusion to form the diploid zygote (7). a-Agglutinin consists of an anchorage subunit (Agalp) and a binding subunit (Aga2p), and a-agglutinin consists of a single subunit (Agaip) (25). The AGal gene contains a domain with significant sequence similarity to members of the immunoglobulin superfamily (49). Because many members of this superfamily are cell adhesion proteins, this homology suggests that a mechanism of cell adhesion may be conserved in yeasts and multicellular eukaryotes (25,46).Transport of the agglutinins to the cell surface occurs through the secretary pathway (40, 41). All three agglutinin structural genes contain N-terminal hydrophobic sequences that are likely to act as signal sequences that direct the proteins into the secretary pathway (3,19,24,33). The C termini of AGal and AGA1 are also hydrophobic (24, 33) and resemble the addition signals for glycosyl-phosphatidylinositol (GPI) anchors (8,12,14). Such anchors have been shown or inferred to be present on several other yeast proteins (2,5,6,18,26,30,42). A 140-kDa form of a-agglutinin has been demonstrated to be GPI anchored to membranes, and the C-terminal hydrophobic domain is essential for membrane association of this form and cell surface localization of mature a-agglutinin (49).Because mature a-agglutinin is anchored in the cell wall rather than to the plasma membrane (1, 19, 23, 35, (212) 772-5227. have characterized intermediates in intracellular transport and cell wall anchorage of a-agglutinin. MATERUILS AND METHOD...

Lipid rafts function in biosynthetic delivery of proteins to the cell surface in yeast

“…First, we examined the DIG association of ␣-agglutinin, which exhibits a dramatic change in size because of extensive glycosylation in the ER and the Golgi (28). After induction of its synthesis by mixing cells of both mating types, both the ER (80 and 140 kDa) and the plasma membrane forms (Ͼ250 kDa) were found in the DIG fraction (not shown).…”

β-Glucosylated proteins in the cell wall of the black yeast Exophiala (Wangiella) dermatitidis

“…These proteins are all rich in serine/threonine and contain a glycosylphosphatidylinositol (GPI) attachment signal (Lu et al, 1995;Van der Vaart et al, 1995). It is believed that the 1,6-p-glucan side chain is linked to the protein through the GPI-anchor (Kapteyn et al, 1994;Lu et al, 1994;Kapteyn et al, 1996). Lu et al (1995) have shown that 1,6-p-glucan becomes attached to a-agglutinin upon incorporation into the cell wall.…”

Identification of a ligand-binding site in an immunoglobulin fold domain of the Saccharomyces cerevisiae adhesion protein alpha-agglutinin.

“…§ a-Agglutinin (Agalp) is composed of a single polypeptide that contains two functional regions, a cell surface anchorage region and a ligand-binding or adhesion region. The C-terminal half consists of a highly glycosylated, serine/threonine-rich domain followed by a hydrophobic signal for attachment of a glycosyl phosphatidylinositol (GPI) anchor, which mediates anchorage to the cell wall (Wojciechowicz et al, 1993;Lu et al, 1994Lu et al, , 1995. The N-terminal half of a-agglutinin is active in binding to its ligand (Wojciechowicz et al, 1993); this adhesion region contains a domain (called domain III; Figure 1A) that shows similarity to immunoglobulin (Ig) fold domains (Williams and Barclay, 1988).…”