Amino Acid Sequence Requirement for Efficient Incorporation of Glycosylphosphatidylinositol-associated Proteins into the Cell Wall of Saccharomyces cerevisiae

Abstract

During cell wall biogenesis in Saccharomyces cerevisiae, some glycosylphosphatidylinositol (GPI)-attached proteins are detached from GPI moieties and bound to β-1,6-glucan of the cell wall. The amino acid sequence requirement for the incorporation of GPI-attached proteins into the cell wall was studied by using reporter fusion proteins. Only the short ω-minus region composed of five amino acids, which is located upstream of the ω site for GPI attachment, determined the cellular localization of the GPI-associated proteins. Within the ω-minus region, amino acid residues at the ω-4 or -5 and ω-2 sites were important for the cell wall incorporation. Yap3p, a well characterized GPI-anchored plasma membrane aspartic protease, was localized in the cell wall when the ω-minus region was mutated to sequences containing Val or Ile at the ω-4 or -5 site and Val or Tyr at the ω-2 site. During cell wall biogenesis in Saccharomyces cerevisiae, some glycosylphosphatidylinositol (GPI)-attached proteins are detached from GPI moieties and bound to β-1,6-glucan of the cell wall. The amino acid sequence requirement for the incorporation of GPI-attached proteins into the cell wall was studied by using reporter fusion proteins. Only the short ω-minus region composed of five amino acids, which is located upstream of the ω site for GPI attachment, determined the cellular localization of the GPI-associated proteins. Within the ω-minus region, amino acid residues at the ω-4 or -5 and ω-2 sites were important for the cell wall incorporation. Yap3p, a well characterized GPI-anchored plasma membrane aspartic protease, was localized in the cell wall when the ω-minus region was mutated to sequences containing Val or Ile at the ω-4 or -5 site and Val or Tyr at the ω-2 site. glycosylphosphatidylinositol hemagglutinin polymerase chain reaction phosphatidylinositol-specific phospholipase C endoglycosidase H. Mannoproteins, one of the components of the yeast cell wall, can be divided into three groups: SDS-extractable mannoproteins, reducing agent-extractable mannoproteins, and glucanase-extractable mannoproteins (1Klis F.M. Yeast. 1994; 10: 851-869Crossref PubMed Scopus (487) Google Scholar, 2Cid V.J. Duran A. del Rey F. Snyder M.P. Nombela C. Sanchez M. Microbiol. Rev. 1995; 59: 345-386Crossref PubMed Google Scholar). The glucanase-extractable mannoproteins are covalently bound to β-1,6-glucan of the cell wall and are released only by glucanase treatment (3De Nobel H. Lipke P.N. Trends Cell Biol. 1994; 4: 42-45Abstract Full Text PDF PubMed Scopus (91) Google Scholar, 4Montijn R.C. van Rinsum J. van Schagen F.A. Klis F.M. J. Biol. Chem. 1994; 269: 19338-19342Abstract Full Text PDF PubMed Google Scholar, 5Lu C.-F. Montijn R.C. Brown J.L. Klis F. Kurjan J. Bussey H. Lipke P.N. J. Cell Biol. 1995; 128: 333-340Crossref PubMed Scopus (188) Google Scholar, 6Kapteyn J.C. Montijn R.C. Vink E. de la Cruz J. Llobell A. Douwes J.E. Shimoi H. Lipke P.N. Klis F.M. Glycobiology. 1996; 6: 337-345Crossref PubMed Scopus (223) Google Scholar). Many genes encoding glucanase-extractable cell wall mannoproteins have been isolated in Saccharomyces cerevisiae, and so far all of them have been identified as GPI-dependent1cell wall proteins (7Lipke P.N. Wojciechowicz D. Kurjan J. Mol. Cell. Biol. 1989; 9: 3155-3165Crossref PubMed Scopus (148) Google Scholar, 8Roy A. Lu C.-F. Marykwas D.L. Lipke P.N. Kurjan J. Mol. Cell. Biol. 1991; 11: 4196-4206Crossref PubMed Scopus (128) Google Scholar, 9Kondo K. Inouye M. J. Biol. Chem. 1991; 266: 17537-17544Abstract Full Text PDF PubMed Google Scholar, 10Shimoi H. Iimura Y. Obata T. J. Biochem. (Tokyo). 1995; 118: 302-311Crossref PubMed Scopus (57) Google Scholar, 11Van der Vaart J.M. Caro L.H.P. Chapman J.W. Klis F.M. Verrips C.T. J. Bacteriol. 1995; 177: 3104-3110Crossref PubMed Google Scholar, 12Teunissen A.W.R.H. Steensma H.Y. Yeast. 1995; 11: 1001-1013Crossref PubMed Scopus (220) Google Scholar, 13Kowalski L.R.Z. Kondo K. Inouye M. Mol. Microbiol. 1995; 15: 341-353Crossref PubMed Scopus (81) Google Scholar, 14Moukadiri I. Armero J. Abad A. Sentandreu R. Zueco J. J. Bacteriol. 1997; 179: 2154-2162Crossref PubMed Google Scholar, 15Van der Vaart J.M. Biesebeke R. Chapman J.W. Toshka H.Y. Klis F.M. Verrips C.T. Appl. Environ. Microbiol. 1997; 63: 615-620Crossref PubMed Google Scholar). GPI-associated proteins have been isolated from various organisms from yeasts and protozoa to mammals (16Field M.C. Menon A.K. Schlesinger M.J. Lipid Modification of Proteins. CRC Press, Inc., Boca Raton, FL1993: 83-134Google Scholar, 17Udenfriend S. Kodukula K. Annu. Rev. Biochem. 1995; 64: 563-591Crossref PubMed Scopus (436) Google Scholar, 18Nosjean O. Briolay A. Roux B. Biochim. Biophys. Acta. 1997; 1331: 153-186Crossref PubMed Scopus (177) Google Scholar). These proteins are structurally related in that they all contain a signal sequence for secretion in the N terminus and a GPI signal for an attachment to a GPI in the C terminus. The GPI-signal region is composed of three domains: a GPI attachment region comprising ω, ω+1, and ω+2 sites; a spacer of 5–10 amino acids; and a hydrophobic stretch of 10–15 amino acids. A protein containing the GPI-signal is cleaved at the ω site, and the resulting carboxyl terminus of the protein becomes covalently bound to a GPI moiety. This reaction occurs in the luminal face of the endoplasmic reticulum and, in yeast, requires GAA1 (19Hamburger D. Egerton M. Riezman H. J. Cell Biol. 1995; 129: 629-639Crossref PubMed Scopus (149) Google Scholar) and GPI8 (20Benghezal M. Benachour A. Rusconi S. Aebi M. Conzelmann A. EMBO J. 1996; 15: 6575-6583Crossref PubMed Scopus (153) Google Scholar) gene products. The GPI-attached proteins are then transported to the cell surface, where they are exposed on the extracytoplasmic face of the plasma membrane. During the transportation from the endoplasmic reticulum to the plasma membrane, the proteins are mannosylated and become GPI-anchored mannoproteins. In protozoa and mammals, the GPI-anchored mannoproteins remain on the plasma membrane and take biological functions related to cell-cell and cell-environment interactions (18Nosjean O. Briolay A. Roux B. Biochim. Biophys. Acta. 1997; 1331: 153-186Crossref PubMed Scopus (177) Google Scholar, 21McConville M. Ferguson M.A.J. Biochem. J. 1993; 294: 305-324Crossref PubMed Scopus (805) Google Scholar, 22Kinoshita T. Inoue N. Takeda J. Adv. Immunol. 1995; 60: 57-103Crossref PubMed Google Scholar). In addition to the above functions, some of the GPI-anchored mannoproteins in the yeast are further processed and are incorporated into the cell wall. The incorporation of GPI-associated proteins into the cell wall is thought to occur in two steps: detachment of a GPI moiety from the protein and linking of the GPI-detached protein to β-1,6-glucan of the cell wall (3De Nobel H. Lipke P.N. Trends Cell Biol. 1994; 4: 42-45Abstract Full Text PDF PubMed Scopus (91) Google Scholar). Our knowledge of this process is very rudimentary. The addition of 30–40 amino acids from the C terminus of GPI-dependent cell wall proteins to a reporter protein directs incorporation of the chimeric proteins into the cell wall (23Van Barkel M.A.A. Caro L.H.P. Montijn R.C. Klis F.M. FEBS Lett. 1994; 349: 135-138Crossref PubMed Scopus (46) Google Scholar, 24Hamada K. Fukuchi S. Arisawa M. Baba M. Kitada K. Mol. Gen. Genet. 1998; 258: 53-59Crossref PubMed Scopus (99) Google Scholar, 25Bourdineaud J.-P. Van der Vaart J.M. Donzeau M. de Sampaio G. Verrips T. Lauquin G.J.-M. Mol. Microbiol. 1998; 27: 85-98Crossref PubMed Scopus (29) Google Scholar). This suggests that the short C-terminal sequences contain all the information necessary for the incorporation of proteins into the cell wall. Therefore, it is possible that there is a gene product that would recognize a sequence or structural motif in the C-terminal region of GPI-associated proteins and direct their final cellular localization. A sequence alignment from our previous study on GPI-dependent cell wall proteins (24Hamada K. Fukuchi S. Arisawa M. Baba M. Kitada K. Mol. Gen. Genet. 1998; 258: 53-59Crossref PubMed Scopus (99) Google Scholar) suggested that there is some sequence the ω-minus to upstream of the ω site, of GPI-dependent cell wall proteins identified in the In this in the amino acid sequences of the ω-minus and determined the sequence requirement for the incorporation of GPI-attached proteins into the cell wall. were in S. Google Scholar). the F. in Scholar) was for T. J. A Scholar) was for the and of was as by T. J. A Scholar). (24Hamada K. Fukuchi S. Arisawa M. Baba M. Kitada K. Mol. Gen. Genet. 1998; 258: 53-59Crossref PubMed Scopus (99) Google Scholar) containing a reporter gene comprising the signal sequence of yeast from and a hemagglutinin was as a The reporter gene is the A the C-terminal amino acids of a GPI-attached protein and of region was by polymerase chain reaction containing a site. The resulting was into the and sites of and then to the yeast of amino acid residues and of chimeric fusion genes were by K. E. K. Arisawa M. Genet. 1997; PubMed Scopus Google Scholar). A sequence was into the of the amino acid sequence M. Yeast. 6: PubMed Scopus Google Scholar). A containing the of the region and the amino acids of was by and into and sites of the sequence M. G. B. EMBO J. 1993; PubMed Scopus Google Scholar) the and sites of the site of A. PubMed Scopus Google Scholar). containing the C-terminal amino acids of and the of region was and into the and on at in the were by and proteins were isolated from the and to as (24Hamada K. Fukuchi S. Arisawa M. Baba M. Kitada K. Mol. Gen. Genet. 1998; 258: 53-59Crossref PubMed Scopus (99) Google Scholar, C. A. Riezman H. J. Biol. Chem. 1993; Full Text PDF PubMed Google Scholar, D. Lu C.-F. Kurjan J. Lipke P.N. Mol. Cell. Biol. 1993; PubMed Google Scholar). were in containing of cell by a a was to the to a final of and the was on for The was a and the was to a at for the was a and into an and a The was and the was proteins in the were as A. Riezman H. C. C. EMBO J. PubMed Scopus Google Scholar). proteins in were of at for and then into a and an in were acid and by and K. A. Mol. Cell. Biol. 1993; PubMed Scopus Google Scholar). was for of the fusion proteins on Cell were isolated from yeast as (24Hamada K. Fukuchi S. Arisawa M. Baba M. Kitada K. Mol. Gen. Genet. 1998; 258: 53-59Crossref PubMed Scopus (99) Google Scholar, C.-F. Kurjan J. Lipke P.N. Mol. Cell. Biol. 1994; PubMed Google G. E. S. Mol. Cell. Biol. 1996; PubMed Google Scholar). were in containing and the cell was a The resulting was a by for at and five The cell were at a final of for at as (24Hamada K. Fukuchi S. Arisawa M. Baba M. Kitada K. Mol. Gen. Genet. 1998; 258: 53-59Crossref PubMed Scopus (99) Google Scholar, M.P. S. Van H. Klis F.M. Yeast. 1993; 9: PubMed Scopus Google Scholar). The was a and the resulting was by and of the proteins on was using the were as C.-F. Montijn R.C. Brown J.L. Klis F. Kurjan J. Bussey H. Lipke P.N. J. Cell Biol. 1995; 128: 333-340Crossref PubMed Scopus (188) Google Scholar). from the cell wall the treatment were at for in containing and was to the to a final of The reaction was at and then further at for a further the addition of the of H. ω sites for GPI attachment are on the by C. A. Riezman H. J. Biol. Chem. 1993; Full Text PDF PubMed Google Scholar) and by and Kodukula S. Kodukula K. Annu. Rev. Biochem. 1995; 64: 563-591Crossref PubMed Scopus (436) Google Scholar). A sequence of GPI-associated proteins suggested that in S. two amino acid and are as ω site residues (24Hamada K. Fukuchi S. Arisawa M. Baba M. Kitada K. Mol. Gen. Genet. 1998; 258: 53-59Crossref PubMed Scopus (99) Google Scholar, L.H.P. H. Van H. Klis F.M. Yeast. 1997; PubMed Scopus Google Scholar). A of the ω site in a well characterized yeast GPI-anchored plasma membrane protein M. E. E. J. Biol. Chem. 1991; 266: Full Text PDF PubMed Google Scholar, C. Conzelmann A. Riezman H. Mol. Cell. Biol. 1991; 11: PubMed Scopus (177) Google of the ω site of the ω site from an ω site or the of GPI attachment, of amino acid residues and in an of the GPI attachment C. A. Riezman H. J. Biol. Chem. 1993; Full Text PDF PubMed Google Scholar). on this of the ω site determined the ω sites of two GPI-dependent cell wall (24Hamada K. Fukuchi S. Arisawa M. Baba M. Kitada K. Mol. Gen. Genet. 1998; 258: 53-59Crossref PubMed Scopus (99) Google Scholar) and der Vaart J.M. Biesebeke R. Chapman J.W. Toshka H.Y. Klis F.M. Verrips C.T. Appl. Environ. Microbiol. 1997; 63: 615-620Crossref PubMed Google and two GPI-anchored plasma membrane (24Hamada K. Fukuchi S. Arisawa M. Baba M. Kitada K. Mol. Gen. Genet. 1998; 258: 53-59Crossref PubMed Scopus (99) Google Scholar) and J. M. Y. J. Biol. Chem. 1995; Full Text Full Text PDF PubMed Scopus Google Scholar, H. S. A. 1995; PubMed Scopus Google Scholar, M. 1995; PubMed Scopus Google Scholar). The C-terminal in mutated amino residues at the ω site were to the reporter and the resulting fusion proteins were in yeast GPI of the fusion proteins was by using of membrane proteins and In the C-terminal sequences of and containing the at the ω site, the fusion proteins were in the and were released into the by the GPI-dependent in fusion proteins of or at the ω site were in the The of proteins were that of the and they were the fusion proteins of or at the ω site were in the as as the The of proteins were and of them were of GPI on GPI the of the ω site for GPI to that in C. A. Riezman H. J. Biol. Chem. 1993; Full Text PDF PubMed Google Scholar). were amino acid at the ω sites for the C-terminal sequences of and it be that for all proteins the N residues as the ω sites for GPI Our previous study (24Hamada K. Fukuchi S. Arisawa M. Baba M. Kitada K. Mol. Gen. Genet. 1998; 258: 53-59Crossref PubMed Scopus (99) Google Scholar) that the addition of the C-terminal amino acids of or to a reporter protein directs the resulting fusion proteins into the cell wall and that the addition of the C-terminal amino acids of or directs their fusion proteins to the plasma membrane. This that the C-terminal amino acid sequences the final cellular localization of GPI-associated proteins. These C-terminal amino acid sequences can be divided into two the and ω-minus the ω site as a The region the residues of ω, ω+1, and ω+2 that as the sequence for GPI a spacer sequence of 5–10 amino acids; and a hydrophobic stretch of 10–15 amino acids. The ω-minus region is to the ω site. which of the two is for of cellular chimeric genes the two were and in yeast and the cellular localization of their chimeric fusion proteins was The chimeric proteins of the ω-minus region from or and the region from or were in the cell wall as as of the and were released only by of the cell wall the the chimeric proteins of the ω-minus region from or and the region from or were in the cell wall were as as in the of and chimeric proteins were determined to be GPI-associated they were released from membrane by the treatment it be that only the ω-minus region is for of the final cellular localization of GPI-associated proteins. ω-minus sequences of amino acids were in A and Therefore, a signal the final cellular localization be located short ω-minus amino acid sequence for the cell wall further the ω-minus of and the ω-minus of the GPI-dependent cell wall proteins are (24Hamada K. Fukuchi S. Arisawa M. Baba M. Kitada K. Mol. Gen. Genet. 1998; 258: 53-59Crossref PubMed Scopus (99) Google Scholar, L.H.P. H. Van H. Klis F.M. Yeast. 1997; PubMed Scopus Google amino acid in the sequence and the of the on the incorporation of and fusion proteins into the cell wall fusion proteins were in the C-terminal sequence of only the sequence to the to sites the for the cell wall incorporation This was to a of the of the fusion proteins released by the treatment of the membrane was that of a fusion protein the sequence Within the sequence a of Val to at the site the incorporation into the cell wall or amino acid the ω-2 site the cell wall incorporation a at the and the ω-2 sites the in the cell wall incorporation for the C-terminal sequence of amino acid only in the sequence to the to sites the for the cell wall incorporation. of the or ω-2 site in the sequence a of the cell wall incorporation and of and ω-2 site residues a of cell wall incorporation it be that the signal for the cell wall incorporation would be located in the sequence of the to sites and that the sequence the and ω-2 sites are the A sequence alignment of the ω-minus in the GPI-dependent cell wall proteins (24Hamada K. Fukuchi S. Arisawa M. Baba M. Kitada K. Mol. Gen. Genet. 1998; 258: 53-59Crossref PubMed Scopus (99) Google Scholar) that residues Ile and Val at the site and residues Tyr and are at the ω-2 site. the requirement of amino acid residues for the cell wall incorporation by using a a stretch of by the ω site and the region of a well characterized GPI-anchored plasma membrane protein J. M. Y. J. Biol. Chem. 1995; Full Text Full Text PDF PubMed Scopus Google Scholar, H. S. A. 1995; PubMed Scopus Google Scholar, M. 1995; PubMed Scopus Google to a this fusion protein was localized in as a GPI-anchored protein a of the fusion protein was in the cell wall as a glucanase-extractable protein The addition of the ω-minus sequence containing the to region from and all GPI-dependent cell wall the of fusion proteins incorporated into the cell wall of incorporation was various the five Val at the and sites and their on the cell wall incorporation of the fusion proteins Only a at the ω-2 site the incorporation of fusion proteins. A was when was to Ile at the ω-2 site was or The of the cell wall incorporation was by the Val or Ile and by the Tyr or was by the in addition to the of to Tyr at the ω-2 site, a of to Ile at the sites was The at the or ω-4 site further the of the cell wall incorporation. In only of the fusion protein was in the cell wall, in and and of the fusion proteins were in the cell wall. The of the cell wall incorporation was by the of to Ile at the or ω-4 site a of to Tyr at the ω-2 site. of the cell wall incorporation was by the of Val at the or ω-4 site and of at the ω-2 site A of the cell wall incorporation was by a of to Val at the ω-4 or site in addition to the of to Val at the ω-2 site all of it be that of Ile or Val at the ω-4 or -5 site and or Val at the ω-2 site to the incorporation of fusion proteins into the cell wall. S. Yap3p, an aspartic protease, is a GPI-anchored plasma membrane protein J. M. Y. J. Biol. Chem. 1995; Full Text Full Text PDF PubMed Scopus Google Scholar, H. S. A. 1995; PubMed Scopus Google Scholar, M. 1995; PubMed Scopus Google Scholar). this the of amino acid residues at the ω-4 or -5 and ω-2 sites directs the protein into the cell wall. the C-terminal sequence of the ω-minus region was The fusion protein this C-terminal sequence was in the cell wall as a glucanase-extractable protein of the ω-minus region from the to or in the fusion protein incorporated into the cell wall These sequences to the to of GPI-dependent cell wall proteins. a from to to the incorporation of the fusion protein in the cell wall, was very the was The protein from membrane was as a was in a of and was released from membrane by treatment that is a GPI-anchored membrane proteins were as GPI-attached membrane proteins incorporation of the and proteins into the cell wall was proteins from the cell wall by glucanase were further the treatment they the of in the Yap3p, of three proteins were released from the cell wall by the glucanase treatment contain mutated sequences of or in the to ω-2 region A containing in the to region was in the cell wall, and was that of the wall incorporation of mutated proteins. of amino acid the sequence of the in the to region of were by of of the protein by and in a of was and and then into a and an in were and and then The proteins were by using cell of of the and proteins. The as in for the proteins. Only of are and and for treatment in and treatment as in B. of cell of the and proteins. The proteins released from the cell wall were and then by using have studied the sequence requirement for incorporation of GPI-attached proteins into the cell wall and the that only the short ω-minus region composed of five amino acids the ω-minus which is located upstream of the ω site for GPI attachment, the cellular localization of the GPI-associated proteins and that the ω-minus sequence amino acid residues at the ω-4 or -5 and ω-2 sites are important for the the and ω-2 Yap3p, a well characterized GPI-anchored plasma membrane protein J. M. Y. J. Biol. Chem. 1995; Full Text Full Text PDF PubMed Scopus Google Scholar, H. S. A. 1995; PubMed Scopus Google Scholar, M. 1995; PubMed Scopus Google localized in the cell wall when the ω-minus sequence of was mutated to sequences containing Val or Ile at the ω-4 or -5 site and Val or Tyr at the ω-2 site. In the previous study (24Hamada K. Fukuchi S. Arisawa M. Baba M. Kitada K. Mol. Gen. Genet. 1998; 258: 53-59Crossref PubMed Scopus (99) Google some sequence in the ω-minus of GPI-dependent cell wall proteins where Val or Ile the site and Tyr is at the ω-2 site. This study their for the cell wall incorporation. a of or at the ω-2 site some to direct fusion proteins into the cell wall, the of the or at the ω-2 site Val or Ile at the site the fusion protein into the cell wall. In of GPI-dependent cell wall proteins and three proteins of Ile and Val and and Val and at the and ω-2 sites (24Hamada K. Fukuchi S. Arisawa M. Baba M. Kitada K. Mol. Gen. Genet. 1998; 258: 53-59Crossref PubMed Scopus (99) Google Scholar). The fusion protein containing the sequence of as the ω-minus sequence of was localized in the cell wall, the fusion protein containing the mutated sequence of was localized in the cell wall The sequence of in and in is a of to to in the localization of the fusion protein is possible that the and residues at the ω-2 and sites for the cell wall incorporation. residues this have been identified in the ω-minus region in some of GPI-anchored plasma membrane proteins (24Hamada K. Fukuchi S. Arisawa M. Baba M. Kitada K. Mol. Gen. Genet. 1998; 258: 53-59Crossref PubMed Scopus (99) Google Scholar, L.H.P. H. Van H. Klis F.M. Yeast. 1997; PubMed Scopus Google Scholar, Lipke P.N. Klis F.M. J. Bacteriol. 1997; 179: PubMed Google Scholar). These residues be a motif as a A protein containing the motif be processed for incorporation into the cell wall and then remain on the plasma membrane a this is the the of the and ω-2 for the cell wall incorporation would be as of the signal by the this would be to the and by the and ω-2 on the cell wall incorporation. the incorporated the fusion proteins into the cell wall that of the Therefore, the be for the cell wall it would be for incorporation of GPI-attached proteins into the cell wall. The incorporation of GPI-associated proteins into the cell wall been thought to occur in two steps: detachment of a from a GPI-attached protein and of the GPI-detached protein to β-1,6-glucan of the cell wall (3De Nobel H. Lipke P.N. Trends Cell Biol. 1994; 4: 42-45Abstract Full Text PDF PubMed Scopus (91) Google Scholar). This β-1,6-glucan is covalently as well as and a cell wall components J.C. Montijn R.C. Vink E. de la Cruz J. Llobell A. Douwes J.E. Shimoi H. Lipke P.N. Klis F.M. Glycobiology. 1996; 6: 337-345Crossref PubMed Scopus (223) Google Scholar, R. E. G. J. J.C. Klis F.M. E. J. Biol. Chem. 1997; Full Text Full Text PDF PubMed Scopus Google Scholar, J.C. R. Montijn R.C. Van H. Llobell A. E. Klis F.M. J. Bacteriol. 1997; 179: PubMed Google Scholar). β-1,6-glucan is to be J.L. B. Bussey H. 1993; PubMed Google it is possible that the of a GPI-detached protein to β-1,6-glucan occurs in or the β-1,6-glucan the study using an β-1,6-glucan this The proteins from the cell wall, β-1,6-glucan R.C. van Rinsum J. van Schagen F.A. Klis F.M. J. Biol. Chem. 1994; 269: 19338-19342Abstract Full Text PDF PubMed Google Scholar, 5Lu C.-F. Montijn R.C. Brown J.L. Klis F. Kurjan J. Bussey H. Lipke P.N. J. Cell Biol. 1995; 128: 333-340Crossref PubMed Scopus (188) Google Scholar, 6Kapteyn J.C. Montijn R.C. Vink E. de la Cruz J. Llobell A. Douwes J.E. Shimoi H. Lipke P.N. Klis F.M. Glycobiology. 1996; 6: 337-345Crossref PubMed Scopus (223) Google Scholar, Barkel M.A.A. Caro L.H.P. Montijn R.C. Klis F.M. FEBS Lett. 1994; 349: 135-138Crossref PubMed Scopus (46) Google Scholar, H. Shimoi H. K. J. Biochem. 1997; Scopus Google it proteins C.-F. Montijn R.C. Brown J.L. Klis F. Kurjan J. Bussey H. Lipke P.N. J. Cell Biol. 1995; 128: 333-340Crossref PubMed Scopus (188) Google Scholar). Therefore, the of the GPI moiety and the linking to β-1,6-glucan would occur in the This suggests that GPI-dependent cell wall proteins are transported to the plasma membrane as a of GPI-attached proteins that are by the GPI moiety. the GPI-attached of the cell wall proteins was in the of and The cell wall incorporation reaction would an and at the of the incorporation detachment of a GPI would of the to the GPI-attached The and ω-2 be related to this in the ω-minus sequence that the cell wall incorporation the of the fusion proteins into the This suggests that the cell wall incorporation in is to of the GPI detachment by the and the GPI-attached A structural of GPI-dependent cell wall proteins that β-1,6-glucan is to the protein a GPI of that the GPI contain and that β-1,6-glucan is bound to one of the residues of the GPI moiety J.C. Montijn R.C. Vink E. de la Cruz J. Llobell A. Douwes J.E. Shimoi H. Lipke P.N. Klis F.M. Glycobiology. 1996; 6: 337-345Crossref PubMed Scopus (223) Google Scholar, C.-F. Kurjan J. Lipke P.N. Mol. Cell. Biol. 1994; PubMed Google Scholar, der Vaart J.M. Biesebeke R. Chapman J.W. Klis F.M. Verrips C.T. Lett. 1996; Scopus Google Scholar). this structural it is possible that the GPI moiety be by residues or the and and then the resulting is bound to as in the protein G. E. S. Mol. Cell. Biol. 1996; PubMed Google the GPI moiety be cleaved by an In this of and residues be the of it to the is to and of a for the incorporation of GPI-attached proteins into the cell wall is for the for the and our are on the of the for of the and for