Altered extent of cross-linking of beta1,6-glucosylated mannoproteins to chitin in Saccharomyces cerevisiae mutants with reduced cell wall beta1,3-glucan content

Abstract

The yeast cell wall contains ␤1,3-glucanase-extractable and ␤1,3-glucanase-resistant mannoproteins. The ␤1,3-glucanase-extractable proteins are retained in the cell wall by attachment to a ␤1,6-glucan moiety, which in its turn is linked to ␤1,3-glucan (J. C. Kapteyn, R. C. Montijn, E. Vink, J. De La Cruz, A. Llobell, J. E. Douwes, H. Shimoi, P. N. Lipke, and F. M. Klis, Glycobiology 6:337-345, 1996). The ␤1,3-glucanase-resistant protein fraction could be largely released by exochitinase treatment and contained the same set of ␤1,6-glucosylated proteins, including Cwp1p, as the ␤1,3-glucanase-extractable fraction. Chitin was linked to the proteins in the ␤1,3-glucanase-resistant fraction through a ␤1,6-glucan moiety. In wild-type cell walls, the ␤1,3-glucanase-resistant protein fraction represented only 1 to 2% of the covalently linked cell wall proteins, whereas in cell walls of fks1 and gas1 deletion strains, which contain much less ␤1,3-glucan but more chitin, ␤1,3-glucanase-resistant proteins represented about 40% of the total. We propose that the increased crosslinking of cell wall proteins via ␤1,6-glucan to chitin represents a cell wall repair mechanism in yeast, which is activated in response to cell wall weakening.The cell wall is crucial for the integrity of Saccharomyces cerevisiae. Its rigid structure maintains the shape of the cell and offers protection against harmful environmental conditions (6, 19). The wall is mainly composed of ␤-glucans and mannoproteins, in addition to smaller amounts of chitin and lipids (6). The glucans, which are interwoven with the chitin fibrils, form the inner skeletal layer of the cell wall, whereas the outer layer consists of mannoproteins. The majority of the cell wall mannoproteins are anchored into the wall through covalent linkages to heteropolymers of ␤1,6-and ␤1,3-glucan (18, 26, 41, 43) (Fig. 1). The ␤1,6-glucosyl moiety of these polymers is phosphodiester-linked to protein as shown by its sensitivity to treatment with ice-cold aqueous hydrofluoric acid (HF) and phosphodiesterases (18). This observation, together with data from other studies, pointed to a glycosylphosphatidylinositol (GPI)-derived structure as the attachment site for ␤1, 6-glucan (3, 23, 40, 42, 45). The ␤1,3/␤1,6-glucan heteropolymer was proposed to be identical to the alkali-soluble ␤1,3/␤1,6-glucan studied by Fleet and Manners (7,8). In S. cerevisiae, this alkali-soluble glucan becomes alkali insoluble through a linkage with chitin (11, 24) via a ␤1,4 bond from the terminal reducing residue of chitin to the nonreducing end of ␤1,3-glucan (20). Furthermore, by digesting cell walls with ␤1,3-glucanase, followed by incubation with exochitinase, a heterogeneous high-molecular-weight complex was isolated (21). Structural studies indicated that in this complex the terminal reducing residue of chitin is linked to ␤1,6-glucan (21). The nonreducing end of the ␤1,6-glucan polymer is bound to the GPI-derived glycan part of a cell wall protein, whereas its reducing terminus is linked to ␤1,3-glucan (...

Scw10p, a cell-wall glucanase/transglucosidase important for cell-wall stability in Saccharomyces cerevisiae

“…An increased level of chitin is one of the typical compensatory mechanisms of cell-wall mutants (Popolo et al, 1997;Garcia-Rodriguez et al, 2000). Chitin is glycosidically linked to non-reducing branches of both 1,3-b-glucan and 1,6-b-glucan (Kollar et al, 1995(Kollar et al, , 1997. It is essential to the insolubility of the wall material, and chitin incorporation results in transfer of the wall material from the alkali-soluble to the alkali-insoluble fraction (Hartland et al, 1994).…”

“…The yeast cell wall is composed of three major components: glucan, mannoproteins and chitin (Kollar et al, 1997; reviewed by Klis et al, 2002). While a 1,3-b-glucan-chitin complex constitutes the inner rigid structure, mannoproteins form the outer layer of the cell wall, and 1,6-b-glucan interconnects all cell-wall components.…”

Dosage rescue by UBC4 restores cell wall integrity in Saccharomyces cerevisiae lacking the myosin type II gene MYO1

“…It has been previously demonstrated by others that cell wall stress can trigger an increase in cross-linking between cell wall proteins via β-1,6-glucan to chitin (Kapteyn et al, 1997) and that a defective incorporation of β-1,3-glucan can lead to an increase in chitin and mannan content in the cell wall (de Nobel et al, 2000;Kapteyn et al, 2001;Yin et al, 2005). These results highlight the importance of glucans, together with chitin and mannoproteins, in preserving cell wall integrity.…”

“…It is expected that myo1 cells will activate a cell wall integrity signalling mechanism that is induced by abnormal cell wall assembly (Cruz et al, 2000;Kapteyn et al, 1997;Osmond et al, 1999;Popolo et al, 1997Popolo et al, , 2001Ram et al, 1998;Valdivieso et al, 2000). The Pkc1p-mediated cell wall integrity signalling pathway (CWIP) is principally responsible for orchestrating changes in the cell wall periodically throughout the cell cycle, as well as in response to various forms of cell wall stress (Heinisch et al, 1999;Levin, 2005;Martin et al, 2005;Vilella et al, 2005).…”

Secretion, interaction and assembly of two O-glycosylated cell wall antigens from Candida albicans

“…This result was expected, as in C. albicans 90 % of the GPI proteins are attached via 1,6-β-glucan to 1,3-β-glucan and the remaining ones are attached through 1,6-β-glucan to chitin (Kapteyn et al, 2000). Similarly in S. cerevisiae, 1-2 % of cell wall proteins are attached directly to chitin through a 1,6-β-glucan moiety , and Cwp1p is found as a 1,6-β-glucanase-extractable and non-extractable, chitin-linked fraction (Van der Vaart et al, 1996 ;Kapteyn et al, 1997). Although the mobility of the highmolecular-mass band was different, the behaviour of the three antigens towards additional digestions or β-elimination was similar to that found in the material released by SDS.…”