Isolation and Composition of the Spore Wall of<i>Saccharomyces cerevisiae</i>

Katohda, Shigeyoshi; Konno, K.; Sasaki, Yoko; Suzuki, Kyoko; Sakamoto, Syuu

doi:10.1080/00021369.1984.10866260

Cited by 9 publications

(9 citation statements)

References 6 publications

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“…The two outer layers, which can be clearly distinguished from the inner layers in electron micrographs after staining with OS04 (1), seem to contribute mostly to the spores' resistance to lytic enzymes, proteases, certain organic solvents, and elevated temperature (2). The inner layers are similar in composition to the vegetative cell wall and consist mostly of glucan and mannan (3,4). The sporulationspecific second outer layer, which is situated beneath the very thin and osmiophilic surface layer, consists of glucosamine.…”

mentioning

confidence: 90%

The sporulation-specific enzymes encoded by the DIT1 and DIT2 genes catalyze a two-step reaction leading to a soluble LL-dityrosine-containing precursor of the yeast spore wall.

Briza

Eckerstorfer

Breitenbach

1994

Proc. Natl. Acad. Sci. U.S.A.

115

111

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Dityrosine is a sporulatlon-specific component ofthe yeast ascospore wall that is essential for the resistance of the spores to adverse environmental conditions. Dityrosne in vivo exists in both the LL and DL configurations and is part of an insoluble macromolecule ofunknown strucre. Here we present data indiating that dityrosine of the yeast spore wall is biosynthesized by a different mehanism than dityrosine in other biological systems-e.g., the hard fertilization membrane of the sea urchin egg. We identified two soluble, low molecular weight LL-dityrosine-containing spore wall precursors in extracts of sporulating cells and one precursor containing L-tyrosine. By expression ofthe previously described sporulation-spedfic genes DITI and DIT2 in vegetative cells, it was shown that DIfi catalyzes the reaction leading from L-tyroSine to the tyrosinecontining precursor. DMT2, which is a member of the cytochrome P450 superfamily, is responsible for the dimerization reaction leading to the dityrosine-containing precursors.Epimerization of LL-to DL-dityrosine is one of the latest steps in spore wall formation and takes place after the dityrosinecontaining precursors are Incorporated into the spore wall. On the basis ofthese findings we suggest a biosynthetic pathway for the top layer of the yeast spore wall.The spore wall of Saccharomyces cerevisiae plays a crucial role in protection of the spores from adverse environmental conditions. The two outer layers, which can be clearly distinguished from the inner layers in electron micrographs after staining with OS04 (1), seem to contribute mostly to the spores' resistance to lytic enzymes, proteases, certain organic solvents, and elevated temperature (2). The inner layers are similar in composition to the vegetative cell wall and consist mostly of glucan and mannan (3, 4). The sporulationspecific second outer layer, which is situated beneath the very thin and osmiophilic surface layer, consists of glucosamine. As shown by NMR measurements, the glucosamine molecules are linked by 1-4 glycosidic linkages, and 95% of them are polymerized into chitosan (3). The surface layer, like the second outer layer, has no equivalent in the vegetative cell wall as well. The major component ofthis layer is the dimerized amino acid dityrosine [2,2'-bishydroxy-5,5'-bis(aaminopropionyl)biphenyl], which forms a highly cross-linked scaffold on the spore surface (5). A detailed analysis of the chemical structure of the spore surface is hindered by the insoluble nature of this scaffold, but a macromolecule consisting mostly of dityrosine in its LL and DL configurations can be solubilized and isolated by partial acid hydrolysis from a purified spore wall preparation (6).Despite some progress in the elucidation of components of the spore wall, biosynthesis and assembly of the spore wall remain unclear. The spore wall arises from the prospore wall, presumably a lipid membrane, that forms around the nuclear lobes during meiosis (7). Electron microscopic investigations of sporulating cells ind...

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mentioning

confidence: 90%

The sporulation-specific enzymes encoded by the DIT1 and DIT2 genes catalyze a two-step reaction leading to a soluble LL-dityrosine-containing precursor of the yeast spore wall.

Briza

Eckerstorfer

Breitenbach

1994

Proc. Natl. Acad. Sci. U.S.A.

115

111

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“…This wall offers increased protection to stress conditions as compared to the wall of vegetative cells (4) and consists of four layers (5,6). The two inner layers are formed by closely juxtaposed glucans and mannans and appear as a single layer very similar in morphology to the vegetative cell wall (6,7). The outermost layer consists of an insoluble macromolecule, probably a protein, which contains a high number of cross-linked tyrosine residues (8 -10).…”

mentioning

confidence: 99%

Two Sporulation-specific Chitin Deacetylase-encoding Genes Are Required for the Ascospore Wall Rigidity of Saccharomyces cerevisiae

Christodoulidou

Bouriotis

Thireos

1996

Journal of Biological Chemistry

110

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The formation of the ascospore wall of Saccharomyces cerevisiae requires the coordinate activity of enzymes involved in the biosynthesis of its components such as chitosan, the deacetylated form of chitin. We have cloned the CDA1 and CDA2 genes which together account for the total chitin deacetylase activity of the organism. We have shown that expression of these genes is restricted to a distinct time period during sporulation. The two genes are functionally redundant, each contributing equally to the total chitin deacetylase activity. Diploids disrupted for both genes sporulate as efficiently as wild type cells, and the resulting mutant spores are viable under standard laboratory conditions. However, they fail to emit the natural fluorescence of yeast spores imparted by the dityrosine residues of the outermost ascospore wall layer. Moreover, mutant spores are relatively sensitive to hydrolytic enzymes, ether, and heat shock, a fact that underscores the importance of the CDA genes for the proper formation of the ascospore wall.The cell wall of Saccharomyces cerevisiae, in spite of its apparent rigidity, is a highly dynamic structure. Its general biological functions include mechanical protection, determination of cell shape, and modulation of selective uptake of molecules. Moreover, yeast are able to alter the composition and/or the structure of their cell wall throughout the different stages of their life cycle such as budding, mating, and sporulation. Such plasticity requires the coordinated regulation of the expression of genes involved in the formation of the cell wall. Although cell wall assembly and its regulation have been extensively studied during vegetative growth (1-3), there is limited information on the composition and formation of the more complex ascospore cell wall.Sporulation is the developmental stage that diploid cells enter when starved for nitrogen and a fermentable carbon source. It proceeds with meiosis followed by the encapsulation of the four haploid nuclei within the spore wall. This wall offers increased protection to stress conditions as compared to the wall of vegetative cells (4) and consists of four layers (5, 6). The two inner layers are formed by closely juxtaposed glucans and mannans and appear as a single layer very similar in morphology to the vegetative cell wall (6, 7). The outermost layer consists of an insoluble macromolecule, probably a protein, which contains a high number of cross-linked tyrosine residues (8 -10). Between these layers a chitosan layer has been identified (6). Chitosan, a ␤-(134)-D-glucosamine homopolymer, was initially detected in the cell wall of Zygomycete species (11) and is produced by the deacetylation of nascent chains of chitin, a ␤-(134)-N-acetyl-D-glucosamine homopolymer produced by the action of chitin synthases (12-14). The deacetylation reaction is catalyzed by the enzyme chitin deacetylase (CDA) 1 (15, 16).Chitin deacetylases are involved either in the formation of the cell wall (12,14,17) or in the deacetylation of chitin oligosaccharides ...

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“…The contents of the cell walls in the 20%NaCl medium show a little quantitative variation from in the cell wall of S. cerevisiae. 14 ) Serious increases in the glucosamine contents of alkali-insoluble fractions obtained from 0% NaCl cell walls were observed (Table III). It is likely that the increases of chitin-glucan complex affected bud scar accumulation, especially in the cells grown in the absence of NaCl before a shift (Fig.…”

Section: Discussionmentioning

confidence: 98%

“…The most suggestive of the differences was in the glucosamine content; this sugar is the major component of the bud scar region, which may be mostly composed of a chitin-like substance, as demonstrated in the cell wall of Saccharomyces cerevisiae. 14 ) The glucosamine contents of the cell walls from the cells shifted to the salt-free medium increased to 2.2-5.5-fold of the levels in the cell walls from the cells shifted to the 20% NaCl medium (Table I). The contents of the cell walls in the 20%NaCl medium show a little quantitative variation from in the cell wall of S. cerevisiae.…”

Section: Discussionmentioning

confidence: 99%

Cell Wall Composition ofZygosaccharomyces rouxiiafter a Shift to a High Concentration of NaCl

Katohda¹,

Taniguchi²,

Sumi³

et al. 1991

Agricultural and Biological Chemistry

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We examined the change of the composition of the cell wall polysaccharides prepared from cells of the salt-tolerant yeast Zygosaccharomyces rouxii grown in two media containing 20% NaCl and 0% NaCI. Comparative analysis of their walls showed that the wall obtained from salt-free medium had greater quantities of alkali-insoluble fraction and smaller quantities of mannan than the walls obtained from 20% NaCl medium. The alkali-insoluble fractions from the cell walls obtained from salt -free medium contained a large amount of glucosamine and a smaller amount of linear P-I,3-glucosidic linkage than the fractions from the cell walls obtained from 20% NaCl medium. Structural analyses showed that the mannans from each cell wall contained an IX-I ,6-mannosidic linked backbone to which single mannose and mannobiose units were connected as side chains by IX-I,2-mannosidic linkages. However, when cells were grown in the presence of 20% NaCI, the side chains of the mannans consisting of a mannobiose unit were largely reduced.These results indicated that the structure of alkali-insoluble glucan and mannan were greatly affected by the presence of NaCl in the final medium.It is well known that Zygosaccharomyces rouxii, a salt-tolerant yeast, is important for the fermentation of shoyu and miso.It has been reported that the shape of Z. rauxii cells change from spherical to spheroidal with an increase in NaCI concentration of the medium and the chemical structure of cell walls may play some role in the regulation of the shape of cells under the salt-stress condition. l • 2 ) In spite of many investigations on the structure of the cell wall mannans from Z. rauxii, 2 -5) there has been no report on the comparison of the other carbohydrate composition of the cell walls under the salt-stress conditions. The purpose of t~is paper was to compare the chemical composition and the structure of cell wall polysaccharides obtained from cells of Z. rauxii before and after a shift to the presence of 20% NaCl and to the absence of NaCI. Materials and MethodsOrganisms and culture methods. Zygosaccharomyces rouxii IFO-505 was maintained in a refrigerator on an agar slant containing (w/v) yeast extract (I %), peptone (I %), glucose (2%) (YEPD), and NaCI (18%), First, the cells were transferred to YEPD-agar medium with or without 20% NaCI, and cultivated at 30°C for 3 days, For preculture, the cells were transferred from the agar media to YEPD medium with (20% NaCI medium) or without salt (salt-free medium), in a test tube conaining \0 ml of the medium, and cultivated at 30°C for 48 hr. The cells obtained were washed with the same fresh medium, separately, and transferred, with an inoculum size of 10 5 cells per ml, to 100 ml of salt-free medium and 20% NaCI medium in 500-ml conical flasks. Late exponential phase cultures were washed by centrifugation (620 x g, 5 min), and shifted to salt-free medium or 20% NaCl.medium by adding an equal density of cells. All liquid cultivations were done at 30 c C on a rotary shaker (220 rpm, 6-cm amplitude). Gro...

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Isolation and Composition of the Spore Wall ofSaccharomyces cerevisiae

Cited by 9 publications

References 6 publications

The sporulation-specific enzymes encoded by the DIT1 and DIT2 genes catalyze a two-step reaction leading to a soluble LL-dityrosine-containing precursor of the yeast spore wall.

The sporulation-specific enzymes encoded by the DIT1 and DIT2 genes catalyze a two-step reaction leading to a soluble LL-dityrosine-containing precursor of the yeast spore wall.

Two Sporulation-specific Chitin Deacetylase-encoding Genes Are Required for the Ascospore Wall Rigidity of Saccharomyces cerevisiae

Cell Wall Composition ofZygosaccharomyces rouxiiafter a Shift to a High Concentration of NaCl

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