Two homologous genes, DCW1 (YKL046c) and DFG5, are essential for cell growth and encode glycosylphosphatidylinositol (GPI)‐anchored membrane proteins required for cell wall biogenesis in Saccharomyces cerevisiae

Kitagaki, Hiroshi; Wu, Hong; Shimoi, Hitoshi; Itō, Kiyoshi

doi:10.1046/j.1365-2958.2002.03244.x

Cited by 126 publications

(115 citation statements)

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“…Thus, it seems that defects in the PKC pathway confer sensitivity to poacic acid, whereas activation of the PKC pathway confers resistance. A deletion mutant of DFG5 (defective for filamentous growth) also was resistant to poacic acid; DFG5 encodes a GPIanchored protein involved in cell wall biogenesis that also has a genetic interaction with Bck1p (32,33).…”

Section: Chemical Genomics Predict That Poacic Acid Targets the Fungamentioning

confidence: 99%

Plant-derived antifungal agent poacic acid targets β-1,3-glucan

Piotrowski

Okada

et al. 2015

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

133

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A rise in resistance to current antifungals necessitates strategies to identify alternative sources of effective fungicides. We report the discovery of poacic acid, a potent antifungal compound found in lignocellulosic hydrolysates of grasses. Chemical genomics using Saccharomyces cerevisiae showed that loss of cell wall synthesis and maintenance genes conferred increased sensitivity to poacic acid. Morphological analysis revealed that cells treated with poacic acid behaved similarly to cells treated with other cell wall-targeting drugs and mutants with deletions in genes involved in processes related to cell wall biogenesis. Poacic acid causes rapid cell lysis and is synergistic with caspofungin and fluconazole. The cellular target was identified; poacic acid localized to the cell wall and inhibited β-1,3-glucan synthesis in vivo and in vitro, apparently by directly binding β-1,3-glucan. Through its activity on the glucan layer, poacic acid inhibits growth of the fungi Sclerotinia sclerotiorum and Alternaria solani as well as the oomycete Phytophthora sojae. A single application of poacic acid to leaves infected with the broad-range fungal pathogen S. sclerotiorum substantially reduced lesion development. The discovery of poacic acid as a natural antifungal agent targeting β-1,3-glucan highlights the potential side use of products generated in the processing of renewable biomass toward biofuels as a source of valuable bioactive compounds and further clarifies the nature and mechanism of fermentation inhibitors found in lignocellulosic hydrolysates.

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Section: Chemical Genomics Predict That Poacic Acid Targets the Fungamentioning

confidence: 99%

Plant-derived antifungal agent poacic acid targets β-1,3-glucan

Piotrowski

Okada

et al. 2015

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

133

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“…For 16 of the 22 additionally identified proteins, GPI modification has been shown or predicted, such as for the cell wall proteins Cwp2, Ssr1, Fit1, Fit2, and Yar066w and Yhr214w, which both show similarity to the sake yeast specific cell wall protein Awa1 ). Dfg5, which shows similarity to bacterial endomannanases (Kitagaki et al, 2002) and is required for filamentous growth, Dse4 which shows similarity to endo-1,3-β-glucanases, and the aspartic protease Yps6 and phospholipase Plb3 are associated with the plasma membrane. The new algorithm therefore seems to be more effective and selective in recognizing GPI proteins than the earlier one.…”

Section: Developing a Fungal-specific Gpi Algorithmmentioning

confidence: 99%

“…and Candida dubliniensis , the Epa1 adhesin of Candida glabrata (Frieman et al, 2002), Gas/Phr homologues in C. glabrata (Weig et al, 2001), Candida maltosa (Nakazawa et al, 2000) and Aspergillus fumigatus (Mouyna et al, 2000), Fem1 of Fusarium oxysporum (Schoffelmeer et al, 2001), MP1 of Penicillium marneffei (Cao et al, 1998), CAP22 of Glomerella cingulata (Hwang and Kolattukudy, 1995), Ag2 of Coccidioides immitis (Zhu et al, 1996), Psu1 (Omi et al, 1999), Yps1 (Ladds and Davey, 2000) and the Dfg5 homologue SPCC970.02 (Kitagaki et al, 2002) of Schizosaccharomyces pombe, and the Dfg5 homologue NCU03 770.1 of Neurospora crassa (Kitagaki et al, 2002). All these proteins have C-terminal regions that conform to the GPI algorithm, which suggests that the algorithm is applicable for identification of GPI proteins in other fungi.…”

Section: Identification Of Fungal Gpi Proteins 791mentioning

confidence: 99%

Genome‐wide identification of fungal GPI proteins

Groot

Hellingwerf

Klis

2003

Yeast

252

278

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Glycosylphosphatidylinositol-modified (GPI) proteins share structural features that allow their identification using a genomic approach. From the known S. cerevisiae and C. albicans GPI proteins, the following consensus sequence for the GPI attachment site and its downstream region was derived: This consensus sequence, which recognized known GPI proteins from various fungi, was used to screen the genomes of the yeasts S. cerevisiae, C. albicans, Sz. pombe and the filamentous fungus N. crassa for putative GPI proteins. The subsets of proteins so obtained were further screened for the presence of an N-terminal signal sequence for the secretion and absence of internal transmembrane domains. In this way, we identified 66 putative GPI proteins in S. cerevisiae. Some of these are known GPI proteins that were not identified by earlier genomic analyses, indicating that this selection procedure renders a more complete image of the S. cerevisiae GPI proteome. Using the same approach, 104 putative GPI proteins were identified in the human pathogen C. albicans. Among these were the proteins Gas/Phr, Ecm33, Crh and Plb, all members of GPI protein families that are also present in S. cerevisiae. In addition, several proteins and protein families with no significant homology to S. cerevisiae proteins were identified, including the cell wallassociated Als, Csa1/Rbt5, Hwp1/Rbt1 and Hyr1 protein families. In Sz. pombe, which has a low level of (galacto)mannan in the cell wall compared to C. albicans and S. cerevisiae, only 33 GPI candidates were identified and in N. crassa 97. BLAST searches revealed that about half of the putative GPI proteins that were identified in Sz. pombe and N. crassa are homologous to known or putative GPI proteins from other fungi. We conclude that our algorithm is selective and can also be used for GPI protein identification in other fungi.

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“…The structure of ScGas2 is composed of two interacting domains as follows, a (␤␣) 8 catalytic domain, abundantly found in other carbohydrate active enzymes; and a C-terminal cysteine-rich domain of the CBM43 family (11) (Fig. 1).…”

Section: Resultsmentioning

confidence: 99%

“…The crystal structure reveals a (␤␣) 8 catalytic core, tightly interacting with the C-terminal CBM43 glucan binding domain. The active site is located in an unusual tyrosine-rich groove, possessing two glutamic acids as catalytic residues.…”

mentioning

confidence: 99%

Molecular Mechanisms of Yeast Cell Wall Glucan Remodeling

Hurtado‐Guerrero

Schüttelkopf

Mouyna

et al. 2009

Journal of Biological Chemistry

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Yeast cell wall remodeling is controlled by the equilibrium between glycoside hydrolases, glycosyltransferases, and transglycosylases. Family 72 glycoside hydrolases (GH72) are ubiquitous in fungal organisms and are known to possess significant transglycosylase activity, producing elongated ␤(1-3) glucan chains. However, the molecular mechanisms that control the balance between hydrolysis and transglycosylation in these enzymes are not understood. Here we present the first crystal structure of a glucan transglycosylase, Saccharomyces cerevisiae Gas2 (ScGas2), revealing a multidomain fold, with a (␤␣) 8 catalytic core and a separate glucan binding domain with an elongated, conserved glucan binding groove. Structures of ScGas2 complexes with different ␤-glucan substrate/product oligosaccharides provide "snapshots" of substrate binding and hydrolysis/transglycosylation giving the first insights into the mechanisms these enzymes employ to drive ␤(1-3) glucan elongation. Together with mutagenesis and analysis of reaction products, the structures suggest a "base occlusion" mechanism through which these enzymes protect the covalent protein-enzyme intermediate from a water nucleophile, thus controlling the balance between hydrolysis and transglycosylation and driving the elongation of ␤(1-3) glucan chains in the yeast cell wall.The cell wall of fungal organisms is a dynamic structure, providing protection against hostile environments, yet also harboring many hydrolytic and toxic molecules required for the fungus to invade its ecological niche (1). Polysaccharides account for over 90% of the cell wall. The central skeletal component of the cell wall common to the vast majority of fungal species is a branched core of ␤(1,3) glucan, linked to chitin via a ␤(1,4) linkage (1). Interchain ␤(1,6) glucosidic linkages account for 3 and 4% of the total glucan linkages in Saccharomyces cerevisiae and Aspergillus fumigatus, respectively (2-4). This core is embedded in a complex of amorphous proteins and/or polysaccharide whose composition is highly species-dependent. The core ␤(1,3) glucan is subjected to continuous synthetic elaboration, degradation, and remodeling by a large arsenal of enzymes, whose activities must be appropriately balanced to provide the cell wall with adequate elasticity to allow growth, budding, or branching and yet sufficient strength to guard against cell lysis (1).Glucan synthase is a protein complex located at the plasma membrane, synthesizing ␤(1,3) glucan from UDP-glucose (65-90% of the total glucan). In cell wall remodeling, glycoside hydrolases and glycosyltransferases/transglycosylases play a crucial role (1, 5). Pure glycoside hydrolases degrade glycans mainly to regulate the plasticity of the cell wall under different circumstances, such as cell division, cell separation, and sporulation (5), whereas glycoside hydrolases with significant transglycosylase activity are capable of forming new glycosidic bonds between oligosaccharides, generating longer or branched polymers. Previous studies have shown...

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Two homologous genes, DCW1 (YKL046c) and DFG5, are essential for cell growth and encode glycosylphosphatidylinositol (GPI)‐anchored membrane proteins required for cell wall biogenesis in Saccharomyces cerevisiae

Cited by 126 publications

References 55 publications

Plant-derived antifungal agent poacic acid targets β-1,3-glucan

Plant-derived antifungal agent poacic acid targets β-1,3-glucan

Genome‐wide identification of fungal GPI proteins

Molecular Mechanisms of Yeast Cell Wall Glucan Remodeling

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