The SC15 protein of <i>Schizophyllum commune</i> mediates formation of aerial hyphae and attachment in the absence of the SC3 hydrophobin

Lugones, Luis G.; Jong, Jan F. de; Vries, O.; Jalving, Ruud; Dijksterhuis, Jan; Wösten, Han A. B.

doi:10.1111/j.1365-2958.2004.04187.x

Cited by 45 publications

(39 citation statements)

References 32 publications

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“…In the homobasidiomycetes, RNA-based gene silencing is a powerful alternative to gene inactivation by homologous recombination, which occurs at low frequency in this group of fungi. For instance, gene inactivation of SC3, SC4, and SC15 in S. commune occurs at a frequency of 1 to 5% (9,14,15). More importantly, functional analysis of genes that are active in the heterokaryotic phase, e.g., in mushroom formation and sporulation, requires inactivation of the gene in both nuclei of the dikaryon.…”

Section: Discussionmentioning

confidence: 99%

“…SC15 and SC3 secretion was monitored by immunodetection after incubation of the colonies on PVDF membrane for 2 and 17 h, respectively. The SC3 and SC15 antisera (9,17) were diluted 10,000-and 5,000-fold, respectively. Goat anti-rabbit-conjugated alkaline phosphatase was used as a second antibody with BCIP (5-bromo-4-chloro-3-indolylphosphate) and nitroblue tetrazolium as substrates (7).…”

Section: Methodsmentioning

confidence: 99%

“…To this end, a hairpin construct of SC15 was made. This gene encodes an abundantly secreted 17-kDa protein of S. commune, which is involved in aerial hyphae formation and attachment in the absence of the SC3 protein (9). This is the first report of RNA-mediated gene silencing in a filamentous homobasidiomycete.…”

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confidence: 96%

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RNA-Mediated Gene Silencing in Monokaryons and Dikaryons of Schizophyllum commune

Jong

Deelstra

Wösten

et al. 2006

Appl Environ Microbiol

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Disruption of genes by homologous recombination occurs at a low frequency in the basidiomycete Schizophyllum commune. For instance, the SC3 and SC15 genes were inactivated at frequencies of 1 and 5%, respectively. As an alternative to disruption, we used gene silencing through the introduction of a hairpin construct. The SC15 gene, which encodes an abundantly secreted structural protein, was silenced at a frequency of 80% in monokaryons of S. commune after introduction of a hairpin construct of the gene. Silencing also occurred in dikaryons in which one of the partners was not a silenced strain. The silencing mechanism resembles RNAi in other filamentous fungi and is a powerful tool for the functional analysis of genes expressed in monokaryons or dikaryons.

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Section: Discussionmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

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RNA-Mediated Gene Silencing in Monokaryons and Dikaryons of Schizophyllum commune

Jong

Deelstra

Wösten

et al. 2006

Appl Environ Microbiol

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“…Purification of SC3-S. commune strain D3 (⌬SC15⌬SC4) resulted from a cross between a strain that contains a disruption of the SC15 gene (20) and a strain containing a disruption of the SC4 gene (21). This strain was grown in production medium (22), and SC3 was purified from the culture medium as described (4,6).…”

Section: Methodsmentioning

confidence: 99%

Assembly of the Fungal SC3 Hydrophobin into Functional Amyloid Fibrils Depends on Its Concentration and Is Promoted by Cell Wall Polysaccharides

Scholtmeijer

Vocht

Rink

et al. 2009

Journal of Biological Chemistry

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Class I hydrophobins function in fungal growth and development by self-assembling at hydrophobic-hydrophilic interfaces into amyloid-like fibrils. SC3 of the mushroom-forming fungus Schizophyllum commune is the best studied class I hydrophobin. This protein spontaneously adopts the amyloid state at the water-air interface. In contrast, SC3 is arrested in an intermediate conformation at the interface between water and a hydrophobic solid such as polytetrafluoroethylene (PTFE; Teflon). This finding prompted us to study conditions that promote assembly of SC3 into amyloid fibrils. Here, we show that SC3 adopts the amyloid state at the water-PTFE interface at high concentration (300 g ml ؊1 ) and prolonged incubation (16 h).Moreover, we show that amyloid formation at both the water-air and water-PTFE interfaces is promoted by the cell wall components schizophyllan (␤(1-3),␤(1-6)-glucan) and ␤(1-3)-glucan. Hydrophobin concentration and cell wall polysaccharides thus contribute to the role of SC3 in formation of aerial hyphae and in hyphal attachment.

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“…Like the class I hydrophobin, the Rep1-1 peptide assembled into a surface-active rigid membrane at the water-air interface. The decrease in water surface tension to 36 mJ m Ϫ2 is similar to that of hydrophobins and other microbial surface-active peptides involved in aerial growth and attachment (35)(36)(37)(38)(39). The water-soluble form of the peptide, which seems to be an oligomer based on its behavior in SDS-PAGE, also aggregated readily in a SDS-insoluble form in the aqueous solution.…”

Section: Discussionmentioning

confidence: 56%

The Filament-specific Rep1-1 Repellent of the Phytopathogen Ustilago maydis Forms Functional Surface-active Amyloid-like Fibrils

Teertstra

Velden

Jong

et al. 2009

Journal of Biological Chemistry

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Repellents of the maize pathogen Ustilago maydis are involved in formation of hydrophobic aerial hyphae and in cellular attachment. These peptides, called Rep1-1 to Rep1-11, are encoded by the rep1 gene and result from cleavage of the precursor protein Rep1 during passage of the secretion pathway. Using green fluorescent protein as a reporter, we here show that rep1 is expressed in filaments and not in the yeast form of U. maydis. In situ hybridization localized rep1 mRNA in the apex of the filament, which correlates with the expected site of secretion of the repellents into the cell wall. We also produced a synthetic peptide, Rep1-1. This peptide reduced the water surface tension to as low as 36 mJ m ؊2 . In addition, it formed amyloid-like fibrils as was shown by negative staining, by thioflavin T fluorescence, and by x-ray diffraction. These fibrils were not soluble in SDS but could be dissociated with trifluoroacetic acid. The repellents in the hyphal cell wall had a similar solubility and also stained with thioflavin T, strongly indicating that they are present as amyloid fibrils. However, such fibrils could not be observed at the hyphal surface. This can be explained by the fact that the Rep1-1 filaments decrease in length at increasing concentrations. Taken together, we have identified the second class of fungal proteins that form functional amyloid-like filaments at the hyphal surface.Ustilago maydis is the causal agent of smut in Zea mays (maize) and Euchlaena mexicana (Mexican teosinte). A filamentous pathogenic dikaryon is formed upon fusion of compatible yeast-like sporidia. Differentiation in the plant leads to the formation of diploid teliospores, which undergo meiosis ultimately resulting in haploid sporidia (see Refs. 1-3).Fusion of haploid cells and development of an infectious dikaryon are controlled by the a and b mating type loci. The a locus regulates cell fusion through a pheromone-based recognition system (4), and the b locus controls post-fusion steps of pathogenic development, including hyphal growth. The latter locus encodes two unrelated homeodomain proteins, bE and bW, that form heterodimers when they are derived from different alleles (5, 6). This heterodimer regulates a number of genes, among which the rep1 gene (7-10). This gene is highly expressed resulting in 2.5% of the mRNA. It encodes a preproprotein that, after processing at KEX2 recognition sites, results in 11 secreted peptides with a high sequence similarity. These peptides are localized in the cell wall of filaments, in an SDSinsoluble, but trifluoroacetic acid-extractable form (10). They are involved in formation of hydrophobic aerial hyphae (10) and in hyphal attachment to hydrophobic surfaces (11); as such they have functionally replaced hydrophobins in U. maydis (11).Hydrophobins, which are not related to the repellents, fulfill a wide spectrum of functions in fungal development (12). They do so by forming an amphipathic protein film, which consists of amyloid-like fibrils (13-15). We here show that the repellent Rep1...

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The SC15 protein of Schizophyllum commune mediates formation of aerial hyphae and attachment in the absence of the SC3 hydrophobin

Cited by 45 publications

References 32 publications

RNA-Mediated Gene Silencing in Monokaryons and Dikaryons of Schizophyllum commune

RNA-Mediated Gene Silencing in Monokaryons and Dikaryons of Schizophyllum commune

Assembly of the Fungal SC3 Hydrophobin into Functional Amyloid Fibrils Depends on Its Concentration and Is Promoted by Cell Wall Polysaccharides

The Filament-specific Rep1-1 Repellent of the Phytopathogen Ustilago maydis Forms Functional Surface-active Amyloid-like Fibrils

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