Mushroom Schizophyllum Commune Research Articles

Effect of fruiting-related genes on the formation of volatile organic compounds in the mushroom Schizophyllum commune

In fungi, little is known about connections between volatile organic compound (VOC) formation and developmental stages that are amongst others triggered by fruiting-related genes (FRGs). We analysed the volatilomes of Schizophyllum commune during different developmental stages in a variety of FRG-deletion strains and wild-type strains. The deletion strains Δtea1Δtea1, Δwc-2Δwc-2 and Δhom2Δhom2 were unable to develop fruiting bodies, and Δfst4Δfst4 formed only rudimentary fruiting body structures. Early developmental stages of these strains were dominated by esters, including methyl 2-methylbutanoate, ethyl 2-methylbutanoate, isobutyl 2-methylpropionate, and 2-methylbutyl acetate, of which the last three were not found in the headspace (HS) of the wild-type samples. Compared to the wild type, in the HS of hom2con samples, that are able to form fruiting bodies, methyl 2-methylbutanoate was the most abundant substance at early stages (68–81% of the total peak area). In contrast to fruiting body forming strains, Δtea1Δtea1, Δwc-2Δwc-2, Δhom2Δhom2 and Δfst4Δfst4 showed less sesquiterpenes in the HS. However, the sesquiterpenes found in the HS of FRG-deletion strains, namely, (E)-nerolidol, δ-cadinene, L-calamenene, α-bisabolol and β-bisabolene, were not present in hom2con or wild-type strains that mainly formed fruiting bodies and barely mycelium. Several sesquiterpenes, including α-guaiene, chamigrene and γ-gurjunene, were only found in presence of fruiting bodies. Our results show remarkable connections between FRGs, fruiting body development and VOC production in S. commune, especially counting for sesquiterpenes. Future studies are needed to reveal whether FRGs directly regulates VOC formation or indirectly by changing the phenotype.

Identification of Long Non-Coding RNAs and Their Target Genes from Mycelium and Primordium in Model Mushroom Schizophyllum commune

Schizophyllum commune has emerged as the most promising model mushroom to study developmental stages (mycelium, primordium), which are two primary processes of fruit body development. Long non-coding RNA (lncRNA) has been proved to participate in fruit development and sex differentiation in fungi. However, potential lncRNAs have not been identified in S. commune from mycelium to primordium developmental stages. In this study, lncRNA-seq was performed in S. commune and 61.56 Gb clean data were generated from mycelium and primordium developmental stages. Furthermore, 191 lncRNAs had been obtained and a total of 49 lncRNAs were classified as differently expressed lncRNAs. Additionally, 26 up-regulated differently expressed lncRNAs and 23 down-regulated between mycelium and primordia libraries were detected. Further, Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis showed that differentially expressed lncRNAs target genes from the MAPK pathway, phosphatidylinositol signal, ubiquitin-mediated proteolysis, autophagy, and cell cycle. This study provides a new resource for further research on the relationship between lncRNA and two developmental stages (mycelium, primordium) in S. commune.

The Transcription Factor Roc1 Is a Key Regulator of Cellulose Degradation in the Wood-Decaying Mushroom Schizophyllum commune.

ABSTRACTWood-decaying fungi of the class Agaricomycetes (phylum Basidiomycota) are saprotrophs that break down lignocellulose and play an important role in nutrient recycling. They secrete a wide range of extracellular plant cell wall degrading enzymes that break down cellulose, hemicellulose, and lignin, the main building blocks of plant biomass. Although the production of these enzymes is regulated mainly at the transcriptional level, no activating regulators have been identified in any wood-decaying fungus in the class Agaricomycetes. We studied the regulation of cellulase expression in the wood-decaying fungus Schizophyllum commune. Comparative genomics and transcriptomics on two wild isolates revealed a Zn2Cys6-type transcription factor gene (roc1) that was highly upregulated during growth on cellulose, compared to glucose. It is only conserved in the class Agaricomycetes. A roc1 knockout strain showed an inability to grow on medium with cellulose as sole carbon source, and growth on cellobiose and xylan (other components of wood) was inhibited. Growth on non-wood-related carbon sources was not inhibited. Cellulase gene expression and enzyme activity were reduced in the Δroc1 strain. ChIP-Seq identified 1474 binding sites of the Roc1 transcription factor. Promoters of genes involved in lignocellulose degradation were enriched with these binding sites, especially those of LPMO (lytic polysaccharide monooxygenase) CAZymes, indicating that Roc1 directly regulates these genes. A conserved motif was identified as the binding site of Roc1, which was confirmed by a functional promoter analysis. Together, Roc1 is a key regulator of cellulose degradation and the first identified in wood-decaying fungi in the phylum Basidiomycota.

Enhancement of Potential Health Benefits of Insoluble Dietary Fiber from the Fruiting Body of Medicinal Mushroom Schizophyllum commune (Agaricomycetes) through Superfine Pulverization.

The objective of the current study is to reveal the influence of particle size of ground Schizophyllum commune and its extracted dietary fiber (DF) on physicochemical and antioxidant properties. Sch. commune powder and the extracted DF was ground by regular and superfine grinding, and their particle sizes were determined using laser diffraction particle size analyzer. The results indicated that superfine grinding could significantly pulverize DF particles to micro-scale; the particle size distribution presented a Gaussian distribution. The soluble DF in Sch. commune was increased effectively with superfine grinding. Sub-micronized insoluble DF showed increased total phenolic content (TPC) and ferric reducing antioxidant power (FRAP). Moreover, with particle size reduction, the oil binding capacity (OBC), nitrite ion absorption capacity (NIAC), cation-exchange capacity (CEC), cholesterol absorption capacity (CAC), and Pb ion adsorption capacity were significantly (p < 0.05) increased and the water retention capacity (WRC), swelling capacity (SC) and Cu, Zn ions adsorption capacity had no significant changes. A kind of health beneficial DF with higher soluble DF content, OBC, NIAC, CEC, CAC, Pb ion adsorption capacity and antioxidant activity (TPC and FRAP) was obtained through superfine grinding. Sch. commune DF could be potentially used as an ingredient for functional food and nutraceuticals.

H3K4me2 ChIP-Seq reveals the epigenetic landscape during mushroom formation and novel developmental regulators of Schizophyllum commune

Mushroom formation represents the most complex multicellular development in fungi. In the model mushroom Schizophyllum commune, comparative genomics and transcriptomics have previously resulted in a regulatory model of mushroom development. However, little is known about the role of epigenetic regulation. We used chromatin immunoprecipitation sequencing (ChIP-Seq) to determine the distribution of dimethylation of lysine 4 on histone H3 (H3K4me2), a mark for transcriptionally active genes, during monokaryotic and dikaryotic development. We identified a total of 6032 and 5889 sites during monokaryotic and dikaryotic development, respectively. The sites were strongly enriched near translation initiation sites of genes. Although the overall epigenetic landscape was similar between both conditions, we identified 837 sites of differential enrichment during monokaryotic or dikaryotic development, associated with 965 genes. Six transcription factor genes were enriched in H3K4me2 during dikaryotic development, indicating that these are epigenetically regulated during development. Deletion of two of these genes (fst1 and zfc7) resulted in arrested development of fruiting bodies, resulting in immature mushrooms. Together these results indicate that H3K4me2 ChIP-Seq is a powerful new tool to map the restructuring of the epigenetic landscape during mushroom development. Moreover, it can be used to identify novel developmental regulators.

A study of chitosan and glucosamine isolated from Sri Lankan local mushroom Schizophyllum commune and oyster mushroom (Pleurotus ostreatus)

Abstract Properties of chitosan extracted from local mushroom Schizophyllum commune and Pleurotus ostreatus were compared. Percentage yield, solubility, moisture-content, ash-content, N-content, water-binding-capacity were 1.73 ± 0.05%/1.22 ± 0.01%, 7.38 ± 0.10%/3.41 ± 0.14%, 6.39 ± 0.20%/8.16 ± 0.42%, 8.19 ± 0.04%/1.63 ± 0.05%, 11.31%/3.02%, 387.13 ± 15.57%/402.57 ± 12.78% respectively for chitosan samples from S. commune followed by P. ostreatus. FBC were varied approximately 250–350% in coconut oil, soy bean oil and sunflower oil. DD% of chitosan using FT-IR and conductometric titration were 53.10% and 60.68% respectively for two species. The yield and purity of glucosamine sample 95.70% and 0.97 ± 0.08% and 58.14% and 0.52 ± 0.04% for Schizophyllum commune and Pleurotus ostreatus were respectively.

High-throughput targeted gene deletion in the model mushroom Schizophyllum commune using pre-assembled Cas9 ribonucleoproteins

Efficient gene deletion methods are essential for the high-throughput study of gene function. Compared to most ascomycete model systems, gene deletion is more laborious in mushroom-forming basidiomycetes due to the relatively low incidence of homologous recombination (HR) and relatively high incidence of non-homologous end-joining (NHEJ). Here, we describe the use of pre-assembled Cas9-sgRNA ribonucleoproteins (RNPs) to efficiently delete the homeodomain transcription factor gene hom2 in the mushroom-forming basidiomycete Schizophyllum commune by replacing it with a selectable marker. All components (Cas9 protein, sgRNA, and repair template with selectable marker) were supplied to wild type protoplasts by PEG-mediated transformation, abolishing the need to optimize the expression of cas9 and sgRNAs. A Δku80 background further increased the efficiency of gene deletion. A repair template with homology arms of 250 bp was sufficient to efficiently induce homologous recombination. This is the first report of the use of pre-assembled Cas9 RNPs in a mushroom-forming basidiomycete and this approach may also improve the genetic accessibility of non-model species.

The wound healing effect of four types of beta-glucan

Beta-glucans, which existed in the cell walls of cereals, bacteria, and fungi, comprise a group of β-d-glucose polysaccharides. We investigated the effects of four kinds of beta-glucan, that are derived from barley, yeast, mushroom, and euglena on wound healing. The migration and viability of keratinocyte or fibroblast were analyzed using the in vitro scratch wound healing assay, invasion assay, MTT assay, and in vivo assay. All the beta-glucans had a significant effect on keratinocyte migration at 20 μM and showed no toxicity on dermal fibroblast. Moreover, treatment of keratinocytes with the beta-glucan derived from the mushroom (Schizophyllum commune) promoted in vivo wound closure. The Integrin/FAK/Src pathway is known to affect cell migration by forming lamellipodia. Beta-glucan from S. commune activates the Integrin/FAK/Src signaling pathway in a time-dependent. Reactive oxygen species are associated with fibroblast differentiation to contract dermal layer and synthesize collagens. We found that fibroblast was activated by increasing NOX4 expression. We propose that beta-glucan derived from mushroom is capable of promoting keratinocyte migration via the induction of FAK/Src phosphorylation there by accelerating wound closure and activating dermal fibroblast differentiation through NADPH oxidase for matrix remodeling.

Identification and expression analysis of Pofst3 suggests a role during Pleurotus ostreatus primordia formation

Only a few transcription factors involved in mushroom development have been reported till now, Therefore, identification of transcription factors in common edible mushroom has commercial and scientific importance. In this study, the Pofst3 gene from Pleurotus ostreatus was cloned and characterized. Bioinformatics analysis showed that Pofst3 protein had 71% sequence similarity with fst3 of model mushroom Schizophyllum commune. Furthermore, the function of Pofst3 gene was analyzed by overexpression and antisense silencing in P.ostreatus via Agrobacterium-mediated transformation. Expression verification of Pofst3 in transformants through qRT-PCR showed that compared with the wild type strains, the transcription level was about 1.26 ∼ 9.59 and 0.01 ∼ 0.30 fold in Pofst3 overexpressing and silencing strains, respectively. Petri dish and bag cultivation tests of transgenic strains showed that the number of primordia and the type of fruiting bodies of Pofst3 overexpressing strains were consistent with the wild type strains, i.e. fewer primordia and larger fruiting bodies; the number of primordia formed by Pofst3 silencing strains were more than those of wild type strains, but fruiting bodies were smaller. It was very likely that Pofst3 was involved in the regulation of P.ostreatus development through inhibiting the formation of clusters of primordia.

Morphological Characteristics and Kinship Relationship of Mushroom Schizophyllum commune Fr.

Schizophyllum commune is a mushroom from the division of Basidiomycota which abundance at rainy season on wood that have been moldered for its habitat. S. commune has been distributed widely at various types of wood habitat (jack fruit, bamboo, rubber, mango etc). Existence of this mushroom has not been known as well as any mushroom are able to be consumed/edible yet. In Java, the societies who have known it as consumption mushroom called it with local name jamur gigit. Jamur gigit is potential to become food material and based on organoleptic test indicates that jamur gigit is more delicious from oyster mushroom. The purpose of research is to investigate the morphology character of S. commune at various growth phases and to study the S. commune relationship in Java. The result of this research is expected can give information about morphology character of S. commune in each its growth phase and to understand S. commune relationship in Java. Sample for research has been collected in 3 areas in Java. The macroscopic morphology character and microscopic character of S. commune were observed. The character that had been obtained is used to compile description and be analyzed by cluster using SPSS 13.0 to study the kinship relationship. The result of research indicates that morphology character of S. commune in Java of fruit body when old and young phase was different, while the microscopic character almost same to the obtained sample. The result of analysis cluster indicated that at similar level of 38,1 % there were two clusters, first cluster was Sidoarjo a S. commune and S. commune Yogyakarta. The second cluster has member the Sidoarjo b S. commune, Malang S. commune, Tangerang S. commune, Semarang S. commune and Kuningan S. commune. © 2015 JNSMR UIN Walisongo. All rights reserved.

Immobilized lipase from Schizophyllum commune ISTL04 for the production of fatty acids methyl esters from cyanobacterial oil

Novel lipase from model mushroom Schizophyllum commune strain ISTL04 produced by solid state fermentation of Leucaena leucocephala seeds, was immobilized onto Celite for enzymatic FAMEs production from cyanobacterial endolith Leptolyngbya ISTCY101. The isolate showed vigorous growth and produced remarkable lipase activity of 146.5 U g(-1) dry solid substrate, without any external lipase inducer. Single-factor experiments were carried out to study the effects of various reaction parameters on the FAMEs yield. The best conditions for enzymatic transesterification as revealed by the results were: 1:3 oil to methanol molar ratio, added at 3h intervals, 12% water content, 1581.5 U g(-1) immobilized lipase, temperature 45 °C, and time 24h. Under these conditions, the maximum FAMEs yield reached 94%. The immobilized lipase was able to produce >90% of the relative FAMEs yield after four repeated transesterification cycles. This immobilized lipase exhibited potential for application in biodiesel industry.

Production of (+)-valencene in the mushroom-forming fungus S. commune

Production of commercially interesting sesquiterpenes was previously examined in plants and microorganisms such as Escherichia coli and Saccharomyces cerevisiae. We here investigate the potential of the mushroom Schizophyllum commune for the production of sesquiterpenes. Genomic analysis of S. commune revealed that the mevalonate pathway required for the synthesis of the farnesyl diphosphate substrate for sesquiterpene production is operational. Introduction of a valencene synthase gene resulted in production of the sesquiterpene (+)-valencene, both in mycelium and in fruiting bodies. Levels of (+)-valencene in culture media of strains containing a mutated RGS regulatory protein gene (thn) were increased fourfold compared to those in wild-type transformants. Up to 16mgL(-1) (+)-valencene was produced in these strains. In addition, the amount of (+)-valencene containing n-dodecane recovered from the culture medium increased sixfold to sevenfold in the thn mutant strains due to the absence of schizophyllan.

Schizophyllan inhibits the development of mammary and hepatic carcinomas induced by 7,12 dimethylbenz(α)anthracene and decreases cell proliferation: comparison with tamoxifen

Breast cancer is one of the leading causes of cancer mortality among women. Some anticancer compounds have been isolated from mushrooms. The aim of the present work was to study the anticancer effects of schizophyllan (SCH), a β-D: -glucan extracted from the mushroom Schizophyllum commune alone or in combination with tamoxifen (TAM) on 7, 12 Dimethylbenz(α)anthracene (DMBA)-induced carcinomas in mice. We isolated SCH from S. commune. Female mice received DMBA, SCH, DMBA+SCH, DMBA+TAM or DMBA+TAM+SCH or vehicles. We studied mice survival, tumour incidence, histopathology, oestrogen receptor (ER) expression, cell proliferation by immunohistochemical detection of proliferating cell nuclear antigen (PCNA), apoptosis by TUNEL assay, as well as caspase-3 expression. DMBA treatment resulted in mammary and hepatocellular carcinomas (HCC). Both SCH and TAM reduced the incidence of DMBA-induced mammary tumours by 85 and 75%, respectively, and equally decreased the PCNA labelling index relative to DMBA. TAM treatment increased the incidence of- and PCNA index in HCCs relative to DMBA, while SCH suppressed these effects. TAM was more effective than SCH in the induction of apoptosis in both mammary and hepatic carcinomas. Caspase-3 levels correlated with the apoptotic index in most experimental groups. Only one dose of SCH had similar therapeutic effects against DMBA-induced mammary carcinomas as 4weeks of TAM treatment. This coupled with the ability of SCH to suppress hepatic lesions associated with TAM treatment provides the rationale for further investigating the combined therapeutic effects of TAM+SCH in preclinical models of ER-positive breast cancer, as well as in liver cancer.

Ras and Rho small G Proteins: Insights from the Schizophyllum commune genome sequence and comparisons to other fungi

Unlike in animal cells and yeasts, the Ras and Rho small G proteins and their regulators have not received extensive research attention in the case of the filamentous fungi. In an effort to begin to rectify this deficiency, the genome sequence of the basidiomycete mushroom Schizophyllum commune was searched for all known components of the Ras and Rho signalling pathways. The results of this study should provide an impetus for further detailed investigations into their role in polarized hyphal growth, sexual reproduction and fruiting body development. These processes have long been the targets for genetic and cell biological research in this fungus.

Transcriptome and Functional Analysis of Mating in the Basidiomycete Schizophyllum commune

In this study, we undertook a functional characterization and transcriptome analysis that enabled a comprehensive study of the mating type loci of the mushroom Schizophyllum commune. Induced expression of both the bar2 receptor and the bap2(2) pheromone gene within 6 to 12 h after mates' contact was demonstrated by quantitative real-time PCR. Similar temporal expression patterns were confirmed for the allelic bbr1 receptor and bbp1 pheromone-encoding genes by Northern hybridization. Interestingly, the fusion of clamp connections to the subterminal cell was delayed in mating interactions in which one of the compatible partners expressed the bar2 receptor with a truncated C terminus. This developmental delay allowed the visualization of a green fluorescent protein (Gfp)-labeled truncated receptor at the cell periphery, consistent with a localization in the plasma membrane of unfused pseudoclamps. This finding does not support hypotheses envisioning a receptor localization to the nuclear membrane facilitating recognition between the two different nuclei present in each dikaryotic cell. Rather, Gfp fluorescence observed in such pseudoclamps indicated a role of receptor-pheromone interaction in clamp fusion. Transcriptome changes associated with mating interactions were analyzed in order to identify a role for pheromone-receptor interactions. We detected a total of 89 genes that were transcriptionally regulated in a mating type locus A-dependent manner, employing a cutoff of 5-fold changes in transcript abundance. Upregulation in cell cycle-related genes and downregulation of genes involved in metabolism were seen with this set of experiments. In contrast, mating type locus B-dependent transcriptome changes were observed in 208 genes, with a specific impact on genes related to cell wall and membrane metabolism, stress response, and the redox status of the cell.

A novel ribonuclease with potent HIV-1 reverse transcriptase inhibitory activity from cultured mushroom Schizophyllum commune

A 20-kDa ribonuclease (RNase) was purified from fresh fruiting bodies of cultured Schizophyllum commune mushrooms. The RNase was not adsorbed on Affi-gel blue gel but adsorbed on DEAE-cellulose and CM-cellulose. It exhibited maximal RNase activity at pH 6.0 and 70°C. It demonstrated the highest ribonucleolytic activity toward poly (U) (379.5 μ/mg), the second highest activity toward poly (C) (244.7 μ/mg), less activity toward poly (A) (167.4 μ/mg), and much weaker activity toward poly (G) (114.5 μ/mg). The RNase inhibited HIV-1 reverse transcriptase with an IC(50) of 65 μM. No effect on [(3)H-methyl]-thymidine uptake by lymphoma MBL2 cells and leukemia L1210 cells was observed at 100 μM concentration of the RNase. A comparison of RNases from S. commune and Volvariella volvacea revealed that they demonstrated some similarities in N-terminal amino acid sequence, optimum pH and polyhomoribonucleotide specificity. However, some differences in chromatographic behavior and molecular mass were observed.

Abstract B53: Cytotoxic effect of 21kDa protein and antitumor effect of 31KDa protein derived from mushroom, schizophyllum commune

Abstract The present study attempts to the purified proteins from mushrooms that way activate and stimulate the proliferation of immune cell, resulting in the onset phagocytosis and nature killer cell (NK cell) activity, and promotes the cytotoxicity of tumor cell. In this study 21 and 30 KDa protein from fermentation liquid of the mushroom Schizophyllum commune were purified that exhibit a distinct cytotoxic capability: The 21 KDa protein was demonstrated to be capable of stimulating the NK cell activity, while the 30 KDa protein prevent the proliferation of Lewis lung carcinoma in vitro. Furthermore it was also forward that 21 KDa protein stimulate the the proliferation of mouse splenocyte at a various concentration levels without agglutinating the red blood cell of ICR mouse. Our result showed that when we used concentration level of 27 µg/ml protein, a significant activity of NK cell was obtained (p&amp;lt; 0.05). In addition 30 KDa protein may activate macrophage RAW264.7 enhance to production of nitric oxide and TNF-α, However, the production of nitric oxide by RAW264.7 was relied on the dose of 30 KDa protein in vitro. Based on the results, the purified proteins protein from Schizophyllum commune were potent immue stimulants and has a potential for development as a novel pharmacological agent. Citation Information: Clin Cancer Res 2010;16(14 Suppl):B53.

Genome sequence of the model mushroom Schizophyllum commune

Much remains to be learned about the biology of mushroom-forming fungi, which are an important source of food, secondary metabolites and industrial enzymes. The wood-degrading fungus Schizophyllum commune is both a genetically tractable model for studying mushroom development and a likely source of enzymes capable of efficient degradation of lignocellulosic biomass. Comparative analyses of its 38.5-megabase genome, which encodes 13,210 predicted genes, reveal the species's unique wood-degrading machinery. One-third of the 471 genes predicted to encode transcription factors are differentially expressed during sexual development of S. commune. Whereas inactivation of one of these, fst4, prevented mushroom formation, inactivation of another, fst3, resulted in more, albeit smaller, mushrooms than in the wild-type fungus. Antisense transcripts may also have a role in the formation of fruiting bodies. Better insight into the mechanisms underlying mushroom formation should affect commercial production of mushrooms and their industrial use for producing enzymes and pharmaceuticals.

Schizolysin, a hemolysin from the split gill mushroom Schizophyllum commune

Abstract A monomeric hemolysin with a molecular mass of 29 kDa was isolated from fresh fruiting bodies of the split gill mushroom Schizophyllum commune. The hemolysin was purified by successive adsorption on DEAE-cellulose, carboxymethyl-cellulose and Q-Sepharose and finally gel filtration on Superdex 75. This demonstrated the N-terminal sequence ATNYNKCPGA, different from those of previously reported fungal and bacterial hemolysins. The hemolysin was stable up to 40 degrees C. Only partial activity remained at 50 and 60 degrees C. Activity was indiscernible at 70 degrees C. A pH of 6.0 was optimal for activity. The hemolytic activity was most potently inhibited by dithiothreitol, sucrose and raffinose, followed by cellobiose, maltose, rhamnose, inulin, lactose, fructose and inositol. The metal ions Cu(2+), Mg(2+), Zn(2+), Al(3+) and Fe(3+) significantly, and Pb(2+) to a lesser extent, curtailed the activity of the hemolysin. The hemolysin inhibited HIV-1 reverse transcriptase with an IC(50) of 1.8 microM.

Fungal GPCR signaling in pheromone response

The mushroom Schizophyllum commune shows several phenotypes linked to pheromone signaling and Ras-dependent development. The genes involved in MAPK signaling, cAMP dependent signaling and Rho signaling all turned on after pheromone recognition in a Ras-dependent manner have been analyzed using the genome sequence. Receptor-pheromone recognition leads to mating and is dependent in mated dikaryons for nuclear migration and clamp cell fusion. In vitro mutagenesis could show domains in the protein involved in ligand recognition. Receptor localization and expression studies could elucidate the involvment in clamp cell fusion. The signal is transmitted through a MAPK cascade and via Ras. Ras constitutive mutant alleles transformed into a wildtype recipient as well as deletion of a RasGap lead to enhanced intracellular signaling. By this approach phenotypes in clamp cel fusion, mushroom development and meiosis, as well as hyphal directional growth can be linked to Ras signaling. The genome sequence allowed us to identify components, which are used for expression studies during pheromone response and mating. A proteome analysis additionally is used to define components involved in pheromone response in this easily tractable fungus.