VI Meeting on Genetics and Cellular Biology of Basidiomycetes (GCBB-VI)

Permanent URI for this collection

http://hdl.handle.net/2454/27028

Browse

Now showing 1 - 20 of 60

Open Access
Agaricus devoniensis complex comprises a group of heterothallic isolates constituting a basis for breeding
(Universidad Pública de Navarra / Nafarroako Unibertsitate Publikoa, 2006) Callac, P.; Spataro, C.; Lataillade, E.; Blasi, P.; Guinberteau, J.
A recent phylogenetic reconstruction of Agaricus section Duploannulati revealed that A. devoniensis and A. subfloccosus are two complexes of species close to A. bisporus. The A. subfloccosus complex comprises two homothallic entities, while the A. devoniensis complex was never studied until now. A sample of 26 isolates, some being unreliably determined, were examined to (i) confirm their identity using a PCR-RFLP marker revealing a characteristic A. devoniensis ITS polymorphism, and (ii) for their ability to fruit in standard conditions used for A. bisporus cultivation. Twenty one isolates were confirmed as A. devoniensis, and only two collections from USA were unable to fruit. The five remaining isolates were excluded from the complex and were unable to fruit; their ITS1+2 regions were sequenced and alignments indicated that four of them were similar to A. campestris and that one belonged to a new entity close to A. bitorquis and A. cappellianus. For the 19 fructifying isolates of the complex, we attempted intrastock and interstock mating tests with single spore isolates: for three isolates, we did not get spore germination; and for seven isolates, we observed partial to complete intersterility between strains. The nine remaining isolates exhibited a unifactorial system of sexual incompatibility for which eight different mating type alleles were detected. Within this group, the heterothallic and presumably interfertile isolates differed in their origin (Greece, France), their habitat (dune, coniferous trees), and their morphology (mean spore length: 5.6 to 6.6 μm); they constitute a diversified genetic basis usable to select smooth white and attractive cultivars for this tasteful edible and cultivable species.
Open Access
Anticancer activity of polysaccharides produced by Pleurotus ostreatus in submerged culture
(Universidad Pública de Navarra / Nafarroako Unibertsitate Publikoa, 2006) Daba, A.; El Demellawy, M.; El Eshasy, H.
It has been known for many years that some compounds produced by edible mushrooms encompass anticancer activities. Most of production methods were based on cultivation of mushroom in solid medium. In the present study Pleurotus ostreatus mycelia were grown in submerged culture. The cultivation of fungal cells in submerged culture resulted in higher growth rate with better control of production process. The bioactive polysaccharides (both intracellular and extracellular) were extracted from culture by solvent repeated precipitation. The polysaccharide structure was determined by examining NMR, IR spectra and the primary structure of the polysaccharide was mainly glucan. The 13C NMR spectral pattern indicated the polysaccharides are highly branched with mainly 1→3 and 1→6 linkage. The results of in vitro anti cancer studies demonstrate that this type of polysaccharides possesses anticancer activity against human oesophageal cancer cell line. Moreover, in the course of in vitro studies, mushroom polysaccharides showed anti-tumour activity and also considered to be biological response modifier because of their mechanism of action through stimulation of the immune system. The polysaccharide activity is especially beneficial in clinics when used as an adjuvant with chemotherapy to decrease its side effect. This work describes production process of anti cancer compound( s) by mushrooms and suitable for pharmaceutical industries.
Open Access
Atypical laccases from the white-rot fungus Pleurotus ostreatus and their application for the treatment of industrial coloured effluents
(Universidad Pública de Navarra / Nafarroako Unibertsitate Publikoa, 2006) Festa, Giovanna; Giardina, Paola; Faraco, Vicenza; Piscitelli, Alessandra; Sannia, Giovanni
White-rot fungi are the most efficient decomposers of lignocellulose because of their capability to synthesize the relevant hydrolytic (cellulases and hemicellulases) and oxidative (laccases, lignin-peroxidases and Mn-peroxidases) extracellular enzymes required to degrade the major components of substrates (cellulose, hemicellulose, and lignin) into low-molecular-weight compounds that can be assimilated in fungal nutrition [1]. Recently, extensive research on these fungi has been conducted with the aim of isolating new organisms able to secrete new enzymes with capability to be used in industrial applications, such as bioremediation of polluted soils and industrial waste-waters, biobleaching and biopulping in pulp and paper industries, textile and food industries, etc. Fungal laccases (benzenediol: oxygen oxidoreductases; EC1.10.3.2) are ligninolytic enzymes that have been isolated from various fungi [2]. They belong to the class of the blue oxidases containing 4 copper atoms/molecule distributed in three different copper binding sites [3, 4]. The type-1 site is responsible for the intense blue colour of the enzyme due to a maximum absorbance at 605 nm; the type-2 site does not exhibit signals in the visible absorbance spectrum; and the type-3 site incorporates two copper centres and is responsible for a band near 330 nm. All these copper ions are involved in the catalytic mechanism. Laccases reduce oxygen to water and simultaneously perform a one electron oxidation of aromatic substrates (polyphenols, methoxysubstituted monophenols, aromatic amines, etc.). These enzymes are present in multiple isoforms, depending on the fungal species and environmental growth conditions [5, 6].
Open Access
Biochemistry of volatile compounds synthesis in Agaricus bisporus
(Universidad Pública de Navarra / Nafarroako Unibertsitate Publikoa, 2006) Combet, E.; Henderson, J.; Eastwood, D.C.; Griffiths, G.; Burton, K.S.
Agaricus bisporus unique flavour is due to the release of a set of eight-carbon volatile compounds, which biosynthetic pathway has not been elucidated yet, despite of the numerous implications of those volatile compounds. Beside their influence on crop quality, they are also important for insect perception and play a part in triggering the switch from vegetative to reproductive growth in mushrooms. 8-carbon volatiles are derived from the oxygenation and the cleavage of the polyunsaturated fatty acid linoleic acid. This reaction has similarities to the plant system, but also major differences. Examination of the enzymic mechanisms and the fatty acid chemistry suggested that the enzyme involved in the oxygenation step could be a lipoxygenase (as found in plants) or a heme-dioxygenase, similar to the recently isolated linoleate diol synthase from Gaeumannomyces graminis. In order to characterise the biochemical pathway leading to eight-carbon volatile production, we investigated fatty acid and lipids distribution in Agaricus bisporus, as well as hydroperoxide and volatile compounds levels. In parallel, we searched for candidate genes susceptible to encode the enzyme responsible for this novel oxidation route in fungi. The combination of analytical methods, such as GC-MS, with a molecular approach based on degenerate PCR and library screening provided us with a broad range of results. These results establish the relation between fatty acids and volatile compounds and enabled us to gain a better understanding of mushroom volatiles biosynthesis and lipid metabolism.
Open Access
Cell wall-associated redox enzymes in white rot fungi
(Universidad Pública de Navarra / Nafarroako Unibertsitate Publikoa, 2006) Dwivedi, R.C.; Zomorrodi, M.; Kües, Ursula; Majcherczyk, A.
Many enzymes of white rot fungi involved in wood degradation belong to the class of redox enzymes. The most important are laccase (copper-containing polyphenol oxidase), lignin peroxidase, manganese- dependent peroxidase and manganese-independent peroxidase. However, the role of these enzymes in wood degradation remains unclear and complex redox processes or unknown redox enzymes also may contribute to this process. Several oxidative enzymes secreted by white rot fungi into the environment have been studied in the past, but little attention has been paid to the cell wall-associated redox enzymes. Cell wall-associated laccase activity in the purified cell walls of copper induced cultures of Trametes versicolor has been found. Laccases have been extracted by establishing new methods for cell wall purification and for protein release from the cell walls of basidiomycetes.
Open Access
Characterisation of Agaricus bisporus response genes to Verticillium fungicola infection
(Universidad Pública de Navarra / Nafarroako Unibertsitate Publikoa, 2006) Costa, A.; Thomas, D.J.I.; Bailey, A.; Foster, G.D.; Challen, M.P.; Mills, P.R.
The mycoparasite Verticillium fungicola is a persistent threat to the cultivation of the mushroom Agaricus bisporus. Mushroom “dry bubble” is characterised by an undifferentiated mass of cells and can result in major crop losses. During the establishment of “dry bubble” substantial changes occur in the biochemistry and physiology of both partners. To enable new insights to be made into the molecular events underlying the disease, work is in progress to identify genes expressed during pathogen infection. Subtractive Suppressive Hybridisation (SSH) has enabled recovery of 65 expressed sequenced tags (ESTs) differentially expressed during infection. After database searches 27 of the genes were identified as most likely from V. fungicola, 25 from A. bisporus and 13 unknown. Bioinformatic analysis suggested that the response genes identified were involved in a range of biological functions that included stress, signalling, protein synthesis and cell wall structure and function. Specific full-length genes will be recovered using cDNA library constructed from lesions of A. bisporus infected with V. fungicola, enabling silencing approaches to be used to further investigate the role of the identified genes in disease. An alternative higher-throughput method of gene function analysis, RNA interference (RNAi) using A. bisporus model genes (URA3, CBX), is also being developed. Silencing constructs expressing RNAi hairpin were transformed into A. bisporus using Agrobacterium tumefaciens and hygromycin resistance. Screening of the transformants by PCR confirmed integration of the silencing construct in 24 transformants. RT-PCR is being used to confirm transcription of the RNAi hairpin. Quantitative PCR will be used to analyse levels of target gene transcripts post RNAi transformation. The role of A. bisporus genes identified, in the infection process, will be determined through infection trails with A. bisporus silenced lines.
Open Access
Computational prediction of protein-coding gene and annotation of DNA sequences with agronomic interest in Pleurotus ostreatus (Oyster Mushroom)
(Universidad Pública de Navarra / Nafarroako Unibertsitate Publikoa, 2006) Palma Dovis, Leopoldo; Peñas Parrila, María Manuela; Ramírez Nasto, Lucía; Pisabarro de Lucas, Gerardo; Producción Agraria; Nekazaritza Ekoizpena
Pleurotus ostreatus, commonly known as oyster mushroom, is a commercially important edible fungus with interesting biotechnological properties. Quantitative trait loci (QTL) analyses are rare in fungi and little is known about their number, position, and genetic structure. Previous studies of our group have allowed the construction of a genetic linkage map of P. ostreatus var. florida, which has provided the basis for performing an efficient QTL analysis. In fact, there is a region of the chromosome VII of P. ostreatus where the most QTLs related to the production and precocity characters have been mapped. These quantitative traits are presumably under the control of a polygenic genetic system and could be associated with some chromosomal regions. The hypothesis of this work is that there is a region in the chromosome VII of protoclon PC15 (monokaryotic parental of the N001 dikaryotic strain) where exist genes which are responsible for the QTLs mentioned above. In order to test this hypothesis, we are developing a molecular QTL analysis through the sequencing of a region with an approximated size of 320 Kbp in chromosome VII (protoclon PC15). For this purpose, a BAC genomic library was constructed and two BAC clones spanning the region of interest are being sequenced. To carry out an efficient computational prediction of protein-coding genes and its annotation on the partial sequences obtained up to date, we have used different Internet resources such as BLASTx, BLASTp, BLASTn, and FGENESH trained on some basidiomycetes genomic data like Phanerochaete chrysosporium and Cryptococcus neoformans (SoftBerry). To our knowledge, this is the firs molecular QTL analysis performed on this edible mushroom.
Open Access
Copper in fruiting body development of Coprinus cinereus
(Universidad Pública de Navarra / Nafarroako Unibertsitate Publikoa, 2006) Navarro González, Mónica; Kilaru, S.; Majcherczyk, A.; Kües, Ursula
The model homobasidiomycete Coprinopsis cinerea grows best at 37°C, but, normally, it produces fruiting bodies only at moderate temperatures around 25-28°C. Light is needed to induce fruiting and also for fruiting body maturation. Cultures kept after fruiting induction predominantly in the dark form structures with an extended stipe and an underdeveloped cap (so-called “etiolated stipes”). In a day/night rhythm, caps develop further, basidia are formed, in which karyogamy and meiosis occurs and of which the basidiospores bud off. Besides light, fruiting body development in basidiomycetes has been repeatedly linked to enzymes belonging to the group of phenoloxidases, in particular the multi-copper containing laccases. However, their roles in fruiting remain unclear. In attempts to induce laccase production in liquid standing cultures at 37°C, to our surprise we found unusual inititation of fruiting body development. However, the abundantly formed primordia did never develop into mature fruiting bodies but into large-sized etiolated stipes, both in dark and in light. Laccase under these conditions was not detected in the medium but bound to the fruiting initiating mycelium. Moreover, enzyme production and etiolated stipe formation correlated with an increase from pH 5.5 to a slightly alkaline pH. Ammonium was found to be produced and nitrate reductase activity has enzymatically been shown. Under normal fruiting conditions, addition of copper to cultures enhances fruiting initiation in time and number. To further unravel the potential involvement of laccases in fruiting as well as of proteins influencing ammonia secretion, we are studying expression of corresponding genes during vegetative growth and fruiting body development. Work in our laboratory is supported by DBU (Deutsche Bundesstiftung Umwelt). MNG holds a CONACYT (Mexico) PhD studentship.
Open Access
The Coprinus cinereus genome project
(Universidad Pública de Navarra / Nafarroako Unibertsitate Publikoa, 2006) Gathman, A.; Lilly, W.; Stajich, Jason E.; Carlson, M.; Murphy, B.; Smith, A.; Fargo, D.; Dietrich, F.; Pukkila, P.J.
Coprinus cinereus is an increasingly attractive basidiomycete model system. Its genome has been sequenced and is publicly available; it is readily cultured in the laboratory on defined media, it has highly synchronous meiosis, and numerous laboratory techniques have been adapted for use with it. The 10X shotgun sequence released by the Whitehead Institute comprises 36 Mb of the 37 Mb of the genome, which have been assembled into 106 supercontigs containing 431 contigs. cDNA libraries have been constructed from two meiotic stages, and 1432 candidate genes have been identified from them. Another set of cDNA libraries has been constructed from vegetative Coprinus cinereus Okayama 7 grown under different environmental conditions, including heat shock, rapamycin treatment, minimal medium, rich medium, and complex carbon and nitrogen sources. 5000 ESTs are being sequenced from these libraries. The EST sequences have been aligned with the genomic sequence, as have known C. cinereus genes from GenBank. Data from known ascomycete gene sequences have been used to train SNAP software to predict a total of 11,340 genes from the remainder of the genome. BlastX and Pfam have been used to assign tentative functions to predicted genes as well as ESTs. tRNA genes have also been identified in the genome. All genomic information is available online via our Gbrowse server.
Open Access
Development of a sporeless strain of oyster mushroom (Pleurotus ostreatus)
(Universidad Pública de Navarra / Nafarroako Unibertsitate Publikoa, 2006) Baars, Johan; Sonnenberg, Anton S.M.; Hendrickx, Patrick
The enormous amounts of spores produced by oyster mushroom (Pleurotus ostreatus) cause lung-related health problems among employees working in oyster mushroom cultivation. If sporeless varieties are used for large-scale cultivation, these lung problems can be avoided. For development of a commercially attractive strain of sporeless oyster mushroom, strain ATCC 58937, a sporeless strain of oyster mushroom was used as a donor of the trait. Microscopic analysis of basidia showed that meiosis was aborted at an early stage. Both nuclear types that constitute strain ATCC 58937 could be retrieved by protoplasting. Protoplasting commercial strain HK35 yielded only one of its nuclear types. Crosses between the ATCC nuclear types and the HK35 nuclear types (either directly or using the Buller phenomenon) yielded normal sporulating strains, indicating that sporelessness was caused by a recessive trait. Among the offspring of crosses between the ATCC nuclear types and the HK35 nuclear types the sporeless trait segregated in a 1 to 1 ratio. The sporeless trait could be mapped and strongly linked genomic markers were developed. The breeding strategy was to successfully introduce sporelessness into both nuclear types of a commercial variety, to achieve a sporeless variety. In a first cross between a sporeless culture and a commercial strain, not only sporelessness was transferred to the commercial variety. Therefore, repeatedly backcrossing the progeny of the first cross with the commercial variety is used to try to restore the original genetic material from the commercial strain as much as possible. Performance of a number of “prototypes” of a sporeless oyster mushroom was tested on commercial mushrooms farms and proved to be satisfactory.
Open Access
Double stranded RNAs in the mushroom virus X complex of Agaricus bisporus
(Universidad Pública de Navarra / Nafarroako Unibertsitate Publikoa, 2006) Mills, P.R.; Challen, M.P.; Soares, A.; Grogan, H.; Maffettone, E.; Burrow, S.
Mushroom virus X (MVX) is a new disease of the commercial mushroom, Agaricus bisporus. When first recorded in 1996 it was associated with localised areas of pin suppression. It is now thought to be responsible for a range of symptoms. It seriously affected the UK mushroom industry in 2000/01 and has now been reported in Ireland, Holland and a number of other mushroom growing countries. The objectives of the work programme put in place to address this problem were to determine the relationship between major dsRNA bands present in the MVX complex and to sequence characterise major components; to develop an RT-PCR diagnostic test for separate viruses within the MVX complex and test commercial spawns and ARP cultures for virus presence; to characterise transmission of dsRNA elements between strains, assess effects of cross infection and determine significance of dsRNA partitioning during transmission through spores; and to determine the rate of spread of MVX into healthy compost from a point source Analysis of diseased mushrooms has identified 26 double stranded RNA (dsRNA) molecules associated with MVX symptoms. Evidence collected to date suggests that MVX might be a complex of viruses, rather than a single virus. Three of these dsRNAs have also been shown to be present in asymptomatic mushrooms. Vertical transmission was shown to occur through production of single spore progeny isolates from infected sporophores. Partitioning of dsRNAs was shown to occur. To determine whether horizontal transmission occurred, dual-culture in vitro tests were performed using donor and acceptor strains. Cultures produced from these transmission lines contained a reduced number of dsRNAs compared with the original donor strains. Sequence data has been generated for several dsRNAs.
Open Access
Ectopic expression of constitutively activated small GTPase cdc42 alters the morphology of haploid and dikaryotic hyphae in the filamentous homobasidiomycete Schizophyllum commune
(Universidad Pública de Navarra / Nafarroako Unibertsitate Publikoa, 2006) Raudaskoski, M.; Salo, V.; Weber, M.; Uuskallio, M.
Cloning of the Cdc42 gene from Schizophyllum commune enabled investigation of the role of ScCdc42 in the regulation of vegetative growth and sexual reproduction in this fungus, which has a well-characterized hyphal cell structure, cytoskeleton and mating system. Ectopic expression of the constitutively active Sccdc42G12V or Sccdc42Q61L allele from native or inducible ScCel1 promoters had dramatic effects on hyphal morphology, cytoskeletal structure and Cdc42 localization, while ectopic over-expression of the wt or the dominant negative ScCdc42D118A allele had no detectable effect. For transformants with constitutively active Sccdc42 tip growth of apical cells in the leading hyphae was normal but polar tip growth in side branches was altered implying different regulation of polarity establishment in the two groups of apical cells. The S. commune genome also contains a gene encoding RacGTPase. Rac1 might regulate the polarized growth of leading hyphae while ScCdc42 regulates the development of side branches in S. commune. In transformants with constitutively active Sccdc42 branch emergence at exceptional sites and isotropic growth next to the septum proved that the branch site selection and subsequent hyphal development are under ScCdc42 control. Poor dikaryotization along with irregular clamp connections in mates with Sccdc42G12V or Sccdc42Q61L allele suggested that Cdc42 also contributes to efficient mating in S. commune. Our results provide strong evidence that Cdc42 is involved in the control of hyphal morphogenesis in filamentous homobasidiomycetes.
Open Access
Enzymatic characterization of a monokaryon population of the edible mushroom, Pleurotus ostreatus with a view to genetic improvement
(Universidad Pública de Navarra / Nafarroako Unibertsitate Publikoa, 2006) Terrón, María del Carmen; López, María; Carbajo, José M.; Pisabarro de Lucas, Gerardo; Ramírez Nasto, Lucía; González Aldo, E.; Producción Agraria; Nekazaritza Ekoizpena
In this work the lignocellulolytic enzymes produced by the edible mushroom Pleurotus ostreatus var. florida were studied. The objective was to know their relationship with the degradation of the biopolymers present in the cell wall of wheat-straw for the purpose of explaining their influence on the production and quality characters of the fruiting bodies. The following enzymatic activities were studied both in solid and submerged culture: Ligninases (Lignin Peroxidase, Manganese Peroxidase (MnP) and Laccase), Cellulases (Glucohydrolases, Glucosidases) and Hemicellulases from the group Arabinofuran- Xylanases (Xylanase, Xilosidase, Glucoronidase, Arabinofuran-Oxidase and Acetylesterase), cooperating enzymes (Glyoxal Oxidase) and feedback enzymes (Glucose Oxidase (GOD), Aryl Alcohol Oxidase (AAO), Tyrosinase (TYR), Veratryl Alcohol Oxydase (VAO), Cellobiose Dehydrogenase (CDH)). The first studies regarding all the mentioned enzymes were performed using the dikayon (N001) and the parental monokarion strains “fast” (PC9) and “slow” (PC15). The studies on all this whole group of enzymes, which are enough representative of the lignocellulolytic complex, let to conclude that (both in solid or submerged culture) the enzymes of major influence in colonizing the natural substrate and also those whose activity-determination better guarantees their further mapping were Laccases, MnP, AAO and TYR. Subsequently these four activities were measured in the monokaryon population being Laccases and MnP, those yielding the best levels in medium-7 (rich in nitrogen). In addition both enzymes allow the discrimination between “fast-” or “slow-” monokaryon strains both in solid medium with several dyes, or in liquid culture in agitation. The analysis of the enzymatic activities detected in the assayed conditions, in the population of “fast” or “slow” strains let to the observation that they map in different places where the loci corresponding to Laccase (pox) and mnp genes are located. These results open the possibility to design more precise studies that could help to establish a correlation between the contribution of the genes already described and the activity of the different ligninolytic enzymes. In addition the results will contribute to know whether in P. ostreatus genome there are new genes or if they correspond with locations that regulate these enzymatic activities, or it is a gene that has a role in the transport system or a kind of effector in the exportation machinery of the protein to the culture medium.
Open Access
Evidence for a 200 gene ribosome and rRNA biosynthesis (rrb) regulon in fungi
(Universidad Pública de Navarra / Nafarroako Unibertsitate Publikoa, 2006) Wade, Christopher; Umbarger, Mark; McAlear, Michael A.
Two challenges of the post genomic era are 1) the need to assign functions to as yet uncharacterized gene products, and 2) the requirement to understand how the expression profiles of large sets of genes are regulated in response to changing environments. Towards these aims, we have used transcriptional profiling analysis to identify and characterize a large set (over 200 genes) of transcriptionally co-regulated genes whose products are involved in rRNA and ribosome biosynthesis. Many of the genes within this set were previously unknown with regards to their function. This RRB regulon is distinct from the ribosomal protein (RP) regulon, and is characterized by a unique pair of conserved promoter motifs. The organization of the RRB regulon appears to be evolutionarily conserved at least from S. cerevisiae to S. pombe. The strategies used to identify and characterize this gene set can be widely used in other organisms to help fulfill the two needs outlined above.
Open Access
Expression of GFP and DsRed in the homobasidiomycete Schizophyllum commune
(Universidad Pública de Navarra / Nafarroako Unibertsitate Publikoa, 2006) Vinck, Arman; Wösten, Han A. B.; Lugones, Luis G.
Expression of GFP and DsRed was studied in the homobasidiomycete Schizophyllum commune. CGFP or sGFP fused to the SC3 promoter resulted in similar steady state mRNA levels. These levels were considerably lower than that of SC3. Despite the low mRNA levels, both GFP variants resulted in fluorescent hyphae. The sGFP expressing strains showed stronger fluorescence than the CGFP expressers. When CGFP was fused to the N-terminal or C-terminal part of the mature SC3 protein, no fluorescence was observed. However, accumulation of GFP
Open Access
Expression of mating type genes in heterologous basidiomycetes
(Universidad Pública de Navarra / Nafarroako Unibertsitate Publikoa, 2006) Srivilai, Prayook; James, Timothy Y.; Vilgalys, Rytas; Chaiseana, Wassana; Kües, Ursula
Mating processes in basidiomycetes are controlled by genes encoding two types of transcription factors (HD1 and HD2) and by genes encoding pheromones and pheromone receptors. For a successful mating reaction, an HD1 protein and an HD2 protein of different specificity have to interact and a pheromone with a pheromone receptor of different specificity. With now having cloned mating type loci from several different basidiomycetes, evolution of these loci and their genes can be addressed by sequence analysis as well as by transformation of genes into heterologous species. Transformation of cloned mating type genes into strains of the same species with different mating type genes can activate mating type controlled development. The A mating type genes of Coprinopsis scobicola and of Coprinellus disseminatus were found to be functional in Coprinopsis cinerea in combination with the endogenous A mating type genes. Moreover, B genes of C. disseminatus in C. cinerea cause peg formation subapical to septa with A-induced clamps cells and fusion of clamp cells with the subapical peg. In several C. cinerea monokaryons, transformed A genes of Schizophyllum commune were not observed to induce clamp cells by interaction with endogenous A genes. However, in crosses of C. cinerea transformants carrying compatible S. commune A genes, clamp cell production has occasionally been observed. To our surprise, when using S. commune A genes or the homologous b genes of Ustilago maydis to transform a specific C. cinerea monokaryon, colonies of transformants may develop faster growing sectors with hyphae having clamp cells. Our laboratory is supported by Deutsche Bundesstiftung Umwelt (DBU) and scholarships by the Mahasarakham University (to PS) and the Rajamangala Institute of Technology (to WC).
Open Access
Expression profiling of natural antisense transcripts in Agaricus bisporus
(Universidad Pública de Navarra / Nafarroako Unibertsitate Publikoa, 2006) Eastwood, D.C.; Sergeant, M.; Mead, A.; Burton, K.S.
Our view of the role of RNA has changed from being a passive intermediary of genetic information to acting as a regulator of gene expression in the form of short-interfering RNA, micro RNA, riboswitches and natural antisense. Long length natural antisense transcripts (NATs) have been identified for genes which are up-regulated after harvest in the fruitbody of the mushroom Agaricus bisporus. These NATs therefore are likely to be involved in the regulation of postharvest events such as development and senescence. A novel quantitative reverse transcriptase PCR technique has been developed. The data have been statistically analysed to produce expression profiles of NATs for six postharvest genes. The average antisense/sense ratios varied by three orders of magnitude, from 8.0 for shs13 to 6 x 10-3 for cruciform DNA binding protein. The expression profiles were found to be highly specific for individual genes, to be dynamic over time and highly variable between neighbouring tissues. This latter characteristic has led to the speculation that NATs may be involved in tissue differentiation. Sequence information of natural antisense transcripts form A. bisporus suggests that they are synthesized from messenger (sense) RNA by RNA-dependent RNA-polymerase. Evidence will be presented that to support the hypothesis that the level of antisense may be controlled by the 3’ processing of sense RNA.
Open Access
Formation of hyphal loops in xylotrophic coprinoid mushrooms
(Universidad Pública de Navarra / Nafarroako Unibertsitate Publikoa, 2006) Badalyan, Suzanna M.; Kües, Ursula
Recent molecular analysis split the traditional genus Coprinus (Homobasidiomycetes) into four distinct genera: Coprinus, Coprinopsis, Coprinellus and Parasola. Coprinoid mushrooms are usually saprotrophic on soil and/or dung of herbivores. However, more than 60 species are able to grow on wood and straw. Xylotrophic mushrooms are forcing a relatively short supply of nitrogen and phosphorous nutrients. Coprinus comatus has been reported to produce specialized structures (“spiny balls”) to penetrate nematodes for nutrient supply (Luo et al. 2004, Mycologia 96, 1218-1224). Nematode traps of other fungi involve adhesive hyphal network and knobs, hyphal loops and snares. Toxin production may support in nematode immobilisation. Nematode-trapping species belong mainly to the mitosporic Deute romy - ce tes, but some are also found amongst Zygomycetes and Basidiomycetes. We have observed hyphal loops in several wood-decaying basidiomycetes, such as Daedalea quercina, Ganoderma lucidum, Lentinula edodes, Piptoporus betulinus and Pleurotus ostreatus. Furthermore, regular and irregular hyphal loops and/or rings were observed in the four clades of Coprinoid species (Coprinus comatus, Coprinellus angulatus, C. bisporus, C. curtus, C. domesticus, C. disseminatus, C. ellissi, C. micaceus, C. xanthothrix, Coprinopsis cinerea, C. gonophylla, C. radians, C. strossmayeri, C. scobicola, and P. plicatilis). Hyphal loops were particularly often formed in Coprinellus species. Such structures were rare in Coprinopsis atramentaria, C. cothurnata, C. romagnesiana, C. psychromorbida and Coprinus patouillardii (an unclassified isolate). It is not clear yet why Basidiomycetes fungi have these structures. Is it that many species have nematode trapping abilities by formation of such structures? Thanks to the DAAD, NATO and the Deutsche Bundesstiftung Umwelt for financial support.
Open Access
Fungal diversity adds value to biotechnology and agriculture
(Universidad Pública de Navarra / Nafarroako Unibertsitate Publikoa, 2006) Zervakis, Georgios I.
Mediterranean countries host rich biological diversity (genetic, population, species, habitats, communities, ecosystems). Until recently research on the fungal diversity was focusing relatively more on phytopathogenic fungi, invertebrate parasites, and saprotrophic and ectomycorrhizal mushrooms (Pezizales, higher Basidiomycetes). For higher Basidiomycetes in particular, detailed inventories and check-lists have been compiled in many western European countries. In the Mediterranean region, however, pertinent data are limited and fragmentary; only recently new information has started to accumulate. Indicative is the case of Greece, where selected ecosystems are studied in respect to their macromycetes diversity, revealing the existence of taxa with significant ecological and economic interest. Prerequisites for the exploitation of biological resources (incl. fungi) is the availability of a large number of individuals with a wide genetic basis, which are correctly identified and suitably evaluated. For example, elucidating taxonomy and clarifying phylogenetic relationships among Pleurotus species has contributed significantly to their widespread use. Large-scale applications related directly (or indirectly) with mushroom resources and their exploitation include the edible mushroom industry, production of medicinal and health-promoting factors, improvement of soil fertility, remediation of soils, enhanced plant growth, suppressiveness of soil-borne pathogens of plants, animal feed, transformation of xenobiotics and antibiotics, biosorption of toxic elements, decolorization of organic pollutants, degradation of industrial and agroforestry wastes, etc. Particular emphasis is given to the upgrade of lignocellulosic wastes and residues through their detoxification and biotransformation into value-added products; among them, soil conditioners and fertilizers generated from spent mushroom substrates conform with the much-sought notion of sustainability in agriculture.
Open Access
Fungal laccase: properties and aplications
(Universidad Pública de Navarra / Nafarroako Unibertsitate Publikoa, 2006) Jönsson, Leif J.
Laccase (EC 1.10.3.2; benzenediol:oxygen oxidoreductase) was first discovered at the end of the 19th century in the sap of Oriental lacquer trees. Later on, the laccase from the white-rot basidiomycete Trametes versicolor was thoroughly characterized using biochemical and biophysical methods. It is an extracellular blue multicopper glycoprotein. The copper ions are involved in the catalytic process, in which a reducing substrate, typically a phenol, is oxidized and molecular oxygen is reduced to water. Today, a multitude of different laccases and laccase genes from various sources have been characterized. The enzyme seems to have different physiological roles in different types of organisms. Several of the best characterized laccases come from basidiomycete fungi causing white-rot decay of wood. These laccases are generally regarded to be associated with the biodegradation of lignin, although more research is needed to shed light on the fundamental molecular mechanisms. Recent advances with regard to the structural and functional diversity of laccases will be discussed in relation to efforts to clarify the physiological roles of the enzyme and to elucidate its potential in various applications, including detoxification, bleaching, and analysis.