Spinning disc confocal microscopical research was conducted on living mating hyphae of the tetrapolar basidiomycete Schizophyllum commune. Haploid strains with either the same or different A and B mating-type genes and expressing differently labelled histone 2B were confronted. In the haploid hyphae histone 2B mCherry and histone 2B EGFP were visualized as red and green nuclei, respectively. In hyphae with the same A but different B genes, the red and green nuclei were observed next to each other. This indicated that nuclear migration between strains, regulated by the B mating type, had taken place. The compatible mating with different A and B genes produced a high number of mixed EFGP/mCherry, yellow nuclei. The mixed nuclei resulted from nearby divisions of nuclei encoding different histones and mating-type genes. During this process, the histones with the different labels were incorporated in the same nuclei, along with the heterodimerized transcription factors encoded by the different A mating-type genes and present around the nuclei. This led to the activation of the A-regulated pathway and indicated that different A genes are important to the cell cycle activation of a compatible mating. Consequently, a yellow nuclear pair stuck together, divided synchronously and proceeded in the migration hyphae towards the colony periphery, where the dikaryotization was promoted by branch formation from the migration hyphae.

The fertilization process of filamentous tetrapolar basidiomycetes is described as a process in which the reciprocal nuclear invasion into the mycelium of a different mating type involves septal dissolution and the migration of nuclei through the foreign colony until a hyphal tip cell is reached. Spinning disc confocal microscopical research on this process was conducted on living hyphae using Schizophyllum commune haploid strains with compatible and hemicompatible A and B mating type genes and, in one of the strains, with nuclei expressing histone 2B mCherry and the other histone 2B labeled with EGFP. In haploid hyphae, the labeled histones stay strictly in the nucleus. In the multinucleate migration hyphae formed in the mating with the same A but different B mating-type genes (A=B≠), mCherry and EGFP nuclei were detected, indicating that the reciprocal nuclear invasion due to the activated B pathway had taken place. In the compatible mating with different A and B mating type genes (A≠B≠) the migration hyphae were occupied by individual mCherry and EGFP nuclei, but also by coexpressed mCherry and EGFP nuclei. These nuclei result from the incorporation of histone H2B-mCherry and -EGFP into the same nucleus and requires nuclear divisions in close proximity activated by the A-regulated pathway. As a consequence, the nuclear pairs will stay together, divide synchronously and proceed in the migration hypha towards the colony periphery. The dikaryotization process is promoted by branch formation from the migration hyphae, together with the movement of the nuclear pairs with an activated A- and B- pathway from the migration hypha into the branch for synchronous nuclear division and clamp cell formation.

Kinesins are essential motor molecules of the microtubule cytoskeleton. All eukaryotic organisms have several genes encoding kinesin proteins, which are necessary for various cell biological functions. During the vegetative growth of filamentous basidiomycetes, the apical cells of long leading hyphae have microtubules extending toward the tip. The reciprocal exchange and migration of nuclei between haploid hyphae at mating is also dependent on cytoskeletal structures, including the microtubules and their motor molecules. In dikaryotic hyphae, resulting from a compatible mating, the nuclear location, synchronous nuclear division, and extensive nuclear separation at telophase are microtubule-dependent processes that involve unidentified molecular motors. The genome of Schizophyllum commune is analyzed as an example of a species belonging to the Basidiomycota subclass, Agaricomycetes. In this subclass, the investigation of cell biology is restricted to a few species. Instead, the whole genome sequences of several species are now available. The analyses of the mating type genes and the genes necessary for fruiting body formation or wood degrading enzymes in several genomes of Agaricomycetes have shown that they are controlled by comparable systems. This supports the idea that the genes regulating the cell biological process in a model fungus, such as the genes encoding kinesin motor molecules, are also functional in other filamentous Agaricomycetes.

The purpose of the present research was to observe in the filamentous basidiomycete Schizophyllum commune, the connection between the nuclear division and polymerization of the contractile actin ring with subsequent formation of septa in living hyphae. The filamentous actin was visualized using Lifeact-mCherry and the nuclei with EGFP tagged histone 2B (H2B). Time-lapse fluorescence microscopy confirmed that in monokaryotic and dikaryotic hyphae, the first signs of the contractile actin ring occur at the site of the nuclear division, in one to two minutes after division. At this stage, the telophase nuclei have moved tens of micrometers from the division site. The actin ring is replaced by the septum in six minutes. The apical cells treated with filamentous actin disrupting drug latrunculin A, had swollen tips but the cells were longer than in control samples due to the absence of the actin rings. The nuclear pairing and association with clamp cell development as well as the clamp cell fusion with the subapical cell was disrupted in latrunculin-treated dikaryotic hyphae, indicating that actin filaments are involved in these processes, also regulated by the A and B mating-type genes. This suggests that the actin cytoskeleton may indirectly be a target for mating-type genes.

Basidiomycetes feature a prolonged dikaryotic life stage. A dispute over open versus closed mitosis could be solved using in vivo fluorescence videomicroscopy of histone 2B::EGFP and Lifeact labeled Schizophyllum commune. It revealed nuclei to condense to approximately one fifth in diameter during mitotic prophase. In addition, the specifics of clamp cell formation typical of many basidiomycetes included an actin network at the future site of nuclear division, which allowed for cessation of nuclear movement and re-localization of one nucleus towards the emerging clamp cell, while the other divided along the hyphal axis. Subsequent fusion of the clamp cell to form the clamp connection restored the close association of the two nuclei in a very fast process after clamp fusion. Septation was preceded by actin patches and vesicles involved in formation of the actin ring.

The availability of genome sequences from both arbuscular and ectomycorrhizal fungi and their hosts has, together with elegant biochemical and molecular biological analyses, provided new information on signal exchange between the partners in mycorrhizal associations. The progress in understanding cellular processes has been more rapid in arbuscular than ectomycorrhizal symbiosis due to its similarities of early processes with Rhizobium -legume symbiosis. In ectomycorrhiza, the role of auxin and ethylene produced by both fungus and host plant is becoming understood at the molecular level, although the actual ligands and receptors leading to ectomycorrhizal symbiosis have not yet been discovered. For both systems, the functions of small effector proteins secreted from the respective fungus and taken up into the plant cell may be pivotal in understanding the attenuation of host defense. We review the subject by comparing cross-talk between fungal and plant partners during formation and establishment of arbuscular and ectomycorrhizal symbioses.

In the filamentous basidiomycetes Coprinopsis cinerea and Schizophyllum commune, mating is regulated by the tetrapolar mating-type system consisting of two unlinked genetic complexes, named A and B. In the nineties, the molecular structure of A and B mating type loci and genes was revealed side by side in C. cinerea and S. commune, first the A complex and quite soon thereafter the B complex genes. The clear molecular structure of C. cinerea mating type genes has led to their use as models for genomic approaches to investigate several other filamentous basidiomycetes. In filamentous fungi, hyphal fusions are important for the distribution of available nutrients in a fungal colony. In ascomycetes and basidiomycetes they are also important for sexual reproduction. These aspects have been approached, especially in filamentous ascomycetes, but in filamentous basidiomycetes the role of fusions in the fungal life cycle has received less attention. Several proteins encoded by the genes required for hyphal fusion in filamentous ascomycetes show homology with proteins forming the striatin-interacting phosphatase and kinase (STRIPAK) complex in eukaryotic cells. Homologs to the genes encoding STRIPAK complex proteins can be identified in C. cinerea and S. commune genomes suggesting that a STRIPAK-like complex could also regulate hyphal fusions of filamentous basidiomycetes. The STRIPAK complex is a conserved signaling complex also homologous to the Saccharomyces cerevisiae Far complex involved in cell cycle arrest at yeast mating, while the fission yeast Schizosaccharomyces pombe SIP complex is involved in transition signaling from mitosis to cytokinesis. In filamentous basidiomycetes the signaling pathway regulated by the B mating type genes, the pheromone response pathway, is assumed to follow the same pattern as in the yeast S. cerevisiae. This poses an interesting question as to the relationship between the STRIPAK complex proteins and the proteins in cellular processes such as cell cycle, septal dissolution, nuclear migration, clamp cell development and fusion, known to take place after hyphal fusions. All these processes are also dependent on the activation of the B and A mating type pathways in C. cinerea and S. commune.

ABSTRACT 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.

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.

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.

Microbiology includes all three domains of life in its research subjects, archaea, bacteria and eukarya. From the eukaryotes, the kingdom of true fungi, the Eumycota, are specifically subjects which can only be investigated by microbiological and molecular genetic techniques. This journal receives and publishes on a regular basis articles dealing with fungi, ascomycetes as well as basidiomycetes, and to a lesser extend also zygomyetes. These organisms differ, in many respects, from the prokaryotic organisms also investigated in microbiology, and therefore warrant specific attention in their own rights. With this Special Issue on “Basic Mycology” we wanted to address the fungi and show their versatility and the basic insights that can be gained from studying these eukaryotic microbes. Fungi create a versatile source of metabolites and enzymes with potential for biotechnological applications. Journal of Basic Microbiology receives, on a regular basis, numerous manuscripts dealing with production of fungal enzymes and metabolites, as well as with fungal biology. Knowing that many of our authors and readers are interested in the fungal laccases, we include here one review which gives insight into the role of lignin modifying enzymes in filamentous basidiomycetes and clarifies the relationships between lignin peroxidases and laccases. For industrial and biotechnological purposes, laccases were, and still are, of major importance, as these fungal oxidoreductases can be used in removal of polyphenols, e.g. in wine and beverages, toxic dyes from waste waters, or in bleaching for paper industry. This is reflected in three original articles in this Special Issue dealing with laccases, which is currently attracting more attention, as the ecological importance of litter degradation is being explored in more detail. Two more papers are dealing with fungal lipases, also showing the ability of fungi for biotechnology. Lipases from microbes also have applications in food, pharmaceutical and cosmetic industry. In both cases, aspergilli were used for the applications of basic mechanisms in fungi. Also using Aspergillus as a model, lectins and the production of a compound known from rose oil were investigated. The latter shows, that the isolation of new strains, in this case from within rose tissue, might reveal interesting insights into basic principles. Not an endophytic, but a pathogenic interaction is seen with rust and smut fungi and their plant hosts. A means to determine geographical variation among different strains is shown for Perenospora . A second review article is dealing with the well‐studied model of using different yeasts, in this case to study metal resistance mechanisms. Here, basic insights into chromium resistance are given. Two original papers within this Special Issue relate to this review: one also dealing with heavy metal resistance versus sensitivity, showing involvement of cyclic AMP in this process. The second paper is also using a yeast, Saccharomyces cerevisiae , where the uptake of fluorescently labeled DNA can be visualized. All in all, with articles covering the fungal kingdom with ascomycetes – from unicellular yeasts through aspergilli and Fusarium – over rust and smut fungi to laccase producing basidiomycetes, we hope that we can provide with this Special Issue on “Basic Mycology” an overview over current research on this special clade of microorganisms. The articles included in Special Issue of Mycology originate from different parts of the world: India, China, South‐America and Europe. In Asian countries the emphasis is, not only deduced from the present articles, on identifying fungal species, the products of which (enzymes or metabolites) could and will have practical use. This makes it sometimes difficult to deduce whether the article would fit better into a biotechnological journal. However, fungi have been and will be an extensive source of new products. Therefore, we have and will welcome this type of articles considered as a first step in a process that might lead to large scale applications for a fungal product. The basic sciences, investigating regulation of fungal physiology and morphology, are being at the heart not only of the Special Issue “Basic Mycology”, but generally for the Journal of Basic Microbiology. The topic is becoming better understood by the help of numerous sequenced fungal genomes, and the editors would welcome more basic research on fungal microorganisms. (© 2010 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim)

ABSTRACT The genome sequences of the basidiomycete Agaricomycetes species Coprinopsis cinerea, Laccaria bicolor, Schizophyllum commune, Phanerochaete chrysosporium, and Postia placenta, as well as of Cryptococcus neoformans and Ustilago maydis, are now publicly available. Out of these fungi, C. cinerea, S. commune, and U. maydis, together with the budding yeast Saccharomyces cerevisiae, have been investigated for years genetically and molecularly for signaling in sexual reproduction. The comparison of the structure and organization of mating type genes in fungal genomes reveals an amazing conservation of genes regulating the sexual reproduction throughout the fungal kingdom. In agaricomycetes, two mating type loci, A, coding for homeodomain type transcription factors, and B, encoding a pheromone/receptor system, regulate the four typical mating interactions of tetrapolar species. Evidence for both A and B mating type genes can also be identified in basidiomycetes with bipolar systems, where only two mating interactions are seen. In some of these fungi, the B locus has lost its self/nonself discrimination ability and thus its specificity while retaining the other regulatory functions in development. In silico analyses now also permit the identification of putative components of the pheromone-dependent signaling pathways. Induction of these signaling cascades leads to development of dikaryotic mycelia, fruiting body formation, and meiotic spore production. In pheromone-dependent signaling, the role of heterotrimeric G proteins, components of a mitogen-activated protein kinase (MAPK) cascade, and cyclic AMP-dependent pathways can now be defined. Additionally, the pheromone-dependent signaling through monomeric, small GTPases potentially involved in creating the polarized cytoskeleton for reciprocal nuclear exchange and migration during mating is predicted.

The genome sequences of the basidiomycete Agaricomycetes species Coprinopsis cinerea, Laccaria bicolor, Schizophyllum commune, Phanerochaete chrysosporium, and Postia placenta, as well as of Cryptococcus neoformans and Ustilago maydis, are now publicly available. Out of these fungi, C. cinerea, S. commune, and U. maydis, together with the budding yeast Saccharomyces cerevisiae, have been investigated for years genetically and molecularly for signaling in sexual reproduction. The comparison of the structure and organization of mating type genes in fungal genomes reveals an amazing conservation of genes regulating the sexual reproduction throughout the fungal kingdom. In agaricomycetes, two mating type loci, A, coding for homeodomain type transcription factors, and B, encoding a pheromone/ receptor system, regulate the four typical mating interactions of tetrapolar species. Evidence for both A and B mating type genes can also be identified in basidio-mycetes with bipolar systems, where only two mating interactions are seen. In some of these fungi, the B locus has lost its self/nonself discrimination ability and thus its specificity while retaining the other regulatory functions in development. In silico analyses now also permit the identification of putative components of the pheromone-dependent signaling pathways. Induction of these signaling cascades leads to development of dikaryotic mycelia, fruiting body formation, and meiotic spore production. In pheromone-dependent signaling, the role of heterotrimeric G proteins, components of a mitogen-activated protein kinase ( MAPK) cascade, and cyclic AMP-dependent pathways can now be defined. Additionally, the pheromone-dependent signaling through monomeric, small GTPases potentially involved in creating the polarized cytoskeleton for reciprocal nuclear exchange and migration during mating is predicted.

A new type of secondary mutation at the Bβ incompatibility locus of Schizophyllum commune is reported. The secondary mutation Bβ7(1–4) was induced by X-rays from the primary mutant allele Bβ7(1). The mutant Bβ7(1–4) allele showed an exceptional pattern of interactions with wild Bβ alleles, which normally behave as though they are completely equivalent and uniform in their mating interactions with other wild alleles and with mutant alleles of the locus. The new Bβ mutation is compatible with three wild Bβ alleles and incompatible with four wild Bβ alleles.

Conifers like Scots pine (Pinus sylvestris) have a complicated root system consisting of morphologically and anatomically different root types, of which the short roots have a very limited ability to elongate. Short roots have an important role in nature since they are able to establish ectomycorrhizal symbiosis, in which the growth of fungal mycelium between the epidermal cells and in the intercellular space between cortical cells leads to formation of dichotomous short roots, which may, through further splitting of the meristem, form coralloid root structures. Dichotomous short roots have been suggested to result from changes in either auxin or ethylene concentrations due to the fungal growth inside the root. NPA, the inhibitor of polar auxin transport, enhances the dichotomization of P. sylvestris short root tips similarly to the fungal growth in the root, thus confirming that auxin plays a role in short root morphogenesis. Ethylene is also known to have an important role in the regulation of root morphogenesis. In future the research dealing with the root system and ectomycorrhiza development in P. sylvestris must take into account that both auxin and ethylene are involved and that there is no contradiction in obtaining the same phenotype with both hormones. The expression analysis of PIN proteins, auxin efflux carriers, could give valuable information about the role of auxin transport in regulating the root growth and morphogenesis of coniferous root system and mycorrhiza.

The SCARECROW (SCR) gene is central to root radial patterning. Its expression has not been investigated in conifers with morphologically different root types. Additional interest in SCR functions in the Pinus sylvestris root system comes from the effect of ectomycorrhiza formation on the short root apical structure. Here, the P. sylvestris SCR gene (PsySCR) was cloned and its expression investigated by northern blot and in situ hybridization of primary, lateral and short roots and mycorrhiza. Short root dichotomization was induced by auxin transport inhibitor (N-1-naphthylphthalamic acid (NPA)). PsySCR has conserved GRAS family protein motifs at the C-terminus and a variable N-terminus. PsySCR expression occurred in young root tissue and mycorrhiza. In root sections the PsySCR signal runs through the tip in initials for stele and root cap column and becomes upwards-restricted to endodermis in all root types. The PsySCR expression pattern suggests for the first time a regulatory role for SCR in maintaining the endodermal characteristics and radial patterning of roots with open meristem organization. The specific PsySCR localization is also an excellent marker for investigation of the dichotomization process in short roots.