The ich1 gene of the mushroom Coprinus cinereus is essential for pileus formation in fruiting

The formation of the pileus in homobasidiomycete fungi is essential for sexual reproduction, because the pileus bears the hymenium, a layer of cells that includes the specialised basidia in which nuclear fusion, meiosis and sporulation occur. The developmental mutant ichijiku of Coprinus cinereus fails to develop a differentiated pileus at the apex of the primordial shaft, which is the basal part of the fruit-body primordia and formed in an early stage of fruit-body differentiation. Genetic analysis indicates that this phenotype is caused by a recessive mutation in a single gene (ich1). The ich1 gene was mapped to chromosome XII using restriction fragment length polymorphism markers and the marker chromosome method, and cloned by complementation using a chromosome-XII-specific cosmid library. The ich1 gene encodes a novel protein of 1,353 amino acids. The Ich1 amino-acid sequence contains nuclear targeting signals, suggesting that the Ich1 protein would function in the nucleus. Northern blot analysis indicates that the ich1 gene is specifically expressed in the pileus of the wild-type fruit-body. No ich1 mRNA was detected in the ichijiku mutant, consistent with loss of the promoter region of ich1 in the mutant genome. These data demonstrate that the ick1 gene product is essential for pileus formation.</p

Strand-Specific RNA-Seq Analyses of Fruiting Body Development in Coprinopsis cinerea

The basidiomycete fungus Coprinopsis cinerea is an important model system for multicellular development. Fruiting bodies of C. cinerea are typical mushrooms, which can be produced synchronously on defined media in the laboratory. To investigate the transcriptome in detail during fruiting body development, high-throughput sequencing (RNA-seq) was performed using cDNA libraries strand-specifically constructed from 13 points (stages/tissues) with two biological replicates. The reads were aligned to 14,245 predicted transcripts, and counted for forward and reverse transcripts. Differentially expressed genes (DEGs) between two adjacent points and between vegetative mycelium and each point were detected by Tag Count Comparison (TCC). To validate RNA-seq data, expression levels of selected genes were compared using RPKM values in RNA-seq data and qRT-PCR data, and DEGs detected in microarray data were examined in MA plots of RNA-seq data by TCC. We discuss events deduced from GO analysis of DEGs. In addition, we uncovered both transcription factor candidates and antisense transcripts that are likely to be involved in developmental regulation for fruiting

The dst1 Gene Involved in Mushroom Photomorphogenesis of Coprinus cinereus Encodes a Putative Photoreceptor for Blue Light

The homobasidiomycete Coprinus cinereus exhibits remarkable photomorphogenesis during fruiting-body development. Under proper light conditions, fruiting-body primordia proceed to the maturation phase in which basidia in the pileus undergo meiosis, producing sexual spores, followed by stipe elongation and pileus expansion for efficient dispersal of the spores. In the continuous darkness, however, the primordia do not proceed to the maturation phase but are etiolated: the pileus and stipe tissues at the upper part of the primordium remain rudimentary and the basal part of the primordium elongates, producing “dark stipe.” In this study we genetically analyzed five strains that produce dark stipes even if light conditions promoting the maturation are given and then characterized one of them, Uar801 (dst1-1). The dst1 gene was cloned as a DNA fragment that rescues the dst1-1 mutation. Dst1 is predicted to be a protein of 1175 amino acids that contains two PAS domains, a coiled-coil structure, and a putative, glutamine-rich, transcriptional activation domain (AD). One of the PAS domains exhibits significant similarity to the LOV domains of known blue-light receptors, suggesting that Dst1 is a blue-light receptor of C. cinereus. The dst1-1 mutation is predicted to truncate the putative AD in the C-terminal region

The Coprinopsis cinerea Tup1 homologue Cag1 is required for gill formation during fruiting body morphogenesis

The pileus (cap) of the fruiting body in homobasidiomycete fungi bears the hymenium, a layer of cells that includes the basidia where nuclear fusion, meiosis and sporulation occur. Coprinopsis cinerea is a model system for studying fruiting body development. The hymenium of C. cinerea forms at the surface of the gills in the pileus. In a previous study, we identified a mutation called cap-growthless1-1 (cag1-1) that blocks gill formation, which yields primordia that never mature. In this study, we found that the cag1 gene encodes a homologue of Saccharomyces cerevisiae Tup1. The C. cinerea genome contains another Tup1 homologue gene called Cc.tupA. Reciprocal tagging of Cag1 and Cc.TupA with green and red fluorescent proteins revealed that the relative ratios of the amounts of the two Tup1 paralogues varied among tissues. Compared with Cc.TupA, Cag1 was preferentially expressed in the gill trama tissue cells, suggesting that the function of Cag1 is required for gill trama tissue differentiation and maintenance. Yeast two-hybrid analysis and co-localisation of Cag1 and Cc.TupA suggested that Cag1 interacts with Cc.TupA in the nuclei of certain cells

Live-cell imaging of septins and cell polarity proteins in the growing dikaryotic vegetative hypha of the model mushroom Coprinopsis cinerea

Abstract The developmental biology underlying the morphogenesis of mushrooms remains poorly understood despite the essential role of fungi in the terrestrial environment and global carbon cycle. The mushroom Coprinopsis cinerea is a leading model system for the molecular and cellular basis of fungal morphogenesis. The dikaryotic vegetative hyphae of this fungus grow by tip growth with clamp cell formation, conjugate nuclear division, septation, subapical peg formation, and fusion of the clamp cell to the peg. Studying these processes provides many opportunities to gain insights into fungal cell morphogenesis. Here, we report the dynamics of five septins, as well as the regulators CcCla4, CcSpa2, and F-actin, visualized by tagging with fluorescent proteins, EGFP, PA-GFP or mCherry, in the growing dikaryotic vegetative hyphae. We also observed the nuclei using tagged Sumo proteins and histone H1. The five septins colocalized at the hyphal tip in the shape of a dome with a hole (DwH). CcSpa2-EGFP signals were observed in the hole, while CcCla4 signals were observed as the fluctuating dome at the hyphal tip. Before septation, CcCla4-EGFP was also occasionally recruited transiently around the future septum site. Fluorescent protein-tagged septins and F-actin together formed a contractile ring at the septum site. These distinct specialized growth machineries at different sites of dikaryotic vegetative hyphae provide a foundation to explore the differentiation program of various types of cells required for fruiting body formation

Strand-Specific RNA-Seq Analyses of Fruiting Body Development in Coprinopsis cinerea.

The basidiomycete fungus Coprinopsis cinerea is an important model system for multicellular development. Fruiting bodies of C. cinerea are typical mushrooms, which can be produced synchronously on defined media in the laboratory. To investigate the transcriptome in detail during fruiting body development, high-throughput sequencing (RNA-seq) was performed using cDNA libraries strand-specifically constructed from 13 points (stages/tissues) with two biological replicates. The reads were aligned to 14,245 predicted transcripts, and counted for forward and reverse transcripts. Differentially expressed genes (DEGs) between two adjacent points and between vegetative mycelium and each point were detected by Tag Count Comparison (TCC). To validate RNA-seq data, expression levels of selected genes were compared using RPKM values in RNA-seq data and qRT-PCR data, and DEGs detected in microarray data were examined in MA plots of RNA-seq data by TCC. We discuss events deduced from GO analysis of DEGs. In addition, we uncovered both transcription factor candidates and antisense transcripts that are likely to be involved in developmental regulation for fruiting