Volvariella turcica, a new species from Turkey, and a multigene phylogeny of Volvariella.

A new species of Volvariella, collected from Aydin Province on the coast of the Aegean Sea in southwestern Turkey, is described as Volvariella turcica, sp. nov., based on morphology and multigene molecular analysis of three nuc rDNA gene regions: internal transcribed spacer ITS1-5.8S-ITS2 (ITS), 28S, and 18S. The new species was found in forests dominated by Pinus brutia and Quercus coccifera and mainly characterized by small basidiomata with a white pileus covered with pale ochre center and an ochre-discoloring volva, small basidiospores, lageniform pleurocystidia, balloon-shaped to clavate cheilocystidia, and stipitipellis hairs that are cylindrical or cylindrical-tortuous with subcapitate or lobe-like projections. A comprehensive description, illustrations, and line drawings are provided, and comparison with morphologically similar and phylogenetically related species is discussed.

Fruiting Body Formation in Volvariella volvacea Can Occur Independently of Its MAT-A-Controlled Bipolar Mating System, Enabling Homothallic and Heterothallic Life Cycles.

Volvariella volvacea is an important crop in Southeast Asia, but erratic fruiting presents a serious challenge for its production and breeding. Efforts to explain inconsistent fruiting have been complicated by the multinucleate nature, typical lack of clamp connections, and an incompletely identified sexual reproductive system. In this study, we addressed the life cycle of V. volvacea using whole genome sequencing, cloning of MAT loci, karyotyping of spores, and fruiting assays. Microscopy analysis of spores had previously indicated the possible coexistence of heterothallic and homothallic life cycles. Our analysis of the MAT loci showed that only MAT-A, and not MAT-B, controlled heterokaryotization. Thus, the heterothallic life cycle was bipolar. Karyotyping of single spore isolates (SSIs) using molecular markers supported the existence of heterokaryotic spores. However, most SSIs were clearly not heterokaryotic, yet contained structural variation (SV) markers relating to both alleles of both parents. Heterokaryons from crossed, self-sterile homokaryons could produce fruiting bodies, agreeing with bipolar heterothallism. Meanwhile, some SSIs with two different MAT-A loci also produced fruiting bodies, which supported secondary homothallism. Next, SSIs that clearly contained only one MAT-A locus (homothallism) were also able to fruit, demonstrating that self-fertile SSIs were not, per definition, secondary homothallic, and that a third life cycle or genetic mechanism must exist. Finally, recombination between SV markers was normal, yet 10 out of 24 SV markers showed 1:2 or 1:3 distributions in the spores, and large numbers of SSIs contained doubled SV markers. This indicated selfish genes, and possibly partial aneuploidy.

The Accordant Trend of Both Parameters (rgs Expression and cAMP Content) Follows the Pattern of Development of Fruiting Body in Volvariella volvacea.

The formation of fruiting body in Volvariella volvacea is affected by endogenous genes and environmental factors. However, its regulation at a molecular level is still poorly understood. To study the genes involved in the formation of fruiting body, we cloned a new regulator of the G protein signaling (RGS) encoding gene (rgs) from V. volvacea. Phylogenetic analysis showed that RGS in V. volvacea and other basidiomycete RGS proteins from Schizophyllum commune and Coprinus cinereus belong to the same clade. In addition, we assayed intracellular cAMP content in the three developmental stages (mycelium, fruiting body primordia, and button). We also found that the expression of rgs was highly positively correlated to the content of intracellular cAMP during fruiting body formation. The conserved protein sequences and expression of rgs, together with high concent of cAMP at primordia tissue, suggested that rgs gene and cAMP may play a crucial role in fruiting body formation in V. volvacea.

A specific type of cyclin-like F-box domain gene is involved in the cryogenic autolysis of Volvariella volvacea.

Cryogenic autolysis is a typical phenomenon of abnormal metabolism in Volvariella volvacea. Recent studies have identified 20 significantly up-regulated genes via high-throughput sequencing of the mRNAs expressed in the mycelia of V. volvacea after cold exposure. Among these significantly up-regulated genes, 15 annotated genes were used for functional annotation cluster analysis. Our results showed that the cyclin-like F-box domain (FBDC) formed the functional cluster with the lowest P-value. We also observed a significant expansion of FBDC families in V. volvacea. Among these, the FBDC3 family displayed the maximal gene expansion in V. volvacea. Gene expression profiling analysis revealed only one FBDC gene in V. volvacea (FBDV1) that was significantly up-regulated, which is located in the FBDC3 family. Comparative genomics analysis revealed the homologous sequences of FBDV1 with high similarity were clustered on the same scaffold. However, FBDV1 was located far from these clusters, indicating the divergence of duplicated genes. Relative time estimation and rate test provided evidence for the divergence of FBDV1 after recent duplications. Real-time RT-PCR analysis confirmed that the expression of the FBDV1 was significantly up-regulated (P < 0.001) after cold-treatment of V. volvacea for 4 h. These observations suggest that the FBDV1 is involved in the cryogenic autolysis of V. volvacea.

[Phylogenomic analysis reveals the significant expansion of gene families of Volvariella volvacea ].

[OBJECTIVE] Cryogenic autolysis of Volvariella volvacea is an unusual phenomenon of abnormal metabolism. The aim of this study was to describe this molecular feature of abnormal metabolism at the genome-level. [METHODS] We used 21 fungal species for the phylogenomic analysis and then selected 9 representative species in basidiomycetes for the comparative genomic analysis. [ RESULTS] The phylogenomic analysis shows that V. volvacea was located at the bottom of the cluster consisting of grass-degrading fungi. Phylogenetic tree shows that basidiomycetes and ascomycetes fungi have independent evolutionary trajectories. Therefore, nine representative species in basidiomycetes were chosen for the comparative genomic analysis. The result shows that compared to other grass-degrading fungi, V. volvacea has the tendency of contraction. The comparison of the number of gene families on a different scale shows that there was a significant expansion of 3 large size ( > 200) gene families (faml, fam4 and fam6) in V. volvacea with their total number significantly more than other species, representing that the molecular feature of V. volvacea is correlated with its abnormal metabolism. [ CONCLUSION] The significant expansion of 3 gene families ( > 200) in V. volvacea indicates the enhancement of their function in specific gene families, which is most likely associated with cryogenic autolysis of V. volvacea.

[Sequence characterization and differential expression of a glutathione S-transferase gene vv-gto1 from Volvariella volvacea].

OBJECTIVE: Based on the analysis of omics data of Volvariella volvacea, a gene encoding glutathione S- transferase (GSTs) named vv-gtol was obtained. To reveal the role of GSTs in the growth and development in edible fungi, the structure, the sequence characters and the expression profile of a GST gene vv-gto1 of Volvariella volvacea were analyzed. METHODS: ZOOM software was used to map sequencing read (reads) from genome and transcriptome against the splicing sequence of genome, to confirm the complete length and the accuracy of the gene sequence, and to visualize gene structure. The MEGA 5.1 was used to do the multiple sequence alignment and phylogenetic tree analysis. Real time fluorescent quantitative PCR was used to determine the expression levels of vv-gtol at different growth periods of Volvariella volvacea. RESULTS: The full sequence of vv-gtol covered 2083 bp, containing 11 exons and 10 introns, and encoded a protein with 356 amino acids. 5'UTR was 305 bp which contains one intron region, and 3'UTR was 86bp. Two intron retentions could be recognized during RNA processing, and the transcripts formed by the intron retention could not translate the correct conservative functional domains. The full-length of vv-gtol had more than 50 accurate positioning genome sequencing reads, suggesting that genome sequencing and assembly results are accurate and reliable. The phylogenetic tree showed that GTO1 of Volvariella volvacea belonged to the subclass I of the Omega class of glutathione S-transferase superfamily, and had the closest relationship with GTO1 and GTO2 in Phanerochaete chrysosporium. The analysis of digital gene expression profiling, fluorescence quantitative PCR and proteomics showed that vv-gtol had the highest expression level in the heterokaryotic hyphae. CONCLUSIONS: This is the first time to obtain a gene encoding glutathione S-transferase from Volvariella volvacea which belongs to Omega class. Our study showed that the gene may play an important role during the special biological functions of heterokaryotic hyphae. This study also suggested that Volvariella volvacea heterokaryotic hyphae in H1521 had stronger resistance ability than other samples. In addition, vv-gto1 could form different alternative splicesome to regulate gene transcription and translation, and ultimately affect the function of the protein.

Genes encoding FAD-binding proteins in Volvariella volvacea exhibit differential expression in homokaryons and heterokaryons.

Flavin adenine dinucleotide (FAD)-binding proteins play a vital role in energy transfer and utilization during fungal growth and mycelia aggregation. We sequenced the genome of Volvariella volvacea, an economically important edible fungus, and discovered 41 genes encoding FAD-binding proteins. Gene expression profiles revealed that the expression levels of four distinctly differentially expressed genes in heterokaryotic strain H1521 were higher than in homokaryotic strains PYd15 and PYd21 combined. These observations were validated by quantitative real-time PCR. The results suggest that the differential expression of FAD-binding proteins may be important in revealing the distinction between homokaryons and heterokaryons on the basis of FAD-binding protein functionality.