Biology subjects

Liu, X.-B.

Publications and source records attributed to Liu, X.-B..

7 recordsLinked to original sources

bioRxiv2026

Functionally dark genes and the transcriptomic landscape of sporulation in a model mushroom-forming fungus

Spores are the primary means of fungal reproduction, contributing to genetic diversity, colonization, and adaptation. Although spore formation is a pivotal part of the fungal life cycle, its genetic underpinnings remain poorly known. In this study, we characterize transcriptomic changes from late meiosis to early basidiospore formation in the mushroom-forming fungus Coprinopsis cinerea, decipher several cellular processes, and identify novel genes involved in this process. We identify distinct trajectories of gene expression, each of which display different functional signals, corresponding to meiotic and morphogenetic processes and transitions between these. Our analyses identify diverse arrays of fungal cell wall modifying carbohydrate-active enzymes, ferritins, a putative catechol-melanin synthesis pathway, as well as components of the mitotic/meiotic apparatus. We present twelve highly conserved genes with roles specific to sexual sporulation in both budding yeast and C. cinerea, indicating deep conservation of the gene networks driving sexual spore formation. Reverse genetics identified three conserved but functionally poorly characterized genes conferring sporeless and spore-poor phenotypes that result from postmeiotic developmental arrests stemming from spore inflation and nuclear migration problems. Overall, this study provides novel insight into basidiomycete spore formation and highlights the cornucopia of novel functions encoded by functionally dark genes.

microbiology↗

bioRxiv2025

Functional characterization of a Con7-related transcription factor in Coprinopsis cinerea indicates evolutionary conservation of morphogenetic roles

Fruiting bodies of mushroom-forming fungi (Agaricomycetes) exhibit the highest degree of multicellular complexity in fungi, yet the molecular underpinnings of their developmental programs remain incompletely understood. Here, we characterize gcd1, a gene encoding a transcription factor in the Con7 subfamily of C2H2-type zinc finger proteins. This subfamily has previously been implicated in pathogenic morphogenesis in Ascomycota, but their role in Agaricomycetes has not previously been addressed. In Coprinopsis cinerea, CRISPR/Cas9-mediated deletion of gcd1 resulted in strains with severely impaired fruiting body morphogenesis, with malformed cap, stipe, and gill tissues. Gcd1 deletion strains lacked universal veil, resembling species with open (gymnocarpous) development. We find that GCD1/Con7 homologs are widely distributed in most Dikarya species and are mostly encoded by a single gene in each species genome. Transcriptome analyses identified several misregulated genes in the {Delta}gcd1 mutant, which pinpoint potential mechanisms underlying its developmental defects as well as provided insights into the morphogenesis of mushroom fruiting bodies. These findings establish GCD1 as a key regulator of multicellular development in C. cinerea and broaden the known functions of Con7-like transcription factors to include fruiting body morphogenesis in Agaricomycetes. Overall, our results and the morphogenetic role of Con7-like transcription factors of Ascomycota suggest functional conservation over half a billion years of evolution.

microbiology↗

bioRxiv2024

A new regulator of sporulation sheds light on spore morphogenesis and ballistospory in mushroom-forming fungi

Sporulation is the most widespread means of reproduction and dispersal in fungi. In the Basidiomycota, sexual spores are produced on specialised cells known as basidia, from which they are discharged forcibly by a powered process called ballistospory, the highest known acceleration in nature. However, the genetics of sporulation, in particular postmeiotic events related to spore morphogenesis and ballistospory, remain poorly known. Here, we explore the genetics of these processes, based on a new, highly conserved transcription factor, Sporulation-Related Regulator 1 (SRR1), and its putative downstream regulatory network. Reverse genetics of Srr1 in the model mushroom Coprinopsis cinerea and commercially produced oyster mushroom indicated a conserved role of Srr1 in sporulation across Agaricomycetes. RNA-Seq analysis and motif-based inference of a hypothetical SRR1 gene regulatory network allowed delimiting putative targets regulated by SRR1 in a direct and indirect manner. Using this network and comparative genomics, we identified genes associated with ballistospory, including a putative SRR1-target chitinase, which was found to be required for normal spore production and morphology. Overall, our study offers new insights into the genetic mechanisms governing postmeiotic spore morphogenesis and ballistospory in the Agaricomycetes.

microbiology↗

bioRxiv2024

Unraveling Morphogenesis, Starvation, and Light Responses in a Mushroom-Forming Fungus, Coprinopsis cinerea, Using Long Read Sequencing and Extensive Expression Profiling

Mushroom-forming fungi (Agaricomycetes) are emerging as pivotal players in several fields, as drivers of nutrient cycling, sources of novel applications, and the group includes some of the most morphologically complex multicellular fungi. Genomic data for Agaricomycetes are accumulating at a steady pace, however, this is not paralleled by improvements in the quality of genome sequence and associated functional gene annotations, which leaves gene function notoriously poorly understood in comparison with other fungi and model eukaryotes. We set out to improve our functional understanding of the model mushroom Coprinopsis cinerea by integrating a new, chromosome-level assembly with high-quality gene predictions and functional information derived from gene-expression profiling data across 67 developmental, stress, and light conditions. The new annotation has considerably improved quality metrics and includes 5- and 3-untranslated regions (UTRs), polyadenylation sites (PAS), upstream ORFs (uORFs), splicing isoforms, conserved sequence motifs (e.g., TATA and Kozak boxes) and microexons. We found that alternative polyadenylation is widespread in C. cinerea, but that it is not specifically regulated across the various conditions used here. Transcriptome profiling allowed us to delineate core gene sets corresponding to carbon starvation, light-response, and hyphal differentiation, and uncover new aspects of the light-regulated phases of life cycle. As a result, the genome of C. cinerea has now become the most comprehensively annotated genome among mushroom-forming fungi, which will contribute to multiple rapidly expanding fields, including research on their life history, light and stress responses, as well as multicellular development.

genomics↗

bioRxiv2023

Snowball: a novel gene family required for developmental patterning in fruiting bodies of mushroom-forming fungi (Agaricomycetes)

The morphogenesis of sexual fruiting bodies of fungi is a complex process determined by a genetically encoded program. Fruiting bodies reached the highest complexity levels in the Agaricomycetes, yet, the underlying genetics is currently poorly known. In this work, we functionally characterized a highly conserved unannotated gene termed snb1, whose expression level increases rapidly during fruiting body initiation. According to phylogenetic analyses, orthologues of snb1 are present in almost all agaricomycetes and may represent a novel conserved gene family that plays a substantial role in fruiting body development. We disrupted snb1 using CRISPR/Cas9 in the agaricomycete model organism Coprinopsis cinerea. Snb1 deletion mutants formed unique, snowball-shaped, rudimentary fruiting bodies that could not differentiate caps, stipes and lamellae. We took advantage of this phenotype to study fruiting body differentiation using RNA-Seq analyses. This revealed differentially regulated genes and gene families that, based on wild-type RNA-Seq data, were upregulated early during development and showed tissue-specific expression, underscoring their potential role in differentiation. Taken together, the novel gene family of snb1 and the differentially expressed genes in the snb1 mutants provide valuable insights into the complex mechanisms underlying developmental patterning in the Agaricomycetes. ImportanceFruiting bodies of mushroom-forming fungi (Agaricomycetes) are complex multicellular structures, with a spatially and temporally integrated developmental program that is, however, currently poorly known. In this study we present a novel, conserved gene family, Snowball (snb), termed after the unique, differentiation-less fruiting body morphology of snb1 knockout strains in the model mushroom Coprinopsis cinerea. Snb is a hitherto unannotated gene that is highly conserved among agaricomycetes and encodes a protein of unknown function. A comparative transcriptomic analysis of the early developmental stages of differentiated wild-type and non-differentiated mutant fruiting bodies revealed conserved differentially expressed genes which may be related to tissue differentiation and developmental patterning fruiting body development.

developmental biology↗

bioRxiv2022

Taxonomic vs genomic fungi: contrasting evolutionary loss of protistan genomic heritage and emergence of fungal novelties

Fungi are among the most ecologically important heterotrophs that have radiated into most niches on Earth and fulfil key ecological services. However, despite intense interest in their origins, major genomic trends characterising the evolutionary route from a unicellular opisthokont ancestor to derived multicellular fungi remain poorly known. Here, we reconstructed gene family evolution across 123 genomes of fungi and relatives and show that a dominant trend in early fungal evolution has been the gradual shedding of protist genes and highly episodic innovation via gene duplication. We find that the gene content of early-diverging fungi is protist-like in many respects, owing to the conservation of protist genes in early fungi. While gene loss has been constant and gradual during early fungal evolution, our reconstructions show that gene innovation showed two peaks. Gene groups with the largest contribution to genomic change included extracellular proteins, transcription factors, as well as ones linked to the coordination of nutrient uptake with growth, highlighting the transition to a sessile osmotrophic feeding strategy and subsequent lifestyle evolution as important elements of early fungal evolution. Taken together, this work provided a highly resolved genome-wide catalogue of gene family changes across fungal evolution. This suggests that the genome of pre-fungal ancestors may have been transformed into the archetypal fungal genome by a combination of gradual gene loss, turnover and two large duplication events rather than by abrupt changes, and consequently, that the taxonomically defined fungal kingdom does not represent a genomically uniform assemblage of extant species characterized by diagnostic synapomorphies.

evolutionary biology↗

bioRxiv2022

Pre-assembled Cas9 ribonucleoprotein-mediated gene deletion identifies the carbon catabolite repressor and its target genes in Coprinopsis cinerea

Cre1 is an important transcription factor that regulates carbon catabolite repression (CCR) and is widely conserved across fungi. This gene has been extensively studied in several Ascomycota species, whereas its role in gene expression regulation in the Basidiomycota remains poorly understood. Here, we identified and investigated the role of cre1 in Coprinopsis cinerea, a basidiomycete model mushroom that can efficiently degrade lignocellulosic plant wastes. We used a rapid and efficient gene deletion approach based on PCR-amplified split-marker DNA cassettes together with in-vitro assembled Cas9-guide RNA ribonucleoproteins (Cas9-RNPs) to generate C. cinerea cre1 gene deletion strains. Gene expression profiling of two independent C. cinerea cre1 mutants showed significant deregulation of carbohydrate metabolism, plant cell wall degrading enzymes (PCWDEs), plasma membrane transporter-related and several transcription factor encoding genes, among others. Our results support the notion that, similarly to reports in the ascomycetes, Cre1 of C. cinerea orchestrates CCR through a combined regulation of diverse genes, including PCWDEs, transcription factors that positively regulate PCWDEs and membrane transporters which could import simple sugars that can induce the expression of PWCDEs. Somewhat paradoxically, though in accordance with other Agaricomycetes, genes related to lignin degradation were mostly downregulated in cre1 mutants, indicating they fall under different regulation than other PCWDEs. The gene deletion approach and the data presented in this paper expand our knowledge of CCR in the Basidiomycota and provide functional hypotheses on genes related to plant biomass degradation. ImportanceMushroom-forming fungi include some of the most efficient degraders of lignocellulosic plant biomass. They degrade dead plant materials by a battery of lignin-, cellulose-, hemicellulose- and pectin-degrading enzymes, the encoding genes of which are under tight transcriptional control. One of the highest-level regulation of these metabolic enzymes is known as carbon catabolite repression, which is orchestrates by the transcription factor Cre1, and ensures that costly lignocellulose-degrading enzyme genes are expressed only when simple carbon sources (e.g. glucose) are not available. Here, we identified the Cre1 ortholog in a litter-decomposer Agaricomycete, Coprinopsis cinerea, knocked it out and characterized transcriptional changes in the mutants. We identified several dozen lignocellulolytic enzyme genes as well as membrane transporters and other transcription factors as putative target genes. These results extend knowledge on carbon catabolite repression to litter decomposer Basidiomycota.

microbiology↗