Fungal Development Research Articles

STRIPAK, a fundamental signaling hub of eukaryotic development.

SUMMARYThe striatin-interacting phosphatase and kinase (STRIPAK) complex is involved in the regulation of many developmental processes in eukaryotic microorganisms and all animals, including humans. STRIPAK is a component of protein phosphatase 2A (PP2A), a highly conserved serine-threonine phosphatase composed of catalytic subunits (PP2Ac), a scaffolding subunit (PP2AA) and various substrate-directing B regulatory subunits. In particular, the B''' regulatory subunit called striatin has evoked major interest over the last 20 years. Studies in fungal systems have contributed substantially to our current knowledge about STRIPAK composition, assembly, and cellular localization, as well as its regulatory role in autophagy and the morphology of fungal development. STRIPAK represents a signaling hub with many kinases and thus integrates upstream and downstream information from many conserved eukaryotic signaling pathways. A profound understanding of STRIPAK's regulatory role in fungi opens the gateway to understanding the multifarious functions carried out by STRIPAK in higher eukaryotes, including its contribution to malignant cell growth.

Genome-Wide Identification and Analysis of Gene Family of Carbohydrate-Binding Modules in Ustilago crameri.

Ustilago crameri is a pathogenic basidiomycete fungus that causes foxtail millet kernel smut (FMKS), a devastating grain disease in most foxtail millet growing regions of the world. Carbohydrate-Binding Modules (CBMs) are one of the important families of carbohydrate-active enzymes (CAZymes) in fungi and play a crucial role in fungal growth and development, as well as in pathogen infection. However, there is little information about the CBM family in U. crameri. Here, 11 CBM members were identified based on complete sequence analysis and functional annotation of the genome of U. crameri. According to phylogenetic analysis, they were divided into six groups. Gene structure and sequence composition analysis showed that these 11 UcCBM genes exhibit differences in gene structure and protein motifs. Furthermore, several cis-regulatory elements involved in plant hormones were detected in the promoter regions of these UcCBM genes. Gene ontology (GO) enrichment and protein-protein interaction (PPI) analysis showed that UcCBM proteins were involved in carbohydrate metabolism, and multiple partner protein interactions with UcCBM were also detected. The expression of UcCBM genes during U. crameri infection is further clarified, and the results indicate that several UcCBM genes were induced by U. crameri infection. These results provide valuable information for elucidating the features of U. crameri CBMs' family proteins and lay a crucial foundation for further research into their roles in interactions between U. crameri and foxtail millet.

The GCN4 Transcription Factor: A Review of Its Functional Progress in Fungi

Nitrogen serves as a pivotal nutrient for the proliferation, maturation, and pathogenicity of fungi. Despite its importance, nitrogen starvation is a common challenge encountered during fungal development and host invasion. A key regulatory transcription factor, known as general control non-derepressible 4 (GCN4), has been characterized in various fungal groups, including model fungal, pathogens, and basidiomycetes. This factor is triggered by nitrogen limitation and subsequently stimulates the expression of a multitude of genes involved in amino acid synthesis, thereby countering the effects of nitrogen deficiency. This paper provides a comprehensive review on the activation mechanisms, the structural characteristics and stability of GCN4, and how GCN4 activates its downstream target genes to regulate the physiological processes of fungi. This study lays the theoretical groundwork for future research endeavors that seek to enhance nitrogen utilization, preserve the delicate balance of carbon–nitrogen metabolism, and stimulate growth, development, and secondary metabolism in fungi, especially under nitrogen-limited conditions.

Multifaceted roles of NADPH oxidases in the growth and pathogenicity of Beauveria bassiana

ABSTRACT Reactive oxygen species (ROS), synthesized by the nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (Nox) complex, are vital molecules in biological cells, influencing various physiological processes such as fungal growth, development, and virulence. Beauveria bassiana, an entomopathogenic fungus, is a promising biopesticide for agricultural, forestry, and urban pest control. This study focuses on the characterization of NADPH oxidases (Noxs) in B. bassiana. Gene expression profiles of Noxs in B. bassiana (BbNoxs) were analysed using RT-qPCR. Knockout strains of single BbNoxA, BbNoxB, BbNoxR, and double BbNoxA and BbNoxB were constructed via homologous recombination, and their phenotypic characteristics were examined. Fungal virulence was evaluated using Galleria mellonella larvae, and infection structures formation and penetration ability were assessed on cicada wings. ROS production and actin assembly during fungal growth and infection were detected using staining and marker methods. Expression analysis revealed significant upregulation of BbNoxs during fungal growth and infection. Compared to the wild-type strain, single knockouts (ΔBbNoxA/B/R) and double knockout (ΔBbNoxAB) of BbNoxs exhibited reduced conidial yields, accelerated conidial germination rates. Deletion of BbNoxB or BbNoxR decreased fungal virulence compared to the WT strain in topical inoculation experiments. Additionally, loss of BbNoxB or BbNoxR impaired infection structures formation, penetration ability, ROS production, and actin aggregation during fungal infection. BbNoxs are crucial for fungal growth, development, and virulence in B. bassiana, playing essential roles in infection structures formation, penetration, ROS production, and actin assembly. Understanding their functions provides insights into B. bassiana’s pathogenic mechanisms.

Functional characterization of extracellular and intracellular catalase-peroxidases involved in virulence of the fungal wheat pathogen Zymoseptoria tritici.

Understanding how pathogens defend themselves against host defence mechanisms, such as hydrogen peroxide (H2O2) production, is crucial for comprehending fungal infections. H2O2 poses a significant threat to invading fungi due to its potent oxidizing properties. Our research focuses on the hemibiotrophic fungal wheat pathogen Zymoseptoria tritici, enabling us to investigate host-pathogen interactions. We examined two catalase-peroxidase (CP) genes, ZtCpx1 and ZtCpx2, to elucidate how Z. tritici deals with host-generated H2O2 during infection. Our analysis revealed that ZtCpx1 was up-regulated during biotrophic growth and asexual spore formation invitro, while ZtCpx2 showed increased expression during the transition from biotrophic to necrotrophic growth and in-vitro vegetative growth. Deleting ZtCpx1 increased the mutant's sensitivity to exogenously added H2O2 and significantly reduced virulence, as evidenced by decreased Septoria tritici blotch symptom severity and fungal biomass production. Reintroducing the wild-type ZtCpx1 allele with its native promoter into the mutant strain restored the observed phenotypes. While ZtCpx2 was not essential for full virulence, the ZtCpx2 mutants exhibited reduced fungal biomass development during the transition from biotrophic to necrotrophic growth. Moreover, both CP genes act synergistically, as the double knock-out mutant displayed a more pronounced reduced virulence compared to ΔZtCpx1. Microscopic analysis using fluorescent proteins revealed that ZtCpx1 was localized in the peroxisome, indicating its potential role in managing host-generated reactive oxygen species during infection. In conclusion, our research sheds light on the crucial roles of CP genes ZtCpx1 and ZtCpx2 in the defence mechanism of Z. tritici against host-generated hydrogen peroxide.

Functional Verification of Transcription Factor comp54181_c0 in Monascus purpureus.

Monacolin K is a valuable secondary metabolite produced after a period of fermentation by Monascus purpureus; however, our current understanding of the regulatory mechanisms of its synthesis remains incomplete. This study conducted functional analysis on the key transcription factor, comp54181_c0, that is involved in the synthesis of monacolin K in Monascus. Mutant strains with either knockout or overexpression of comp54181_c0 were constructed using CRISPR/Cas9. A comparison between the knockout and overexpression strains revealed changes in fungal morphology and growth, with a significant increase in the production of Monascus pigments and monacolin K when comp54181_c0 was absent. Real-time fluorescence quantitative PCR analysis revealed that comp54181_c0 significantly influenced the transcription of key genes related to monacolin K biosynthesis in Monascus. In conclusion, our study elucidates the crucial role of comp54181_c0 in Monascus, enriches our understanding of fungal secondary metabolite development and regulation, and provides a foundation for the development and regulation of Monascus and monacolin K production.

MoHG1 Regulates Fungal Development and Virulence in Magnaporthe oryzae

Magnaporthe oryzae causes rice blast disease, which threatens global rice production. The interaction between M. oryzae and rice is regarded as a classic model for studying the relationship between the pathogen and the host. In this study, we found a gene, MoHG1, regulating fungal development and virulence in M. oryzae. The ∆Mohg1 mutants showed more sensitivity to cell wall integrity stressors and their cell wall is more easily degraded by enzymes. Moreover, a decreased content of chitin but higher contents of arabinose, sorbitol, lactose, rhamnose, and xylitol were found in the ∆Mohg1 mutant. Combined with transcriptomic results, many genes in MAPK and sugar metabolism pathways are significantly regulated in the ∆Mohg1 mutant. A hexokinase gene, MGG_00623 was downregulated in ∆Mohg1, according to transcriptome results. We overexpressed MGG_00623 in a ∆Mohg1 mutant. The results showed that fungal growth and chitin contents in MGG_00623-overexpressing strains were restored significantly compared to the ∆Mohg1 mutant. Furthermore, MoHG1 could interact with MGG_00623 directly through the yeast two-hybrid and BiFC. Overall, these results suggest that MoHG1 coordinating with hexokinase regulates fungal development and virulence by affecting chitin contents and cell wall integrity in M. oryzae, which provides a reference for studying the functions of MoHG1-like genes.

The divergence of DHN-derived melanin pathways in Metarhizium robertsii.

The important role of dihydroxynaphthalene-(DHN) melanin in enhancing fungal stress resistance and its importance in fungal development and pathogenicity are well-established. This melanin also aids biocontrol fungi in surviving in the environment and effectively infecting insects. However, the biosynthetic origin of melanin in the biocontrol agents, Metarhizium spp., has remained elusive due to the complexity resulting from the divergence of two DHN-like biosynthetic pathways. Through the heterologous expression of biosynthetic enzymes from these two pathways in baker's yeast Saccharomyces cerevisiae, we have confirmed the presence of DHN biosynthesis in M. roberstii, and discovered a novel naphthopyrone intermediate, 8, that can produce a different type of pigment. These two pigment biosynthetic pathways differ in terms of polyketide intermediate structures and subsequent modification steps. Stress resistance studies using recombinant yeast cells have demonstrated that both DHN and its intermediates confer resistance against UV light prior to polymerization; a similar result was observed for its naphthopyrone counterpart. This study contributes to the understanding of the intricate and diverse biosynthetic mechanisms of fungal melanin and has the potential to enhance the application efficiency of biocontrol fungi such as Metarhizium spp. in agriculture.

The yeast Dothiora sorbi IOJ-3 naturally produced various filamentous sectors with distinct abilities by undergoing DNA demethylation

Some fungi have demonstrated the ability to adapt rapidly to changing environments by exhibiting morphological plasticity, a trait influenced by species and environmental factors. Here, an anamorphic yeast strain IOJ-3 exhibited unique sectorization characteristics, naturally producing diverse filamentous sectors when cultivated on potato dextrose agar (PDA) medium or natural culture medium for durations exceeding 13 days. The strain IOJ-3 and its filamentous sectors were identified as Dothiora sorbi. The morphology of the sectors was consistent and heritable. The life cycle of strain IOJ-3 was investigated through microscopic observation, emphasizing the development of conidiogenous cells as a crucial stage, from which filamentous sectors originate. Some physiological characteristics of IOJ-3 and filamentous sectors are compared, and strain IOJ-3 has a higher antibiotic tolerance than two filamentous sectors, IOJ-3a expands faster on the culture medium, and IOJ-3b can penetrate cellophane. A transcriptomic analysis was conducted to investigate the differentially expressed genes between the yeast form IOJ-3 and its two filamentous sectors, revealing a total of 594 genes that exhibited consistent differential expression relative to IOJ-3, including 44 silencing genes in IOJ-3 that were activated. Gene Ontology analysis indicated that these differentially expressed genes were primarily associated with the cellular component category. Furthermore, adding 5-Azacytidine accelerated filamentous sectorization and increased the proportion of filamentous cells of strain IOJ-3 in PD liquid media, suggesting that the filamentous sectorization observed in strain IOJ-3 is linked to processes of DNA demethylation. In conclusion, this study sheds light on the biological characteristics of D. sorbi regarding morphological transitions and provides substantial direction for exploring genes related to fungal filamentous development.

Zinc finger protein LjRSDL regulates arbuscule degeneration of arbuscular mycorrhizal fungi in Lotus japonicus.

In arbuscular mycorrhizal (AM) symbiosis, appropriate regulation of the formation, maintenance, and degeneration of the arbuscule are essential for plants and fungi. In this study, we identified a Cysteine-2/Histidine-2 zinc finger protein (C2H2-ZFP)-encoding gene in Lotus japonicus named Regulator of Symbiosome Differentiation-Like (LjRSDL) that is required for arbuscule degeneration. Evolutionary analysis showed that homologs of LjRSDL exist in mycorrhizal flowering plants. We obtained ProLjRSDL::GUS transgenic hairy roots and showed that LjRSDL was strongly upregulated upon AM colonization, particularly at 18 days post AM fungi inoculation and specifically expressed in arbuscular-containing cells. The mycorrhization rate increased in the ljrsdl mutant but decreased in LjRSDL overexpressed L. japonicus. Interestingly, we observed higher proportions of large arbuscule in the ljrsdl mutant but lower proportions of larger arbuscule in LjRSDL overexpressing plants. Transcriptome analyses indicated that genes involved in arbuscule degeneration were significantly changed upon the dysregulation of LjRSDL and that LjRSDL-dependent regulation in AM symbiosis is mainly via the hormone signal transduction pathway. LjRSDL, therefore, represents a C2H2-ZFP that negatively regulates AM symbiosis. Our study provides insight into understanding plant-AM fungal communication and AM symbiosis development.

Transcriptome Analysis Reveals the Effect of Oyster Mushroom Spherical Virus Infection in Pleurotus ostreatus.

Oyster mushroom spherical virus (OMSV) is a mycovirus that inhibits mycelial growth, induces malformation symptoms, and decreases the yield of fruiting bodies in Pleurotus ostreatus. However, the pathogenic mechanism of OMSV infection in P. ostreatus is poorly understood. In this study, RNA sequencing (RNA-seq) was conducted, identifying 354 differentially expressed genes (DEGs) in the mycelium of P. ostreatus during OMSV infection. Verifying the RNA-seq data through quantitative real-time polymerase chain reaction on 15 DEGs confirmed the consistency of gene expression trends. Both Gene Ontology and Kyoto Encyclopedia of Genes and Genomes analyses highlighted the pivotal role of primary metabolic pathways in OMSV infection. Additionally, significant changes were noted in the gene expression levels of carbohydrate-active enzymes (CAZymes), which are crucial for providing the carbohydrates needed for fungal growth, development, and reproduction by degrading renewable lignocellulose. The activities of carboxymethyl cellulase, laccase, and amylase decreased, whereas chitinase activity increased, suggesting a potential mechanism by which OMSV influenced mycelial growth through modulating CAZyme activities. Therefore, this study provided insights into the pathogenic mechanisms triggered by OMSV in P. ostreatus.

The Cryptochrome CryA Regulates Lipid Droplet Accumulation, Conidiation, and Trap Formation via Responses to Light in Arthrobotrys oligospora.

Light is a key environmental factor affecting conidiation in filamentous fungi. The cryptochrome/photolyase CryA, a blue-light receptor, is involved in fungal development. In the present study, a homologous CryA (AoCryA) was identified from the widely occurring nematode-trapping (NT) fungus Arthrobotrys oligospora, and its roles in the mycelial growth and development of A. oligospora were characterized using gene knockout, phenotypic comparison, staining technique, and metabolome analysis. The inactivation of AocryA caused a substantial decrease in spore yields in dark conditions but did not affect spore yields in the wild-type (WT) and ∆AocryA mutant strains in light conditions. Corresponding to the decrease in spore production, the transcription of sporulation-related genes was also significantly downregulated in dark conditions. Contrarily, the ∆AocryA mutants showed a substantial increase in trap formation in dark conditions, while the trap production and nematode-trapping abilities of the WT and mutant strains significantly decreased in light conditions. In addition, lipid droplet accumulation increased in the ∆AocryA mutant in dark conditions, and the mutants showed an increased tolerance to sorbitol, while light contributed to the synthesis of carotenoids. Finally, AoCryA was found to affect secondary metabolic processes. These results reveal, for the first time, the function of a homologous cryptochrome in NT fungi.

Phenology, voltinism, and brood development of Xylosandrus (Coleoptera: Curculionidae) ambrosia beetles in New York.

The ambrosia beetles Xylosandrus germanus (Blanford) and Xylosandrus crassiusculus (Motschulsky) are nonnative pests in orchards and nurseries in North America. They construct galleries in the sapwood of stressed woody hosts and culture a symbiotic fungus as food for their offspring. Preventing attacks is preferred but a better understanding of their biology may elucidate additional avenues for control. Recent phenological studies are lacking for X. germanus that is common in New York, and biological information on brood and gallery development is lacking for the less abundant X. crassiusculus. We conducted both outdoor rearing and laboratory studies to better understand the timing of key events in the maturation of their galleries, particularly associated with the symbiotic fungi. Two and a partial third summer generation were consistently observed over 2 yr for X. germanus; and thus, 3 flights of adult females (foundresses) occurred each summer from mid-April to late September. In both the field and laboratory, initial growth of the symbiotic fungus occurs within a few days of gallery initiation. The rapid development of the reproductive tract and oviposition by X. germanus appears to be stimulated by the presence of the fungus. Fungal, reproductive, and brood development are similar for the related X. crassiusculus in laboratory studies; the 2 species mainly appear to differ in size. The lag between beetle colonization and reproduction currently seems too brief to be exploited for management. Disruption of other stages in gallery development should also be explored to minimize the beetle damage if attacks cannot be prevented.

Transcriptomic Insights into the Physiological Aspects of the Saprotrophic Fungus Penicillium citrinum During the Spoilage of Tobacco Leaves

Penicillium citrinum is one of the most prevalent tobacco spoilage fungi. However, the mechanisms underlying fungal growth on tobacco leaves remain largely unknown. In this study, transcriptomic analyses were performed to reveal the genome-wide expression profiles of P. citrinum growing on tobacco leaves. First, a comparative analysis was conducted between two sets of transcriptomic data from P. citrinum growing on chemically defined media and tobacco leaves. Enrichment analyses showed that differentially regulated genes were mainly associated with carbohydrate degradation (e.g., cellulose, pectin, and xylan) and the catabolism of fatty acids and aromatic compounds. Comparative transcriptomic analyses between different time points indicated that the fungal transcriptome varied dynamically during the spoilage process, and the enriched terms were associated with small-molecule degradation and fungal development. Enrichment analyses indicated that more up-regulated genes appeared in all enriched Gene Ontology terms. Notably, more organelles significantly contributed to further fungal growth on tobacco leaves. In conclusion, P. citrinum activates a comprehensive transcriptome that changes dynamically when causing tobacco mildew.

Inhibitory Activity of Propolis Extracts From Stingless Bees (Meliponini) on the Development of Phytopathogenic Fungi: in Vitro Evaluation

Objective: The present study aimed to assess the alcoholic extracts of propolis from Tetragona clavipes, Scaptotrigona bipunctata, Tetragonisca angustula, and Melipona quadrifasciata. Theoretical Framework: There is growing interest in the bioactive metabolites of propolis and honey produced by different species of the group of native stingless bees (NSB) belonging to the Meliponini tribe. Method: The extracts were prepared in a proportion of 30 g of propolis to 70 mL of 96 % v/v ethanol (1:2). Afterwards, its chemical constituents were characterized by HPLC. A PDA medium was used to evaluate the antifungal activity, with the extracts added to the melting medium (55 °C) in different concentrations. The effect of increasing concentrations (1.6 %, 3.2 %, 6.4 %, and 12.0 % v/v) of extracts on the mycelial growth of the fungi Sclerotinia sclerotiorum, Fusarium sp., Colletotrichum gloeosporioides, Botrytis cinerea, and Botryosphaeria sp. was evaluated. Fungal development was determined by measuring mycelial diameter on the 14th day of inoculation. Results and Discussion: The antioxidant activity and the contents of flavonoids and phenolic compounds were identified in all samples of propolis extracts from ANSF. The T. clavipes propolis extract had the highest content of phenolic compounds (5,593 mgEAG·100 mL-1). Among the chemical compounds, naringin, gallic acid, hesperidin, and ferulic acid were identified in greater quantities. The fungi evaluated in this work were partially inhibited by most SNB alcoholic extracts, mainly at concentrations of 1.6 % and 3.2 % v/v compared to the control treatment (ethanol 96 % v/v). Changes in the hyphal network and the reduction or absence of conidia were also observed when combined with different alcoholic propolis extracts and the control with 96 % v/v ethanol. Research Implications: The results demonstrate that the chemical components of the alcoholic extracts of NSB propolis have the potential for controlling phytopathogenic fungi of agricultural interest. Originality/Value: This study contributes by assessing and highlighting the potential applications of NSB as natural products in the alternative control of phytopathogenic fungi of agricultural interest.

Involvement of LaeA and Velvet Proteins in Regulating the Production of Mycotoxins and Other Fungal Secondary Metabolites.

Fungi are rich sources of secondary metabolites of agrochemical, pharmaceutical, and food importance, such as mycotoxins, antibiotics, and antitumor agents. Secondary metabolites play vital roles in fungal pathogenesis, growth and development, oxidative status modulation, and adaptation/resistance to various environmental stresses. LaeA contains an S-adenosylmethionine binding site and displays methyltransferase activity. The members of velvet proteins include VeA, VelB, VelC, VelD and VosA for each member with a velvet domain. LaeA and velvet proteins can form multimeric complexes such as VosA-VelB and VelB-VeA-LaeA. They belong to global regulators and are mainly impacted by light. One of their most important functions is to regulate gene expressions that are responsible for secondary metabolite biosynthesis. The aim of this mini-review is to represent the newest cognition of the biosynthetic regulation of mycotoxins and other fungal secondary metabolites by LaeA and velvet proteins. In most cases, LaeA and velvet proteins positively regulate production of fungal secondary metabolites. The regulated fungal species mainly belong to the toxigenic fungi from the genera of Alternaria, Aspergillus, Botrytis, Fusarium, Magnaporthe, Monascus, and Penicillium for the production of mycotoxins. We can control secondary metabolite production to inhibit the production of harmful mycotoxins while promoting the production of useful metabolites by global regulation of LaeA and velvet proteins in fungi. Furthermore, the regulation by LaeA and velvet proteins should be a practical strategy in activating silent biosynthetic gene clusters (BGCs) in fungi to obtain previously undiscovered metabolites.

The G-protein alpha subunit AaGA1 positively regulates vegetative growth, appressorium-like formation, and pathogenicity in Alternaria alternata.

The Gα subunit is a major component of heterotrimeric G proteins, which play a crucial role in the development and pathogenicity of several model fungi. However, its detailed function in the causal agent of pear black spot (Alternaria alternata) is unclear. Our aim was to understand the characteristics and functions of AaGA1 in A. alternata. AaGA1 was cloned from A. alternata in this study, which encodes 353 amino acids and has a "G-alpha" domain. Mutant ΔAaGA1 resulted in reduced vegetative growth, conidiation, and spore germination. Especially, mutant ΔAaGA1 produced only fewer conidia on the V8A medium, and spore formation-related genes AbaA, BrlA, and WetA were significantly downregulated. More tolerance against cell wall-inhibiting agents was observed after the deletion of AaGA1. Moreover, AaGA1 deletion led to a significant reduction in melanin and toxin production. Interestingly, deletion of AaGA1 resulted in defective appressorium-like formations, complete loss of the ability to penetrate cellophane, and decreased infection on non-wound inoculated tobacco leaves. Cell wall-degrading enzyme-related genes PME, CL, Cut2, and LC were significantly downregulated in mutant ΔAaGA1 mutant, significantly reducing virulence on wound-inoculated pear fruits. The G protein alpha subunit AaGA1 is indispensable for fungal development, appressorium-like formations, and pathogenicity in A. alternata.

Characterization of PEBP-like Genes and Function of Capebp1 and Capebp5 in Fruiting Body Regeneration in Cyclocybe aegerita.

Phosphatidylethanolamine-binding proteins (PEBPs) play a crucial role in the growth and development of various organisms. Due to the low sequence similarity compared to plants, humans, and animals, the study of pebp genes in fungi has not received significant attention. The redifferentiation of fruiting bodies is exceedingly rare in fungal development. Hitherto, only a few studies have identified the Capebp2 gene as being associated with this phenomenon in Cyclocybe aegerita. Thus, exploring the role of pebp genes in fruiting body development is imperative. In the present study, four Capebp genes (Capebp1, Capebp3, Capebp4, and Capebp5) were cloned from the AC0007 strain of C. aegerita based on genome sequencing and gene prediction. The findings indicate that the pebp family, in C. aegerita, comprises a total of five genes. Moreover, the sequence similarity was low across the five CAPEBP protein sequences in C. aegerita, and only a few conserved sequences, such as HRY and RHF, were identical. Expression analyses revealed that, similarly to Capebp2, the four Capebp genes exhibit significantly higher expression levels in the fruiting bodies than in the mycelium. Furthermore, overexpressed and RNA interference Capebp1 or Capebp5 transformants were analyzed. The results demonstrate that overexpression of Capebp1 or Capebp5 could induce the regeneration of the lamella or fruiting body, whereas the knockdown of Capebp1 or Capebp5 could lead to the accelerated aging of fruiting bodies. These findings highlight a significant role of Capebp genes in the generation of C. aegerita fruiting bodies and provide a foundation for further exploration into their involvement in basidiomycete growth and development.

FgVAC1 is an Essential Gene Required for Golgi-to-Vacuole Transport and Fungal Development in Fusarium graminearum

Fusarium graminearum is an important plant pathogen that causes head blight in cereal crops such as wheat, barley, and rice worldwide. In this study, we identified and functionally characterized FgVAC1, an essential gene in F. graminearum that encodes a Rab5 effector involved in membrane tethering functions. The essentiality of FgVAC1 was confirmed through a conditional promoter replacement strategy using the zearalenone-inducible promoter (PZEAR). Cytological analyses revealed that FgVac1 colocalizes with FgRab51 on early endosomes and regulates the proper transport of the vacuolar hydrolase FgCpy1 to the vacuole. Suppression of FgVAC1 led to inhibited vegetative growth, reduced asexual and sexual reproduction, decreased deoxynivalenol (DON) biosynthesis, and diminished pathogenicity. Our findings highlight the significant role of FgVac1 in vacuolar protein sorting, fungal development, and plant infection in F. graminearum.

Glyoxal oxidase-mediated detoxification of reactive carbonyl species contributes to virulence, stress tolerance, and development in a pathogenic fungus.

Reactive carbonyl and oxygen species (RCS/ROS), often generated as metabolic byproducts, particularly under conditions of pathology, can cause direct damage to proteins, lipids, and nucleic acids. Glyoxal oxidases (Gloxs) oxidize aldehydes to carboxylic acids, generating hydrogen peroxide (H2O2). Although best characterized for their roles in lignin degradation, Glox in plant fungal pathogens are known to contribute to virulence, however, the mechanism underlying such effects are unclear. Here, we show that Glox in the insect pathogenic fungus, Metarhizium acridum, is highly expressed in mycelia and during formation of infection structures (appressoria), with the enzyme localizing to the cell membrane. MaGlox targeted gene disruption mutants showed RCS and ROS accumulation, resulting in cell toxicity, induction of apoptosis and increased autophagy, inhibiting normal fungal growth and development. The ability of the MaGlox mutant to scavenge RCS was significantly reduced, and the mutant exhibited increased susceptibility to aldehydes, oxidative and cell wall perturbing agents but not toward osmotic stress, with altered cell wall contents. The ΔMaGlox mutant was impaired in its ability to penetrate the host cuticle and evade host immune defense resulting in attenuated pathogenicity. Overexpression of MaGlox promoted fungal growth and conidial germination, increased tolerance to H2O2, but had little to other phenotypic effects. Transcriptomic analyses revealed downregulation of genes related to cell wall synthesis, conidiation, stress tolerance, and host cuticle penetration in the ΔMaGlox mutant. These findings demonstrate that MaGlox-mediated scavenging of RCS is required for virulence, and contributes to normal fungal growth and development, stress resistance.