The transcription factor Ron1 is required for chitin metabolism, asexual development and pathogenicity in Beauveria bassiana, an entomopathogenic fungus.

Ndt80-like transcription factor Ron1 is best known for its essential role in the regulation of N-acetylglucosamine (GlcNAc) catabolism. Ron1 was again found to be essential for sensing GlcNAc in Beauveria bassiana. Importantly, our study revealed that Ron1 is involved in the metabolic processes of chitin and asexual development. To further investigate the novel functions of Ron1 in B. bassiana, extracellular chitinase activity in the ΔRon1 mutant was found to decrease by 84.73% compared with wild type. The deletion of Ron1 made it difficult for the fungus to accumulate intracellular GlcNAc. Furthermore, transcriptomic analysis revealed that Ron1 exerted a significant effect on global transcription and positively regulated genes encoding chitin metabolism in respond to chitin nutrition. Yeast one-hybrid assay confirmed that Ron1 could bind to specific cis-acting elements in the promoters of chitinase and hexokinase. In addition, ΔRon1 displayed an impaired chitin component of the cell wall, with a chitin synthetase (ChsVII) predicted to function downstream of Ron1. Finally, the virulence of ΔRon1 mutant was significantly reduced in the Galleria mellonella insect model through cuticle infection or cuticle bypassing infection. These data functionally characterize Ron1 in B. bassiana and expand our understanding of how the transcription factor Ron1 works in pathogens.

The Tudor Domain-Containing Protein BbTdp1 Contributes to Fungal Cell Development, the Cell Cycle, Virulence, and Transcriptional Regulation in the Insect Pathogenic Fungus Beauveria bassiana.

Beauveria bassiana is an insect pathogenic fungus that serves as a model system for exploring the mechanisms of fungal development and host-pathogen interactions. Clinical and experimental studies have indicated that SND1 is closely correlated with the progression and invasiveness of common cancers as a potential oncogene, but this gene has rarely been studied in fungi. Here, we characterized the contributions of an SND1 ortholog (Tdp1) by constructing a BbTdp1 deletion strain and a complemented strain of B. bassiana. Compared with the wild-type (WT) strain, the ΔBbTdp1 mutant lost conidiation capacity (∼87.7%) and blastospore (∼96.3%) yields, increased sensitivity to chemical stress (4.4 to 54.3%) and heat shock (∼44.2%), and decreased virulence following topical application (∼24.7%) and hemocoel injection (∼40.0%). Flow cytometry readings showed smaller sizes of both conidia and blastospores for ΔBbTdp1 mutants. Transcriptomic data revealed 4,094 differentially expressed genes (|log2 ratio| > 2 and a q value of <0.05) between ΔBbTdp1 mutants and the WT strain, which accounted for 41.6% of the total genes, indicating that extreme fluctuation in the global gene expression pattern had occurred. Moreover, deletion of BbTdp1 led to an abnormal cell cycle with a longer S phase and shorter G2/M and G0/G1 phases of blastospores, and enzyme-linked immunosorbent assay confirmed that the level of phosphorylated cyclin-dependent kinase 1 (Cdk1) in the ΔBbTdp1 strain was ∼31.5% lower than in the WT strain. In summary, our study is the first to report that BbTdp1 plays a vital role in regulating conidia and blastospore yields, fungal morphological changes, and pathogenicity in entomopathogenic fungi. IMPORTANCE In this study, we used Beauveria bassiana as a biological model to report the role of BbTdp1 in entomopathogenic fungi. Our findings indicated that BbTdp1 contributed significantly to cell development, the cell cycle, and virulence in B. bassiana. In addition, deletion of BbTdp1 led to drastic fluctuations in the transcriptional profile. BbTdp1 can be developed as a novel target for B. bassiana development and pathogenicity, which also provides a framework for the study of Tdp1 in other fungi.

BbWor1, a Regulator of Morphological Transition, Is Involved in Conidium-Hypha Switching, Blastospore Propagation, and Virulence in Beauveria bassiana.

Morphological transition is an important adaptive mechanism in the host invasion process. Wor1 is a conserved fungal regulatory protein that controls the phenotypic switching and pathogenicity of Candida albicans. By modulating growth conditions, we simulated three models of Beauveria bassiana morphological transitions, including CTH (conidia to hyphae), HTC (hyphae to conidia), and BTB (blastospore to blastospore). Disruption of BbWor1 (an ortholog of Wor1) resulted in a distinct reduction in the time required for conidial germination (CTH), a significant increase in hyphal growth, and a decrease in the yield of conidia (HTC), indicating that BbWor1 positively controls conidium production and negatively regulates hyphal growth in conidium-hypha switching. Moreover, ΔBbWor1 prominently decreased blastospore yield, shortened the G0/G1 phase, and prolonged the G2/M phase under the BTB model. Importantly, BbWor1 contributed to conidium-hypha switching and blastospore propagation via different genetic pathways, and yeast one-hybrid testing demonstrated the necessity of BbWor1 to control the transcription of an allergen-like protein gene (BBA_02580) and a conidial wall protein gene (BBA_09998). Moreover, the dramatically weakened virulence of ΔBbWor1 was examined by immersion and injection methods. Our findings indicate that BbWor1 is a vital participant in morphological transition and pathogenicity in entomopathogenic fungi. IMPORTANCE As a well-known entomopathogenic fungus, Beauveria bassiana has a complex life cycle and involves transformations among single-cell conidia, blastospores, and filamentous hyphae. This study provides new insight into the regulation of the fungal cell morphological transitions by simulating three models. Our research identified BbWor1 as a core transcription factor of morphological differentiation that positively regulates the production of conidia and blastospores but negatively regulates hyphal growth. More importantly, BbWor1 affects fungal pathogenicity and the global transcription profiles within three models of growth stage transformation. The present study lays a foundation for the exploration of the transition mechanism of entomopathogenic fungi and provides material for the morphological study of fungi.

Involvement of BbTpc1, an important Zn(II)2Cys6 transcriptional regulator, in chitin biosynthesis, fungal development and virulence of an insect mycopathogen.

Chitin is one of the major components of the fungal cell wall and contributes to the mechanical strength and shape of the fungal cell. Zn(II)2Cys6 transcription factors are unique to the fungal kingdom and have a variety of functions in some fungi. However, the mechanisms by which Zn(II)2Cys6 proteins affect entomopathogenic fungi are largely unknown. Here, we characterized the Zn(II)2Cys6 transcription factor BbTpc1 in the insect pathogenic fungus Beauveria bassiana. Disruption of BbTpc1 resulted in a distinct changes in vegetative growth and septation patterns, and a significant decrease in conidia and blastospore yield. The ΔBbTpc1 mutant displayed impaired resistance to chemical stresses and heat shock and attenuated virulence in topical and intrahemocoel injection assays. Importantly, the ΔBbTpc1 mutant had an abnormal cell wall with altered wall thickness and chitin synthesis, which were accompanied by transcriptional repression of the chitin synthetase family genes. In addition, comparative transcriptomics revealed that deletion of BbTpc1 altered fungal asexual reproduction via different genetic pathways. These data revealed that BbTpc1 regulates fungal development, chitin synthesis and biological control potential in B. bassiana.