A fungal sirtuin modulates development and virulence in the insect pathogen, Beauveria bassiana.

Chromatin transitions are mediated in part by acetylation/deacetylation post-translational modifications of histones. Histone deacetylases, e.g. sirtuins (Sir-proteins), repress transcription via promotion of heterochromatin formation. Here, we characterize the Sir2 class III histone deacetylase (BbSir2) in the environmentally and economically important fungal insect pathogen, Beauveria bassiana. BbSir2 is shown to contribute to the deacetylation of lysine residues on H3 and H4 histones. Targeted gene knockout of BbSir2 resulted in impaired asexual development, reduced abilities to utilize various carbon/nitrogen sources, reduced tolerance to oxidative, heat, and UV stress, and attenuated virulence. ΔBbSir2 cells showed disrupted cell cycle development and abnormal hyphal septation patterns. Proteomic protein acetylation analyses of wild type and ΔBbSir2 cells revealed the differential abundance of 462 proteins and altered (hyper- or hypo-) acetylation of 436 lysine residues on 350 proteins. Bioinformatic analyses revealed enrichment in pathways involved in carbon/nitrogen metabolism, cell cycle control and cell rescue, defence and mitochondrial functioning. Critical targets involved in virulence included LysM effector proteins and a benzoquinone oxidoreductase implicated in detoxification of cuticular compounds. These data indicate broad effects of BbSir2 on fungal development and stress response, with identification of discrete targets that can account for the observed (decreased) virulence phenotype.

Two sirtuin proteins, Hst3 and Hst4, modulate asexual development, stress tolerance, and virulence by affecting global gene expression in Beauveria bassiana.

Beauveria bassiana is a widely used entomopathogenic fungus in insect biological control applications. In this study, we investigated the role of two sirtuin homologs, BbHst3 and BbHst4, in the biological activities and pathogenicity of B. bassiana. Our results showed that deletion of BbHst3 and/or BbHst4 led to impaired sporulation, reduced (~50%) conidial production, and decreased tolerance to various stresses, including osmotic, oxidative, and cell wall-disturbing agents. Moreover, BbHst4 plays dominant roles in histone H3-K56 acetylation and DNA damage response, while BbHst3 is more responsible for maintaining cell wall integrity. Transcriptomic analyses revealed significant changes (>1,500 differentially expressed genes) in gene expression patterns in the mutant strains, particularly in genes related to secondary metabolism, detoxification, and transporters. Furthermore, the ΔBbHst3, ΔBbHst4, and ΔBbHst3ΔBbHst4 strains exhibited reduced virulence in insect bioassays, with decreased (~20%) abilities to kill insect hosts through topical application and intra-hemocoel injection. These findings highlight the crucial role of BbHst3 and BbHst4 in sporulation, DNA damage repair, cell wall integrity, and fungal infection in B. bassiana. Our study provides new insights into the regulatory mechanisms underlying the biological activities and pathogenicity of B. bassiana and emphasizes the potential of targeting sirtuins for improving the efficacy of fungal biocontrol agents.IMPORTANCESirtuins, as a class of histone deacetylases, have been shown to play important roles in various cellular processes in fungi, including asexual development, stress response, and pathogenicity. By investigating the functions of BbHst3 and BbHst4, we have uncovered their critical contributions to important phenotypes in Beauveria bassiana. Deletion of these sirtuin homologs led to reduced conidial yield, increased sensitivity to osmotic and oxidative stresses, impaired DNA damage repair processes, and decreased fungal virulence. Transcriptomic analyses showed differential expression of numerous genes involved in secondary metabolism, detoxification, transporters, and virulence-related factors, potentially uncovering new targets for manipulation and optimization of fungal biocontrol agents. Our study also emphasizes the significance of sirtuins as key regulators in fungal biology and highlights their potential as promising targets for the development of novel antifungal strategies.

Contributions of a Histone Deacetylase (SirT2/Hst2) to Beauveria bassiana Growth, Development, and Virulence.

Sirtuins are a class of histone deacetylases that promote heterochromatin formation to repress transcription. The entomopathogenic fungus Beauveria bassiana contains six sirtuin homologs. The class III histone deacetylase, BbSir2, has been previously shown to affect the regulation of carbon/nitrogen metabolism and asexual development, with only moderate effects on virulence. Here, we examine another class III histone deacetylase (BbSirT2) and show that it contributes to deacetylation of lysine residues on histone H4-K16ac. Directed gene-knockout of BbSirT2 dramatically reduced conidiation, the ability of the fungus to metabolize a range of carbon and nitrogen sources, and tolerances to oxidative, heat, and UV stress and significantly attenuated virulence in both intrahemocoel injection and topical bioassays using the Greater wax moth (Galleria mellonella) as the insect host. ΔBbSirT2 cells showed alterations in cell cycle development and hyphal septation and produced morphologically aberrant conidia. Comparative transcriptomic analyses of wild type versus ΔBbSirT2 cells indicated differential expression of 1148 genes. Differentially expressed genes were enriched in pathways involved in cell cycle and rescue, carbon/nitrogen metabolism, and pathogenesis. These included changes in the expression of polyketide synthases (PKSs) and LysM effector proteins that contribute to degradation of host toxins and target host pathways, respectively. These data indicate contributions of BbSirT2 in helping to mediate fungal stress and development, with the identification of affected gene targets that can help account for the observed reduced virulence phenotype.

The Spt10 GNAT Superfamily Protein Modulates Development, Cell Cycle Progression and Virulence in the Fungal Insect Pathogen, Beauveria bassiana.

Chromatin remodeling is mediated in part by post-translational acetylation/deacetylation modifications of histones. Histone acetyltransferases (HATs), e.g., members of the GNAT/MYST superfamily, activate gene transcription via promotion of euchromatin formation. Here, we characterized a GNAT family HAT, Spt10 (BbSpt10), in the environmentally and economically important fungal insect pathogen, Beauveria bassiana. Targeted gene knockout of BbSpt10 resulted in impaired asexual development and morphogenesis; reduced abilities to utilize various carbon/nitrogen sources; reduced tolerance to heat, fungicides, and DNA damage stress; and attenuated virulence. The ΔBbSpt10 mutant showed disrupted cell cycle development and abnormal hyphal septation patterns. Transcriptome analyses of wild type and ΔBbSpt10 cells revealed the differential expression of 373 genes, including 153 downregulated and 220 upregulated genes. Bioinformatic analyses revealed downregulated genes to be enriched in pathways involved in amino acid metabolism, cellular transportation, cell type differentiation, and virulence, while upregulated genes were enriched in carbon/nitrogen metabolism, lipid metabolism, DNA process, and cell rescue, defense, and virulence. Downregulated virulence genes included hydrophobins, cellular transporters (ABC and MFS multidrug transporters) and cytochrome P450 detoxification genes. These data indicated broad effects of BbSpt10 on fungal development, multi-stress response, and virulence.