Calcium Signaling Enhances Bostrycin Biosynthesis in Nigrospora sphaerica HCH285: Transcriptomic Insights into Metabolic Regulation

Abstract

Background: Bostrycin, an anthraquinone secondary metabolite produced by Nigrospora sphaerica , exhibits broad-spectrum antitumor and antimicrobial activities, but its low production limits further development and application. Calcium ions (Ca²⁺) are important intracellular signaling molecules that regulate fungal physiology and secondary metabolism, but their role in bostrycin biosynthesis remains unclear. This study aimed to investigate the effect of exogenous Ca 2+ on bostrycin production and to elucidate the underlying molecular response using transcriptomic analysis. Methods: Ca²⁺, Mg²⁺, Na⁺, and K⁺ (supplemented as chloride salts) were screened for their ability to promote bostrycin production in N. sphaerica HCH285. The optimal Ca²⁺ concentration and addition time were determined by measuring bostrycin production using HPLC. Biomass accumulation, sporulation capacity, and extracellular nucleic acid and protein leakage were evaluated to characterize the physiological effects of Ca 2+ treatment.Control and Ca²⁺-treated cultures were subjected to transcriptome sequencing, followed by GO and KEGG enrichment analyses and qRT-PCR validation. Results: Among all tested metal ions, Ca 2+ showed the strongest stimulatory effect on bostrycin biosynthesis, with 4 g/L Ca 2+ producing the highest yield of bostrycin with 63.6 ± 0.31 μg/mL, representing a 2.55-fold increase over the control. Ca 2+ supplementation also significantly increased fungal biomass and sporulation capacity, suggesting that enhanced growth contributed to the elevated metabolite production. Increased extracellular nucleic acid and protein contents indicated altered cell integrity or secretion activity following Ca 2+ treatment.Transcriptomic analysis identified DEGs most significantly enriched in metabolic pathways, including oxidative phosphorylation, while genes related to polyketide metabolism and ubiquinone biosynthesis were also annotated. Several genes associated with ion transport, energy metabolism, and secondary metabolite biosynthesis were significantly upregulated. Among them, wbpA, talA, cho1, and aroB were further validated by qRT-PCR, indicating that Ca²⁺ supplementation triggered broad metabolic reprogramming associated with bostrycin accumulation. Conclusions: Exogenous Ca 2+ effectively enhances bostrycin production in N. sphaerica HCH285 through coordinated physiological and transcriptional responses. Transcriptomic analysis suggests that Ca 2+ supplementation promotes energy metabolism, ion transport, and secondary metabolic processes associated with bostrycin biosynthesis. These findings provide new insights into calcium-mediated regulation of fungal secondary metabolism and offer a potential strategy for improving bostrycin production through fermentation optimization and metabolic engineering.

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