Identification of the Antifungal Metabolite Chaetoglobosin P From Discosia rubi Using a Cryptococcus neoformans Inhibition Assay: Insights Into Mode of Action and Biosynthesis

<i>Cryptococcus neoformans</i> is an important human pathogen with limited options for treatments. We have interrogated extracts from fungal fermentations to find <i>Cryptococcus</i>-inhibiting natural products using assays for growth inhibition, differential thermosensitivity, and synergy with existing antifungal drugs. Extracts from fermentations of strains of <i>Discosia rubi</i> from eastern Texas showed anticryptococcal bioactivity with preferential activity in agar zone of inhibition assays against <i>C. neoformans</i> at 37°C versus 25°C. Assay-guided fractionation led to the purification and identification of chaetoglobosin P as the active component of these extracts. Genome sequencing of these strains revealed a biosynthetic gene cluster consistent with chaetoglobosin biosynthesis and β-methylation of the tryptophan residue. Proximity of genes of the actin-binding protein twinfilin-1 to the chaetoglobosin P and K gene clusters suggested a possible self-resistance mechanism involving twinfilin-1 which is consistent with the predicted mechanism of action involving interference with the polymerization of the capping process of filamentous actin. A <i>C. neoformans</i> mutant lacking twinfilin-1 was hypersensitive to chaetoglobosin P. Chaetoglobosins also potentiated the effects of amphotericin B and caspofungin on <i>C. neoformans</i>.