The isocyanide functional group is important in the synthesis of many organic compounds and is found in natural products produced by plants, bacteria, marine invertebrates, and fungi. The antimicrobial activities of isocyanide compounds have been documented for almost 70 years, however, the biochemical targets and mechanisms of action remain poorly defined. We report antimicrobial activity of 4-para-nitrophenyl-isocyanide (p-NPIC) against a model fungus, Saccharomyces cerevisiae, and the human fungal pathogen Candida albicans. To identify the cellular and molecular targets of p-NPIC, we screened the non-essential single gene-deletion collection of S. cerevisiae. We aimed to identify genes which, when absent, rendered the resulting strain incapable of growth on solid media containing 1.5 µM p-NPIC. We identified 167 strains that were hypersensitive to p-NPIC and determined the minimum inhibitory concentration of p-NPIC for each of these mutant strains. The most sensitive deletion-strains (MIC \u3c 3.0 µM in liquid media) were enriched in mitochondrial functions including the mitochondrial type II fatty acid synthase, lipoic acid biosynthesis and protein lipoylation, synthesis and assembly of iron-sulfur clusters, and assembly and maintenance of cytochrome c oxidase. The identification of essentially all components of the proton-pumping vacuolar membrane ATPase, as well as some pH sensitive components of membrane lipid biosynthesis, also suggest a role for regulation of cytoplasmic pH as a key determinant of p-NPIC tolerance. Taken together, these results suggest that mitochondrial metal homeostasis and reactive oxygen scavenging are disrupted by p-NPIC treatment and provide new information about the potential mechanisms of action of isocyanide natural products.
Advisor: Wayne R. Riekho
