Dissecting the Mechanism of Action of a Novel Antifungal Peptide

摘要

There is an urgent need for novel treatments for Candida infections. The utility of antimicrobial peptides for antifungal therapy has garnered interest in recent years. One promising family of peptides is the Histatins, a family of naturally-occurring peptides secreted into the oral cavity that display antimicrobial activity. Histatin 5 is a twenty-four amino acid peptide with strong antifungal activity. Studies from our laboratory have identified a small histatin-derived peptide, KM29, that yields fungicidal activity 10-fold greater than Histatin 5 against multiple Candida species. Our laboratory has focused on understanding the mechanism of action of KM29 to further develop it as a therapeutic agent for oral and systemic candidiasis. To this end, a genetic screen was carried out using the available genome-wide deletion collection in S. cerevisiae. Our goal was to use this as a subrogates species to learn about the killing mechanism used by KM29 in Candida species. Analysis of the mutants revealed a significant presence of genes involved in mitochondrial function conferring increased resistance to KM29. We hypothesized that the S. cerevisiae mutants affected in different aspects of mitochondrial function will be more resistant to KM29 either because there is less ROS production due to their defective mitochondria, or less ATP production, which in turn may decrease peptide uptake and/or mitochondrial localization. We observed concentration dependent ROS production after exposure to KM29, however, this ROS production was loosely correlated with cell death. We also observed mitochondrial membrane potential depolarization and mitochondrial fission after exposure to KM29, indicating impairment of mitochondrial function. Additionally, we observed that the respiratory status of yeast cells inversely regulates KM29 fungicidal activity by influencing KM29 uptake. In conclusion, these studies provide valuable insights into the mechanism of action of KM29 and of cationic peptides in general.