The role of Pep4 protease in Cryptococcus neoformans cell survival and virulence factors

Cryptococcosis is a systemic mycosis caused by Cryptococcus neoformans with significant clinical importance, mainly affecting immunodeficient patients. The treatment options are limited to a few drugs, and resistance to them has been reported. Therefore, research is essential to broaden knowledge regarding the biology of this yeast, aiming to identify traits that could serve as new targets for antifungal drugs. This study aims to expand the current understanding of the autophagy process in this pathogenic yeast. Autophagy is a conserved intracellular degradation and recycling process among eukaryotes, indispensable in cellular homeostasis. In Saccharomyces cerevisiae, the PEP4 gene encodes a protease required during the final stages of autophagy, playing a role in the maturation and activation of vacuolar hydrolases, which contributes to cell survival under conditions of nutritional deprivation and stress. However, PEP4 has never been studied in C. neoformans. Thus, we evaluated the impact of PEP4 deletion on the expression of virulence factors and the cell response to multiple stress conditions. Our results demonstrated that the pep4Δ mutant exhibited attenuated virulence in Galleria mellonella and a decreased fungal burden in macrophages. Notably, we observed the accumulation of autophagic bodies in the pep4Δ strain under nutrient starvation, suggesting a defect in the final steps of autophagic degradation. These findings suggest that the Pep4 protein of C. neoformans plays a crucial role in vacuolar function and the adaptation and survival of yeast cells under stressful conditions, as well as in the host–pathogen interaction.