Oxidative Stress Causes Vacuolar Fragmentation in the Human Fungal Pathogen Cryptococcus neoformans.

Donghyeun Kim,Moonyong Song,James W Kronstad,Yoojeong Choi,Won Hee Jung,Eunsoo Do

doi:10.3390/jof7070523

Abstract

Vacuoles are dynamic cellular organelles, and their morphology is altered by various stimuli or stresses. Vacuoles play an important role in the physiology and virulence of many fungal pathogens. For example, a Cryptococcus neoformans mutant deficient in vacuolar functions showed significantly reduced expression of virulence factors such as capsule and melanin synthesis and was avirulent in a mouse model of cryptococcosis. In the current study, we found significantly increased vacuolar fragmentation in the C. neoformans mutants lacking SOD1 or SOD2, which respectively encode Zn, Cu-superoxide dismutase and Mn-superoxide dismutase. The sod2 mutant showed a greater level of vacuole fragmentation than the sod1 mutant. We also observed that the vacuoles were highly fragmented when wild-type cells were grown in a medium containing high concentrations of iron, copper, or zinc. Moreover, elevated temperature and treatment with the antifungal drug fluconazole caused increased vacuolar fragmentation. These conditions also commonly cause an increase in the levels of intracellular reactive oxygen species in the fungus, suggesting that vacuoles are fragmented in response to oxidative stress. Furthermore, we observed that Sod2 is not only localized in mitochondria but also in the cytoplasm within phagocytosed C. neoformans cells, possibly due to copper or iron limitation.

Highlights

The vacuole is an important cellular organelle in fungi, and it performs diverse functions including the storage of ions, amino acids, phosphate, and carbohydrates
We studied the effect of deletion of SOD1 and SOD2 on vacuole morphology and the response to oxidative stress to determine the role of superoxide dismutase (SOD) in the physiology of C. neoformans
The functionality of the fusion proteins was confirmed by growing the cells under high concentrations of NaCl and in the presence of a superoxide anion generating agent menadione, the conditions of which reduced the growth of the sod1 and sod2 mutants [7,18]

Summary

Introduction

The vacuole is an important cellular organelle in fungi, and it performs diverse functions including the storage of ions, amino acids, phosphate, and carbohydrates. The vacuole morphology dynamically adapts to intra- and extracellular environmental conditions through fusion (forming a single enlarged vacuole) and fission, which is called fragmentation. Vacuole fusion is induced by nutrient limitation or hypotonic treatment, whereas vacuole fragmentation is triggered by hyperosmotic shock, endoplasmic reticulum stress, or lactic acid stress in the model fungus Saccharomyces cerevisiae [1,2]. Examples include Fab, a lipid kinase that produces phosphatidylinositol 3,5-bisphosphate [PI(3,5)P2 ] from phosphatidylinositol 3-phosphate in S. cerevisiae. The S. cerevisiae mutant lacking the FAB1 gene exhibits an enlarged vacuole, indicating the involvement of the protein in vacuole fragmentation [4].

Methods

Results

Conclusion