|M.Sc Student||Avila Cristina|
|Subject||Hemolytic Activity in C. Albicans|
|Department||Department of Medicine||Supervisor||Professor Daniel Kornitzer|
|Full Thesis text|
Fungal pathogens can cause devastating diseases in immunocompromised individuals. The most common opportunistic pathogen is Candida albicans. Like all pathogens, it has developed many strategies to acquire nutrients from the human host. To acquire iron, C. albicans has the ability to use hemoglobin as a source, by deploying a network of extracellular proteins that extract heme from hemoglobin and internalize it into the cell. However, to access hemoglobin, C. albicans must be able to lyse the erythrocytes, a function known as hemolysis. In the first part of this study we identified the aspartyl proteinases SAP1 and SAP3 as C. albicans genes that can confer hemolytic activity to the non-pathogenic yeast S. cerevisiae. The aspartyl proteinases are extracellular hydrolytic enzymes encoded by a family of 10 SAP genes. However, deletion of SAP3 from the C. albicans genome using CRISPR/Cas9 did not cause a significant reduction in the hemolytic activity. We suggest that multiple SAPs might mediate the hemolytic activity of C. albicans, which can therefore not be affected by a single SAP gene knockout.
In the second part of this study, we focused on the ESCRT-dependent endocytic pathway which is required for heme internalization. We identified RIM8, a β-arrestin gene, as being involved in heme-iron utilization. β-arrestins are involved in ubiquitin-mediated internalization of transmembrane channels and receptors. Deletion of RIM8 resulted in a strong reduction in the ability to utilize hemoglobin-iron source, suggesting a direct role in the heme internalization pathway. However, the rim8 phenotype was dependent on the nature of the iron chelator used, suggesting that the role of Rim8 depends on the presence or absence of additional transition metals.
Given the role of ubiquitin in ESCRT pathway and arrestin functions, in the third part of this study we screened for ubiquitin ligases that are potentially involved in the heme-iron utilization pathway. One mutant, rsp5, was identified that is defective in hemoglobin utilization, suggesting a possible role for this HECT-type ubiquitin ligase in endocytosis of the C. albicans heme receptor.