Superoxide dismutases and mitochondrial oxidative stress in Candida albicans.

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 DESCRIPTION (provided by applicant): From bacteria to humans, the superoxide dismutases (SODs) enzymes play important roles in oxidative stress resistance by catalyzing the disproportionation of superoxide to oxygen and hydrogen peroxide. A typical eukaryotic cell contains a Cu/Zn SOD1 that resides largely in the cytosol with a small fraction localized to the intermembrane space (IMS) of the mitochondria, while the mitochondrial matrix contains a distinct manganese SOD2. Such partitioning of copper and manganese SODs has been well conserved throughout evolution but a striking deviation can be seen with the human fungal pathogen, Candida albicans. C. albicans uniquely expresses both a manganese and copper containing SOD in the same cytosolic compartment. The Culotta lab has recently shown that these two SODs are differentially expressed according to growth state and copper. With rapidly dividing cells and abundant copper, Cu/Zn SOD1 predominates, while in long-term stationary phase when copper becomes limiting, cells switch from Cu/Zn SOD1 to Mn SOD3. This switch is evident in laboratory cultures of C. albicans as well as in infection models for disseminated candidiasis. However, the biological rationale for this switch in SOD enzymes is completely unknown. If the pathogen is susceptible to copper depravation, why has it retained Cu/Zn SOD1? My hypothesis is that Cu/Zn SOD1 but not Mn SOD3, protects C. albicans from mitochondrial oxidative stress when cells are rapidly dividing. Interestingly, C. albicans undergoes a switch in mitochondrial respiration that appears to accompany the switch in SOD enzymes. Rapidly dividing cells utilize the conventional copper dependent cytochrome C oxidase (COX) driven respiration that is predicted to generate ROS; stationary phase cells use an alternative oxidase (AOX) that does not require copper and is predicted to generate low ROS. I predict that during COX respiration, Cu/Zn SOD1 enters the mitochondria to guard against mitochondrial ROS, while during AOX respiration, there is less need for an IMS SOD and Mn SOD3 remains cytosolic. This possible connection between SOD1, SOD3 and mitochondrial respiration and oxidative stress will be addressed as follows: Aim 1: To understand the connection between C. albicans SOD1 and SOD3 and mitochondrial respiration. Conditions for COX and AOX respiration will be optimized and isolation of mitochondria during these conditions will be used to probe for SOD1 and SOD3 in the IMS of the mitochondria. We will also test the effects of high and low intracellular copper on mitochondrial respiration and mitochondrial uptake of SOD1 and SOD3. Aim 2: To determine the role of SOD1 and SOD3 in mitochondrial oxidative stress protection. We will compare whole cell and mitochondrial ROS produced in COX versus AOX respiration. We will investigate a role for SOD1 and SOD3 in mitochondrial oxidative stress protection by monitoring protein oxidation and mitochondrial function. Together these basic science studies on SOD enzymes and mitochondrial biology in C. albicans can provide important insight as to how this pathogenic yeast can adapt and survive long term in a human host.