Molecular Mechanisms of Stress-Induced Mitochondrial Damage

The main objective of my research program is to identify the molecular mechanisms of mitochondrial dysfunction in pathological stress conditions. Proper mitochondrial functioning involves the production of the energy source, in the form of ATP molecules, through the process of oxidative phosphorylation (OXPHOS). During OXPHOS, the oxidation of high-energy substrates in the enzymes of the inner mitochondrial membrane (IMM) is coupled with the creation of an electrochemical gradient across the membrane, referred to as the mitochondrial membrane potential (ΔΨm). The energy of ΔΨm is used by the ATP synthase enzyme (ATPase) to phosphorylate ADP and produce ATP. The essential condition for proper OXPHOS is the tightly regulated transport of ions and solutes across the IMM and its integrity. Numerous pathologies have been shown to disturb the integrity of the IMM and disrupt the production of energy required for tissue preservation. Up to now, the only partially identified mechanism of mitochondrial damage is mitochondrial permeability transition (mPT). mPT is believed to occur through the opening of a pore (mPTP) in the IMM stimulated by an elevation of Ca2+ and reactive oxygen species (ROS) concentrations in the mitochondrial matrix. mPTP opening enables uncontrolled permeability in IMM, which is associated with mitochondrial depolarization (a drop in ΔΨm), swelling, and leads to the energy starvation of cells and tissues, subsequently resulting in cell death. Inhibition of mPT is highly protective for tissues in conditions ranging from stroke and heart attack to neurodegenerative disorders and diabetes, highlighting the importance of preserving mitochondrial functions. Recently, I applied original experimental approaches: ex vivo patch clamp and holographic imaging to address the mechanisms of mitochondrial dysfunction. The advantage of these methodologies lies in their ability to detect mPTP independently of mitochondrial depolarization or swelling. I discovered that stress-induced mitochondrial damage and cell death might occur through both mPTP opening and mechanisms independent of mPTP formation. Interestingly, mPTP and non-mPTP mechanisms share similar induction stressors and features. In this project, I propose to employ an interdisciplinary methodological combination of my novel functional approaches with high-resolution Cryo imaging, structural proteomics, and genetic modifications to investigate novel mPTP-independent mechanisms of mitochondrial dysfunction and identify the protein complex responsible for mPTP formation. This paradigm shift from the uniform mPTP mechanism to multiple pathways of mitochondrial damage and unique methodological advancement will allow me to identify the link between the mechanisms of intracellular Ca2+ and ROS signaling, the mode of mitochondrial dysfunction, and mitochondria- associated cell death within cellular and tissue models of pathological stress.