Accumulation of mitochondrial unfolded protein is a pathological threat insufficiently mentioned about the pathophysiological contribution and intrinsic counter mechanisms. This accumulation leads to mitochondrial dysfunction and cellular stress. Hence, in response to this stress, mitochondrial selective autophagy known as mitophagy is potentially induced as a protective mechanism to eliminate the damaged mitochondria and aggregates unfolded protein. As another quality control machinery of mitochondria, mitochondria-derived vesicles (MDVs) can also be released from the damaged mitochondria. MDVs are small vesicles derived from the mitochondrial membrane that contain various cargo molecules. The aim of my thesis project was studying mitophagy and release of MDVs in cellular models of inducible unfolded protein accumulation in mitochondrial matrix and intermembrane space. During this thesis the mitochondrial unfolded protein model was generated by targeting luciferase mutant, which has shown as unfolded proteins, into mitochondria. Moreover, Immunofluorescence techniques were used during this project in order to visualize and differentiate MDVs. Also, in order to track the mitophagy, we have generated an optimized system utilizing a different mitophagy sensor. Our results demonstrated well-defined MDVS including PDH-positive MDVs and TOM20-Positive MDVs. We have also proved that the enhanced system provides a valuable alternative mitophagy sensor in which mitochondria-targeting EGFP conjugated mTagBFP2 can be used instead of mCherry and in order to take advantage of the blue color instead of red.
Mitophagy and release of mitochondrial derived vesicles in cellular models of inducible unfolded protein accumulation in mitochondrial matrix and intermembrane space.
NEJADFARD, DELARAM
2022/2023
Abstract
Accumulation of mitochondrial unfolded protein is a pathological threat insufficiently mentioned about the pathophysiological contribution and intrinsic counter mechanisms. This accumulation leads to mitochondrial dysfunction and cellular stress. Hence, in response to this stress, mitochondrial selective autophagy known as mitophagy is potentially induced as a protective mechanism to eliminate the damaged mitochondria and aggregates unfolded protein. As another quality control machinery of mitochondria, mitochondria-derived vesicles (MDVs) can also be released from the damaged mitochondria. MDVs are small vesicles derived from the mitochondrial membrane that contain various cargo molecules. The aim of my thesis project was studying mitophagy and release of MDVs in cellular models of inducible unfolded protein accumulation in mitochondrial matrix and intermembrane space. During this thesis the mitochondrial unfolded protein model was generated by targeting luciferase mutant, which has shown as unfolded proteins, into mitochondria. Moreover, Immunofluorescence techniques were used during this project in order to visualize and differentiate MDVs. Also, in order to track the mitophagy, we have generated an optimized system utilizing a different mitophagy sensor. Our results demonstrated well-defined MDVS including PDH-positive MDVs and TOM20-Positive MDVs. We have also proved that the enhanced system provides a valuable alternative mitophagy sensor in which mitochondria-targeting EGFP conjugated mTagBFP2 can be used instead of mCherry and in order to take advantage of the blue color instead of red.File | Dimensione | Formato | |
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https://hdl.handle.net/20.500.12608/51281