Novel Mitochondrial And Lysosomal Probes For Probing Organelle Homeostasis

Both mitochondria and lysosomes are important organelles for maintaining cell homeostasis.Mitochondrial membrane potential is critical for mitochondrial functions and cell performance.Loss or attenuation of mitochondrial membrane potential often leads to up-regu lation of ROS generation,autophagy and cell death,etc.Therefore,studying the changes of mitochondrial membrane potential under different stress states is crucial for understanding the roles of mitochondrial membrane potential in different physiological processes and various diseases.Lysosomes are acidic organelles and contain different kinds of acid hydrolase,serving as the essential waste treatment system in cells.Lysosomal parameters,such as pH,locations,quantity and volume,undergo significant changes in cell stress and myriad diseases.Therefore,imaging and tracking the changes of lysosome parameters under stress conditions would facilitate investigation on the biological processes related to lysosomal homeostasis.This dissertation is divided into three chapters.The first chapter firstly introduces the biological significance of mitochondrial membrane potential on organelle homeostasis,and pros/cons of current methods for studying mitochondrial membrane potential.Then we the described the development of Staadinger reaction for bioorthogonal probes in bioimaging.Finally,we introduced the biological functions of lysosomes and available fluorescent probes for lysosomes.In the second chapter,we designed a non-fluorescent probe AZF-TPP,which undergoes the Staudinger reaction to generate an anionic fluorescent probe(F-TPP)to achieve a "signal-on" analytical imaging of mitochondria.Compared to classical cationic "always-on" fluorescent probes specific for mitochondria,this organelle-oriented bioorthogonal imaging approach effectively discern the change of mitochondrial membrane potential under normal and different stress states.-With high sensitivity and selectivity,AZF-TPP represents a new perspective for real-time monitoring of the dynamic changes of mitochondria.In the third chapter,we studied and synthesized a fluorescent molecule ROXSA with good photostability,which is stably retained irrespective of lysosomal pH change.Distinct from widely used fluorescent probes prone to dissipation from stressed lysosomes,ROXSA enables long-term tracking of stressed lysosomes(0-120 h)with STORM super-resolution imaging,revealing time-lapsed lysosomes changes,dynamic fusion/fission of lysosomes during mitophagy,and engulfment of mitochondria into lysosomes.