All-in-One Fluorescent Probe: Enabling Simultaneous Tracking of Multiple Dynamic Organellar Perturbations during...

Area of Science:

• Biochemistry
• Cell Biology
• Molecular Medicine

Background:

• Ferroptosis is a regulated cell death pathway driven by lipid peroxidation.
• Understanding ferroptosis requires tools to monitor multiple organelles and their microenvironments simultaneously.
• Current tools are limited in their ability to provide concurrent, multi-organelle visualization.

Purpose of the Study:

• To develop a novel single fluorescent probe for simultaneous monitoring of mitochondria, lysosomes, and lipid droplets.
• To utilize this probe to visualize the dynamic changes during ferroptosis.
• To investigate the role of SLC7A11 in ferroptosis by assessing its impact on organelle homeostasis.

Main Methods:

• Synthesis and characterization of a novel single fluorescent probe, ATBI.
• Simultaneous multicolor imaging of mitochondria, lysosomes, and lipid droplets using ATBI.
• Fluorescence lifetime imaging microscopy (FLIM) to quantify viscosity changes.
• Treatment with Erastin to induce ferroptosis and observation of organelle dynamics.
• Assessment of SLC7A11's effect on organelle morphology and viscosity during ferroptosis.

Main Results:

• ATBI allows simultaneous, spectrally distinct visualization of mitochondria/lysosomes (~719 nm) and lipid droplets (~425 nm).
• Mitochondria and lysosomes are distinguishable by morphology and fluorescence lifetime, enabling viscosity quantification.
• Erastin-induced ferroptosis is characterized by lipid droplet accumulation, increased lysosomal viscosity, and decreased mitochondrial viscosity.
• SLC7A11 expression restored normal organelle morphology and viscosity homeostasis, suggesting a protective role.

Conclusions:

• The novel probe ATBI is a versatile tool for multi-organelle imaging and microenvironment analysis.
• Visualizing ferroptosis dynamics revealed key organelle perturbations and viscosity changes.
• Maintaining subcellular organelle morphology and microenvironmental stability is crucial for resisting ferroptosis, with SLC7A11 playing a significant role.