1. Fluorescent Biosensors for Multiplexed Imaging of Phosphoinositide Dynamics.
- Author
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Hertel, Fabian, Li, Simin, Chen, Mingyuan, Pott, Lutz, Mehta, Sohum, and Zhang, Jin
- Subjects
Biosensing Techniques ,Cell Line ,Cell Membrane ,Escherichia coli ,Fluorescence Resonance Energy Transfer ,Fluorescent Dyes ,Humans ,Kinetics ,Luminescent Proteins ,Optical Imaging ,Phosphatidylinositols ,Protein Domains ,Signal Transduction - Abstract
Phosphoinositides constitute a critical family of lipids that regulate numerous cellular processes. Phosphatidylinositol 4,5-bisphosphate (PIP2) is arguably the most important plasma membrane phosphoinositide and is involved in regulating diverse processes. It is also the precursor of phosphatidylinositol 3,4,5-trisphosphate (PIP3), which is critical for growth factor signaling, as well as membrane polarization and dynamics. Studying these lipids remains challenging, because of their compartmentalized activities and location-dependent signaling profiles. Here, we introduce several new genetically encoded fluorescent biosensors, including FRET-based and dimerization-dependent fluorescent protein (ddFP)-based biosensors, that enable real-time monitoring of PIP2 levels in live cells. In addition, we developed a red fluorescent biosensor for 3-phosphoinositides that can be co-imaged with the green PIP2 indicator. Simultaneous visualization of the dynamics of PIP2 and 3-phosphoinositides in the same cell shows that plasma membrane PIP3 formation upon EGF stimulation is coupled to a decrease in the local pool of PIP2.
- Published
- 2020