Visualizing PIEZO1 Localization and Activity in hiPSC-Derived Single Cells and Organoids with HaloTag Technology.

Area of Science:

• Biophysics
• Cell Biology
• Biotechnology

Background:

• PIEZO1 channels are crucial mechanosensors involved in various physiological processes.
• Visualizing endogenous PIEZO1 activity and localization is essential for understanding its function.
• Existing methods may lack the resolution or applicability for studying PIEZO1 in complex human systems.

Purpose of the Study:

• To develop a novel chemogenetic platform for visualizing and monitoring endogenous PIEZO1 activity in human cells and tissues.
• To enable physiologically and clinically relevant studies on human PIEZO1.
• To facilitate the study of PIEZO1 mechanotransduction in three-dimensional human tissue models.

Main Methods:

• Genetically engineered human induced pluripotent stem cells (hiPSCs) to express a HaloTag fused to endogenous PIEZO1.
• Utilized advanced imaging techniques, including lightsheet microscopy.
• Employed Ca2+-sensitive HaloTag ligands for non-invasive monitoring of channel activity.

Main Results:

• Established a hiPSC line with precisely visualized PIEZO1 localization dynamics.
• Achieved non-invasive, real-time monitoring of PIEZO1 channel activity with high temporal resolution.
• Demonstrated molecular-scale imaging of PIEZO1 in hiPSC-derived neural organoids.

Conclusions:

• The PIEZO1-HaloTag hiPSC platform offers a powerful tool for studying mechanotransduction in human systems.
• This technology facilitates the investigation of PIEZO1's role in health and disease.
• Advances potential for elucidating disease mechanisms and developing targeted therapeutics for PIEZO1-related conditions.