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Endogenous Piezo1 Can Confound Mechanically Activated Channel Identification and Characterization
Adrienne E Dubin1, Swetha Murthy1, Amanda H Lewis2
1Howard Hughes Medical Institute, Department of Neuroscience, Dorris Neuroscience Center, The Scripps Research Institute, La Jolla, CA 92037, USA.
Neuron
|April 21, 2017
Summary
Mechanically activated (MA) currents were not detected in cells lacking endogenous Piezo1, suggesting Piezo1 is essential for observed MA currents. This highlights the need for naive cells lacking endogenous channels in mechanotransduction studies.
Area of Science:
- Cellular Electrophysiology
- Mechanobiology
- Molecular Biology
Background:
- Heterologous expression in naive cells is standard for characterizing mechanically activated (MA) currents.
- Recent studies proposed TMEM150c and Piezo1 N-terminal domains as MA channel components based on this method.
Purpose of the Study:
- To validate findings on TMEM150c and Piezo1 as MA channels.
- To emphasize the critical need for endogenous MA channel-free systems in mechanotransduction research.
Main Methods:
- Replication of heterologous expression experiments for TMEM150c and Piezo1 constructs.
- CRISPR/Cas9 inactivation of endogenous Piezo1 in HEK cells.
- Electrophysiological recordings to measure MA currents.
Main Results:
- Replicated constructs showed modest MA currents in a small fraction of naive HEK cells.
- These MA currents were absent in HEK cells with CRISPR/Cas9 inactivated endogenous Piezo1.
- The findings challenge the proposed roles of TMEM150c and Piezo1's N-terminus.
Conclusions:
- Observed MA currents are dependent on endogenous Piezo1.
- Naive cells lacking endogenous MA channels are crucial for accurate assessment of candidate mechanotransduction proteins.
- This study underscores the importance of rigorous controls in mechanobiology research.
