PIEZO1-HaloTag hiPSCs: Bridging Molecular, Cellular and Tissue Imaging

Gabriella A Bertaccini; Elizabeth L Evans; Jamison L Nourse; George D Dickinson; Gaoxiang Liu; Ignasi Casanellas; Sayan Seal; Alan T Ly; Jesse R Holt; Shijun Yan; Elliot E Hui; Mitradas M Panicker; Srigokul Upadhyayula; Ian Parker; Medha M Pathak

doi:10.1101/2023.12.22.573117

PIEZO1-HaloTag hiPSCs: Bridging Molecular, Cellular and Tissue Imaging

bioRxiv [Preprint]. 2023 Dec 23:2023.12.22.573117. doi: 10.1101/2023.12.22.573117.

Authors

Gabriella A Bertaccini^{1

2}, Elizabeth L Evans^{1

2}, Jamison L Nourse^{1

2}, George D Dickinson¹, Gaoxiang Liu³, Ignasi Casanellas^{1

2}, Sayan Seal³, Alan T Ly^{1

2}, Jesse R Holt^{1

2

4}, Shijun Yan⁵, Elliot E Hui⁵, Mitradas M Panicker^{1

2}, Srigokul Upadhyayula^{3

6

7}, Ian Parker^{1

8}, Medha M Pathak^{1

2

4

5}

Affiliations

¹ Department of Physiology and Biophysics, University of California, Irvine, Irvine, CA.
² Sue and Bill Gross Stem Cell Research Center, University of California, Irvine, Irvine, CA.
³ Advanced Bioimaging Center, Department of Molecular and Cell Biology, University of California, Berkeley, United States.
⁴ Center for Complex Biological Systems, University of California, Irvine, Irvine, CA.
⁵ Department of Biomedical Engineering, University of California, Irvine, Irvine, CA.
⁶ Molecular Biophysics and Integrated Bioimaging Division, Lawrence Berkeley National Laboratory, Berkeley, United States.
⁷ Chan Zuckerberg Biohub, San Francisco, United States.
⁸ Department of Neurobiology and Behavior, University of California, Irvine, Irvine, CA.

Abstract

PIEZO1 channels play a critical role in numerous physiological processes by transducing diverse mechanical stimuli into electrical and chemical signals. Recent studies underscore the importance of endogenous PIEZO1 activity and localization in regulating mechanotransduction. To enable physiologically and clinically relevant human-based studies, we genetically engineered human induced pluripotent stem cells (hiPSCs) to express a HaloTag fused to endogenous PIEZO1. Combined with super-resolution imaging, our chemogenetic approach allows precise visualization of PIEZO1 in various cell types. Further, the PIEZO1-HaloTag hiPSC technology allows non-invasive monitoring of channel activity via Ca²⁺-sensitive HaloTag ligands, with temporal resolution approaching that of patch clamp electrophysiology. Using lightsheet imaging of hiPSC-derived neural organoids, we also achieve molecular scale PIEZO1 imaging in three-dimensional tissue samples. Our advances offer a novel platform for studying PIEZO1 mechanotransduction in human cells and tissues, with potential for elucidating disease mechanisms and development of targeted therapeutics.

Publication types

Preprint

Abstract

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