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Visualizing mechanical tension across membrane receptors with a fluorescent sensor

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Abstract

We report a fluorescence-based turn-on sensor for mapping the mechanical strain exerted by specific cell-surface proteins in living cells. The sensor generates force maps with high spatial and temporal resolution using conventional fluorescence microscopy. We demonstrate the approach by mapping mechanical forces during the early stages of regulatory endocytosis of the ligand-activated epidermal growth factor receptor (EGFR).

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Figure 1: Design and response of the EGFR tension sensor.
Figure 2: Characterization and quantification of the EGFR tension sensor.

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Acknowledgements

We thank A. Mattheyses (Emory University) for the CLC-eGFP plasmid and R. Nahta (Emory University Winship Cancer Institute) for the HCC1143 cells. We acknowledge the Emory University Winship Cancer Institute for support. K.S.S. acknowledges the Georgia Cancer Coalition Cancer Research Award for its support.

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Authors and Affiliations

  1. Department of Chemistry, Emory University, Atlanta, Georgia, USA

    Daniel R Stabley, Carol Jurchenko, Stephen S Marshall & Khalid S Salaita

Authors
  1. Daniel R Stabley
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  2. Carol Jurchenko
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  3. Stephen S Marshall
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  4. Khalid S Salaita
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Contributions

D.R.S. adapted the FRET surface sensor for use with human cells expressing the EGFR and performed the majority of the human cell experiments. C.J. developed the force sensor and performed the quantitative characterization of the zero-force sensor conformation and its components. S.S.M. optimized and performed the CLC-eGFP transfections. K.S.S. devised the overall experimental strategy. D.R.S., C.J. and K.S.S. wrote and edited the manuscript.

Corresponding author

Correspondence to Khalid S Salaita.

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Competing interests

The authors declare no competing financial interests.

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–13 (PDF 10105 kb)

Supplementary Video 1

Animation showing the mechanism of sensor function. (AVI 3308 kb)

Supplementary Video 2

Movie showing cell activation of the force sensor. (AVI 513 kb)

Supplementary Video 3

Movie showing clathrin colocalization with force sensor activation. (AVI 4616 kb)

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Stabley, D., Jurchenko, C., Marshall, S. et al. Visualizing mechanical tension across membrane receptors with a fluorescent sensor. Nat Methods 9, 64–67 (2012). https://doi.org/10.1038/nmeth.1747

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  • DOI: https://doi.org/10.1038/nmeth.1747

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