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Fluorescence fluctuations of quantum-dot sensors capture intracellular protein interaction dynamics

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Abstract

We extend the in vitro principle of co-immunoprecipitation to quantify dynamic protein interactions in living cells. Using a multiresolution implementation of fluorescence correlation spectroscopy to achieve maximal temporal resolution, we monitored the interactions of endogenous bait proteins, recruited by quantum dots, with fluorescently tagged prey. With this approach, we analyzed the rapid physiological regulation of protein kinase A.

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Figure 1: Quantum dot–based visual immunoprecipitation (QD-VIP).
Figure 2: Optimizing the temporal resolution of QD-VIP by multiparametric analysis and mrFCS.
Figure 3: Monitoring PKA response to physiological stimulations.

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References

  1. Niethammer, P. et al. PLoS Biol. 5, e29 (2007).

    Article  Google Scholar 

  2. Carlson, C.R. et al. J. Biol. Chem. 281, 21535–21545 (2006).

    Article  CAS  Google Scholar 

  3. Bacia, K., Kim, S.A. & Schwille, P. Nat. Methods 3, 83–89 (2006).

    Article  CAS  Google Scholar 

  4. Lohse, M.J. et al. Trends Pharmacol. Sci. 29, 159–165 (2008).

    Article  CAS  Google Scholar 

  5. Houge, G., Steinberg, R.A., Ogreid, D. & Doskeland, S.O. J. Biol. Chem. 265, 19507–19516 (1990).

    CAS  PubMed  Google Scholar 

  6. Nikolaev, V.O., Gambaryan, S., Engelhardt, S., Walter, U. & Lohse, M.J. J. Biol. Chem. 280, 1716–1719 (2005).

    Article  CAS  Google Scholar 

  7. Violin, J.D. et al. J. Biol. Chem. 283, 2949–2961 (2008).

    Article  CAS  Google Scholar 

  8. Huang, L.J. & Taylor, S.S. J. Biol. Chem. 273, 26739–26746 (1998).

    Article  CAS  Google Scholar 

  9. Seet, B.T., Dikic, I., Zhou, M.M. & Pawson, T. Nat. Rev. Mol. Cell Biol. 7, 473–483 (2006).

    Article  CAS  Google Scholar 

  10. Peyker, A., Rocks, O. & Bastiaens, P.I. ChemBioChem 6, 78–85 (2005).

    Article  CAS  Google Scholar 

  11. Digman, M.A. et al. Biophys. J. 89, 1317–1327 (2005).

    Article  CAS  Google Scholar 

  12. Digman, M.A. et al. Biophys. J. 88, L33–L36 (2005).

    Article  CAS  Google Scholar 

  13. Kolin, D.L. & Wiseman, P.W. Cell Biochem. Biophys. 49, 141–164 (2007).

    Article  CAS  Google Scholar 

  14. Hebert, B., Costantino, S. & Wiseman, P.W. Biophys. J. 88, 3601–3614 (2005).

    Article  CAS  Google Scholar 

  15. Zamir, E. & Bastiaens, P.I. Nat. Chem. Biol. 4, 643–647 (2008).

    Article  CAS  Google Scholar 

  16. Zaccolo, M. et al. Nat. Cell Biol. 2, 25–29 (2000).

    Article  CAS  Google Scholar 

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Acknowledgements

This work was supported by the European Molecular Biology Organization (fellowship to E.Z.) and the Federation of European Biochemical Societies (fellowship to P.H.M.L.). H.E.G. and P.H.M.L. were also supported by the Center for Systems Biology co-financed by European Regional Development Fund and the state of North-Rhine Westphalia. E.Z. was also supported by the EU integrated project grant “Interaction Proteome”. Part of the work was conducted at the Cell Biology and Biophysics Unit, European Molecular Biology Laboratory. We thank M. Zaccolo (Dulbecco Telethon Institute, Venetian Institute of Molecular Medicine, Padova) for the PKA-Cα-EYFP plasmid, S.S. Taylor and R.Y. Tsien (University of California, San Diego) for the mGFP-PKA-RIα plasmid, S. Gentz and C.F. Becker (Max Planck Institute of Molecular Physiology, Dortmund) for the synthesis of the RIAD peptide, M.A. Hink for helpful discussions and A. Krämer for help with manuscript preparation.

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Author notes

  1. Eli Zamir and Piet H M Lommerse: These authors contributed equally to this work.

Authors and Affiliations

  1. Department of Systemic Cell Biology, Max Planck Institute of Molecular Physiology, Dortmund, Germany

    Eli Zamir, Piet H M Lommerse, Ali Kinkhabwala, Hernán E Grecco & Philippe I H Bastiaens

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  1. Eli Zamir
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  2. Piet H M Lommerse
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  3. Ali Kinkhabwala
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  4. Hernán E Grecco
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  5. Philippe I H Bastiaens
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Contributions

E.Z., P.H.M.L. and P.I.H.B. devised the method. E.Z. and P.H.M.L. performed the experiments and analyzed the data. E.Z., P.H.M.L., A.K. and H.E.G. developed the analysis. E.Z., P.H.M.L. and P.I.H.B. wrote the paper.

Corresponding author

Correspondence to Philippe I H Bastiaens.

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The authors declare no competing financial interests.

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Supplementary Figures 1–4, Supplementary Tables 1–2, Supplementary Notes 1–2 (PDF 2918 kb)

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Zamir, E., Lommerse, P., Kinkhabwala, A. et al. Fluorescence fluctuations of quantum-dot sensors capture intracellular protein interaction dynamics. Nat Methods 7, 295–298 (2010). https://doi.org/10.1038/nmeth.1441

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  • DOI: https://doi.org/10.1038/nmeth.1441

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