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Tracking protein aggregation and mislocalization in cells with flow cytometry

Abstract

We applied pulse-shape analysis (PulSA) to monitor protein localization changes in mammalian cells by flow cytometry. PulSA enabled high-throughput tracking of protein aggregation, translocation from the cytoplasm to the nucleus and trafficking from the plasma membrane to the Golgi as well as stress-granule formation. Combining PulSA with tetracysteine-based oligomer sensors in a cell model of Huntington's disease enabled further separation of cells enriched with monomers, oligomers and inclusion bodies.

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Figure 1: Use of PulSA to track changes in protein localization in cells.
Figure 2: PulSA combined with tetracysteine sensors of Htt oligomerization selectively separated cells enriched in Htt monomers, oligomers or inclusion bodies.

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References

  1. Hatters, D.M. IUBMB Life 60, 724–728 (2008).

    Article  CAS  Google Scholar 

  2. Silveira, J.R. et al. Nature 437, 257–261 (2005).

    Article  CAS  Google Scholar 

  3. Bucciantini, M. et al. Nature 416, 507–511 (2002).

    Article  CAS  Google Scholar 

  4. Arrasate, M., Mitra, S., Schweitzer, E.S., Segal, M.R. & Finkbeiner, S. Nature 431, 805–810 (2004).

    Article  CAS  Google Scholar 

  5. Johnston, J.A., Ward, C.L. & Kopito, R.R. J. Cell Biol. 143, 1883–1898 (1998).

    Article  CAS  Google Scholar 

  6. Kaganovich, D., Kopito, R. & Frydman, J. Nature 454, 1088–1095 (2008).

    Article  CAS  Google Scholar 

  7. De Vos, W.H., Van Neste, L., Dieriks, B., Joss, G.H. & Van Oostveldt, P. Cytometry A 77A, 64–75 (2010).

    Google Scholar 

  8. MacDonald, M.E. et al. Cell 72, 971–983 (1993).

    Article  Google Scholar 

  9. Hoffman, R.A. Curr. Protoc. Cytom. 1.23.21–21.23.17 (2009).

  10. Deng, H.X. et al. Science 261, 1047–1051 (1993).

    Article  CAS  Google Scholar 

  11. Ramdzan, Y.M. et al. Chem. Biol. 17, 371–379 (2010).

    Article  CAS  Google Scholar 

  12. Ossato, G. et al. Biophys. J. 98, 3078–3085 (2010).

    Article  CAS  Google Scholar 

  13. Lajoie, P. & Snapp, E.L. PLoS ONE 5, e15245 (2011).

    Article  Google Scholar 

  14. McKenna, B.K., Evans, J.G., Cheung, M.C. & Ehrlich, D.J. Nat. Methods 8, 401–403 (2011).

    Article  CAS  Google Scholar 

  15. Rizzo, M.A., Springer, G.H., Granada, B. & Piston, D.W. Nat. Biotechnol. 22, 445–449 (2004).

    Article  CAS  Google Scholar 

  16. Olshina, M.A. et al. J. Biol. Chem. 285, 21807–21816 (2010).

    Article  CAS  Google Scholar 

  17. Johannes, L., Tenza, D., Antony, C. & Goud, B. J. Biol. Chem. 272, 19554–19561 (1997).

    Article  CAS  Google Scholar 

  18. Mallard, F. et al. J. Cell Biol. 143, 973–990 (1998).

    Article  CAS  Google Scholar 

Download references

Acknowledgements

This work was funded by grants to D.M.H. (Australian National Health and Medical Research Council project grant 566640). D.M.H. is funded as a Grimwade fellow, supported by the Miegunyah Trust. D.C.H.N. is funded as a CR Roper fellow. The SOD1-GFP fusion constructs were provided by B.J. Turner (Florey Neurosciences Institute, Australia). HSPA1A (Hsp70) and DNAJB1 (Hsp40) in the pCMV vector was provided by P. Muchowski (Gladstone Institute of Neurological Disease and University of California, San Francisco).

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Y.M.R., S.P., C.P.Z.C., I.H.W.N., N.P.C., A.W.P., D.C.H.N. and A.R.O. performed experiments and/or helped interpret data. Y.M.R., M.A.B., D.C.H.N. and P.A.G. contributed in the design of some experiments. D.M.H. conceived of the method, oversaw implementation of experiments and wrote the manuscript.

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Correspondence to Danny M Hatters.

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The authors declare no competing financial interests.

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Supplementary Figures 1–6, Supplementary Note 1 (PDF 1664 kb)

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Ramdzan, Y., Polling, S., Chia, C. et al. Tracking protein aggregation and mislocalization in cells with flow cytometry. Nat Methods 9, 467–470 (2012). https://doi.org/10.1038/nmeth.1930

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