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Tracking single molecules at work in living cells

Nature Chemical Biology volume 10, pages 524–532 (2014)Cite this article

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Abstract

Methods for imaging and tracking single molecules conjugated with fluorescent probes, called single-molecule tracking (SMT), are now providing researchers with the unprecedented ability to directly observe molecular behaviors and interactions in living cells. Current SMT methods are achieving almost the ultimate spatial precision and time resolution for tracking single molecules, determined by the currently available dyes. In cells, various molecular interactions and reactions occur as stochastic and probabilistic processes. SMT provides an ideal way to directly track these processes by observing individual molecules at work in living cells, leading to totally new views of the biochemical and molecular processes used by cells whether in signal transduction, gene regulation or formation and disintegration of macromolecular complexes. Here we review SMT methods, summarize the recent results obtained by SMT, including related superresolution microscopy data, and describe the special concerns when SMT applications are shifted from the in vitro paradigms to living cells.

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Figure 1: A schematic model of the dynamic PM obtained by SMT.
Figure 2: SMT revealed that class A GPCRs continually form dimers and disintegrate into monomers within fractions of a second.
Figure 3: The archipelago architecture of the FA proposed on the basis of the SMT results.
Figure 4: LAT, a key adaptor molecule of immune signal transduction, might work in various dynamic states, as suggested by SMT and superresolution microscopy.

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Acknowledgements

The authors would like to thank K. Kanemasa for his help in preparing figures and all of the members of the Kusumi lab for extensive discussion. This research was supported in part by Grants-in-Aid for scientific research and for priority areas from the Ministry of Education, Culture, Sports, Science and Technology (MEXT) of Japan to R.S.K., T.K.F., T.A.T. and A.K. The WPI-iCeMS of Kyoto University is supported by the World Premiere Research Center Initiative of the MEXT.

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Authors and Affiliations

  1. Institute for Integrated Cell-Material Sciences (WPI-iCeMS), Kyoto University, Kyoto, Japan

    Akihiro Kusumi, Taka A Tsunoyama, Kohichiro M Hirosawa, Rinshi S Kasai & Takahiro K Fujiwara

  2. Institute for Frontier Medical Sciences, Kyoto University, Kyoto, Japan

    Akihiro Kusumi, Taka A Tsunoyama & Rinshi S Kasai

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  1. Akihiro Kusumi
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  2. Taka A Tsunoyama
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  3. Kohichiro M Hirosawa
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  5. Takahiro K Fujiwara
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Correspondence to Akihiro Kusumi.

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Kusumi, A., Tsunoyama, T., Hirosawa, K. et al. Tracking single molecules at work in living cells. Nat Chem Biol 10, 524–532 (2014). https://doi.org/10.1038/nchembio.1558

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