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Dynein achieves processive motion using both stochastic and coordinated stepping

Nature Structural & Molecular Biology volume 19, pages 193–200 (2012)Cite this article

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Abstract

Processivity, the ability of single molecules to move continuously along a track, is a fundamental requirement of cargo-transporting molecular motors. Here, we investigate how cytoplasmic dynein, a homodimeric, microtubule-based motor, achieves processive motion. To do this, we developed a versatile method for assembling Saccharomyces cerevisiae dynein heterodimers, using complementary DNA oligonucleotides covalently linked to dynein monomers labeled with different organic fluorophores. Using two-color, single-molecule microscopy and high-precision, two-dimensional tracking, we find that dynein has a highly variable stepping pattern that is distinct from all other processive cytoskeletal motors, which use 'hand-over-hand' mechanisms. Uniquely, dynein stepping is stochastic when its two motor domains are close together. However, coordination emerges as the distance between motor domains increases, implying that a tension-based mechanism governs these steps. This plasticity may allow tuning of dynein for its diverse cellular functions.

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Figure 1: Dynein structure and constructs used in this study.
Figure 2: Two-dimensional stepping analysis of GST–dynein homodimers.
Figure 3: DNA-based dynein heterodimers are functional and step similarly to protein-based dynein homodimers.
Figure 4: Two-color tracking of dynein stepping.
Figure 5: Spatial arrangement of dynein motor domains during the two-head-bound state.
Figure 6: Dynein steps are stochastic at short head-to-head spacing and coordinated as head-to-head spacing increases.

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Acknowledgements

We thank X. Su and D. Pellman (Harvard Medical School) for providing purified kinesin-8; S. Zou for technical assistance; A. Carter, S. Churchman, A. Gennerich, Y. Goldman, A. Hendricks, J. Huang, A. Leschziner and A. Roberts for critical comments on the manuscript; F. Aguet, A. Besser, M. Vilela and G. Danuser for discussions of data analysis; M. Bagonis for early work on oligomer-SNAP linking; A. Leschziner for help with figure design; and A. Carter for providing MATLAB code. W.Q. is supported by a postdoctoral fellowship from the American Heart Association. S.L.R.-P. is funded by the Rita Allen Foundation, the Harvard Armenise Foundation and a US National Institutes of Health New Innovator award (1 DP2 OD004268-01).

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Author notes

  1. Weihong Qiu and Nathan D Derr: These authors contributed equally to this work.

Authors and Affiliations

  1. Department of Cell Biology, Harvard Medical School, Boston, Massachusetts, USA

    Weihong Qiu, Nathan D Derr, Brian S Goodman & Samara L Reck-Peterson

  2. Dana Farber Cancer Institute, Boston, Massachusetts, USA

    Nathan D Derr & William Shih

  3. Department of Biochemistry and Molecular Pharmacology, Harvard Medical School, Boston, Massachusetts, USA

    Nathan D Derr & William Shih

  4. Department of Molecular Structural Biology, Max Planck Institute of Biochemistry, Martinsried, Germany

    Elizabeth Villa

  5. Physiology Course, Marine Biological Laboratory, Woods Hole, Massachusetts, USA

    Elizabeth Villa & David Wu

  6. Geffen School of Medicine, University of California, Los Angeles, Los Angeles, California, USA

    David Wu

  7. Wyss Institute, Harvard University, Boston, Massachusetts, USA

    William Shih

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  1. Weihong Qiu
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Contributions

W.Q. and N.D.D. contributed equally. W.Q., N.D.D., W.S. and S.L.R.-P. designed the experiments. W.Q., N.D.D. and B.S.G. conducted the experiments and analyzed the data. W.Q., N.D.D., B.S.G. and S.L.R.-P. wrote the paper. E.V. and D.W. wrote the two-dimensional particle tracking code.

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Correspondence to Samara L Reck-Peterson.

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Qiu, W., Derr, N., Goodman, B. et al. Dynein achieves processive motion using both stochastic and coordinated stepping. Nat Struct Mol Biol 19, 193–200 (2012). https://doi.org/10.1038/nsmb.2205

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