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Assembly dynamics of microtubules at molecular resolution

Abstract

Microtubules are highly dynamic protein polymers1 that form a crucial part of the cytoskeleton in all eukaryotic cells. Although microtubules are known to self-assemble from tubulin dimers, information on the assembly dynamics of microtubules has been limited, both in vitro2,3 and in vivo4,5, to measurements of average growth and shrinkage rates over several thousands of tubulin subunits. As a result there is a lack of information on the sequence of molecular events that leads to the growth and shrinkage of microtubule ends. Here we use optical tweezers to observe the assembly dynamics of individual microtubules at molecular resolution. We find that microtubules can increase their overall length almost instantaneously by amounts exceeding the size of individual dimers (8 nm). When the microtubule-associated protein XMAP215 (ref. 6) is added, this effect is markedly enhanced and fast increases in length of about 40–60 nm are observed. These observations suggest that small tubulin oligomers are able to add directly to growing microtubules and that XMAP215 speeds up microtubule growth by facilitating the addition of long oligomers. The achievement of molecular resolution on the microtubule assembly process opens the way to direct studies of the molecular mechanism by which the many recently discovered microtubule end-binding proteins regulate microtubule dynamics in living cells7,8,9.

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Figure 1: Measuring growth dynamics of MTs with optical tweezers.
Figure 2: High-resolution details of growth and shrinkage events.
Figure 3: Quantifying the sizes of the large steps with our step-fitting algorithm.
Figure 4: MT end mechanics.

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Acknowledgements

We thank T. Hyman and T. Mitchison for discussions; K. Kinoshita for help with the purification of XMAP215; S. Tans, K. Kuipers and D. Drechsel for a critical reading of the manuscript; and M. Footer for the gift of axonemes. This work is part of the research program of the Stichting voor Fundamenteel Onderzoek der Materie (FOM), which is supported financially by the Nederlandse Organisatie voor Wetenschappelijk Onderzoek (NWO).

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Correspondence to Marileen Dogterom.

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Supplementary information

Supplementary Methods 1

Additional methods used in this work. (PDF 24 kb)

Supplementary Methods 2

Key-hole trap for MT length measurements. The file also contains Supplementary Figure A1 and one reference. (PDF 112 kb)

Supplementary Methods 3

Step fitting algorithm. The file also contains Supplementary Figures C1–C3. (PDF 184 kb)

Supplementary Data

Dynamics of freely growing microtubules: effect of XMAP215. The file also contains Supplementary Figure B1, Supplementary Table 1 and one reference. (PDF 42 kb)

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Kerssemakers, J., Laura Munteanu, E., Laan, L. et al. Assembly dynamics of microtubules at molecular resolution. Nature 442, 709–712 (2006). https://doi.org/10.1038/nature04928

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