Abstract
Background: The labile nature of microtubules is critical for establishing cellular morphology and motility, yet the molecular basis of assembly remains unclear. Here we use optical tweezers to track microtubule polymerization against microfabricated barriers, permitting unprecedented spatial resolution. Results: We find that microtubules exhibit extensive nanometer-scale variability in growth rate and often undergo shortening excursions, in some cases exceeding five tubulin layers, during periods of overall net growth. This result indicates that the guanosine triphosphate (GTP) cap does not exist as a single layer as previously proposed. We also find that length increments (over 100 ms time intervals, n = 16,762) are small, 0.81 ± 6.60 nm (mean ± standard deviation), and very rarely exceed 16 nm (about two dimer lengths), indicating that assembly occurs almost exclusively via single-subunit addition rather than via oligomers as was recently suggested. Finally, the assembly rate depends only weakly on load, with the average growth rate decreasing only 2-fold as the force increases 7-fold from 0.4 pN to 2.8 pN. Conclusions: The data are consistent with a mechanochemical model in which a spatially extended GTP cap allows substantial shortening on the nanoscale, while still preventing complete catastrophe in most cases.
| Original language | English (US) |
|---|---|
| Pages (from-to) | 1445-1455 |
| Number of pages | 11 |
| Journal | Current Biology |
| Volume | 17 |
| Issue number | 17 |
| DOIs | |
| State | Published - Sep 4 2007 |
Bibliographical note
Funding Information:The authors would like to thank Edgar Meyhofer for valuable discussion and comments on this work and Yan Chen, Joachim Mueller, and Erkan Tuzel for helpful discussions. The chemical-engineering clean room at the University of Michigan supplied lithography equipment and expertise. This work was supported by the grants to A.J.H. from the Burroughs Wellcome Fund and the National Science Foundation MCB-0334835. H.T.S. was supported by the Whitaker Foundation, and M.K.G. is supported by National Institutes of Health National Research Service Award (NRSA) grant EB005568. D.J.O. is supported by National Institutes of Health grant GM071522 and by National Science Foundation grant MCB-0615568. The authors are grateful to the Minnesota Supercomputing Institute for providing computing resources and to David Do for technical assistance in lateral cap model simulations.
Keywords
- CELLBIO