Changes in seam number and location induce holes within microtubules assembled from porcine brain tubulin and in Xenopus egg cytoplasmic extracts
- Université de Rennes 1, CNRS, France
- University of Colorado Boulder, United States
- Paul Scherrer Institute, Switzerland
Abstract
Microtubules are tubes of about 25 nm in diameter that are critically involved in a variety of cellular functions including motility, compartmentalization, and division. They are considered as pseudo-helical polymers whose constituent ab-tubulin heterodimers share lateral homotypic interactions, except at one unique region called the seam. Here, we used a segmented sub-tomogram averaging strategy to reassess this paradigm and analyze the organization of the ab-tubulin heterodimers in microtubules assembled from purified porcine brain tubulin in the presence of GTP and GMPCPP, and in Xenopus egg cytoplasmic extracts. We find that in almost all conditions, microtubules incorporate variable protofilament and/or tubulin subunit helical-start numbers, as well as variable numbers of seams. Strikingly, the seam number and location vary along individual microtubules, generating holes of one to a few subunits in size within their lattices. Together, our results reveal that the formation of mixed and discontinuous microtubule lattices is an intrinsic property of tubulin that requires the formation of unique lateral interactions without longitudinal ones. They further suggest that microtubule assembly is tightly regulated in a cytoplasmic environment.
Data availability
Sub-tomogram averages and extracts from cryo-electron tomograms presented in the figures have been deposited onto the EMDB and are listed in Supplementary Table 1 with reference to the corresponding figures and videos. All the tilt series, tomograms, models and motiv lists used to reconstruct the microtubule segments in PEET have been deposited onto the EMPIAR (Supplementary Table 2).
Article and author information
Author details
Funding
Agence Nationale de la Recherche (ANR-16-CE11-0017-01)
- Denis Chrétien
Agence Nationale de la Recherche (ANR-18-CE13-0001-01)
- Denis Chrétien
Human Frontier Science Program (CDA00019/1019-C)
- Romain Gibeaux
Swiss National Science Fondation (310030_192566)
- Michel O Steinmetz
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Ethics
Animal experimentation: All animal experimentation in this study was performed according to our animal use protocol APAFiS #26858-2020072110205978 approved by the Animal Use Ethic Committee (#7, Rennes, France) and the French Ministry of Higher Education, Research and Innovation. Mature Xenopus laevis female frogs were obtained from the CRB Xénope (Rennes, France) and ovulated with no harm to the animals with at least a 6-month rest interval between ovulations.
Copyright
© 2022, Guyomar et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
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Changes in seam number and location induce holes within microtubules assembled from porcine brain tubulin and in Xenopus egg cytoplasmic extracts
eLife 11:e83021.
https://doi.org/10.7554/eLife.83021
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