Quantitative mapping of dense microtubule arrays in mammalian neurons

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Abstract

The neuronal microtubule cytoskeleton underlies the polarization and proper functioning of neurons, amongst others by providing tracks for motor proteins that drive intracellular transport. Different subsets of neuronal microtubules, varying in composition, stability and motor preference, are known to exist, but the high density of microtubules has so far precluded mapping their relative abundance and three-dimensional organization. Here we use different super-resolution techniques (STED, Expansion Microscopy) to explore the nanoscale organization of the neuronal microtubule network in rat hippocampal neurons. This revealed that in dendrites acetylated microtubules are enriched in the core of the dendritic shaft, while tyrosinated microtubules are enriched near the plasma membrane, thus forming a shell around the acetylated microtubules. Moreover, using a novel analysis pipeline we quantified the absolute number of acetylated and tyrosinated microtubules within dendrites and found that they account for 65-75% and ~20-30% of all microtubules, respectively, leaving only few microtubules that do not fall in either category. Because these different microtubule subtypes facilitate different motor proteins, these novel insights help to understand the spatial regulation of intracellular transport.

Data availability

All quantitative data is available on Figshare:https://doi.org/10.6084/m9.figshare.c.5306546.v2Software is available on Zenodo:https://doi.org/10.5281/zenodo.4281064https://doi.org/10.5281/zenodo.4534715https://doi.org/10.5281/zenodo.4534721

Article and author information

Author details

Eugene A Katrukha

Department of Biology, Utrecht University, Utrecht, Netherlands

Competing interests
The authors declare that no competing interests exist.
Daphne Jurriens

Biology, Utrecht University, Utrecht, Netherlands

Competing interests
The authors declare that no competing interests exist.
Desiree M Salas Pastene

Biology, Utrecht University, Utrecht, Netherlands

Competing interests
The authors declare that no competing interests exist.
Lukas C Kapitein

Biology, Utrecht University, Utrecht, Netherlands

For correspondence
l.kapitein@uu.nl

Competing interests
The authors declare that no competing interests exist.

"This ORCID iD identifies the author of this article:" 0000-0001-9418-6739

Funding

H2020 European Research Council (819219)

Lukas C Kapitein

ZonMw (91217002)

Daphne Jurriens
Lukas C Kapitein

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Ethics

Animal experimentation: Culturing of neurons has been approved by the ethical commitee (DEC) of Utrecht University and by the Centrale Commissie Dierproeven of the Dutch government (permit application AVD1080020173404). The ethical committee (DEC) is independent and must review any experimental use of animals in the Netherlands.

Copyright

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.