Growth cone advance requires EB1 as revealed by genomic replacement with a light-sensitive variant
- University of California, San Francisco, United States
- Erasmus MC, Netherlands
Abstract
A challenge in analyzing dynamic intracellular cell biological processes is the dearth of methodologies that are sufficiently fast and specific to perturb intracellular protein activities. We previously developed a light-sensitive variant of the microtubule plus end tracking protein EB1 by inserting a blue light-controlled protein dimerization module between functional domains. Here, we describe an advanced method to replace endogenous EB1 with this light-sensitive variant in a single genome editing step, thereby enabling this approach in human induced pluripotent stem cells (hiPSCs) and hiPSC-derived neurons. We demonstrate that acute and local optogenetic EB1 inactivation in developing cortical neurons induces microtubule depolymerization in the growth cone periphery and subsequent neurite retraction. In addition, advancing growth cones are repelled from areas of blue light exposure. These phenotypes were independent of the neuronal EB1 homolog EB3, revealing a direct dynamic role of EB1-mediated microtubule plus end interactions in neuron morphogenesis and neurite guidance.
Data availability
Raw data have been deposited to Dryad: doi:10.7272/Q6CF9NC5
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Data for: Growth cone advance requires EB1 as revealed by genomic replacement with a light-sensitive variantDryad Digital Repository, doi:10.7272/Q6CF9NC5.
Article and author information
Author details
Funding
National Cancer Institute (R21 CA224194)
- Torsten Wittmann
National Institute of Neurological Disorders and Stroke (R01 NS107480)
- Torsten Wittmann
National Institutes of Health (S10 RR026758)
- Torsten Wittmann
National Institutes of Health (S10 OD028611)
- Torsten Wittmann
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Copyright
© 2023, Dema 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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Growth cone advance requires EB1 as revealed by genomic replacement with a light-sensitive variant
eLife 12:e84143.
https://doi.org/10.7554/eLife.84143
