Abstract
Neuronal polarization occurs shortly after mitosis. In neurons differentiating in vitro, axon formation follows the segregation of growth-promoting activities to only one of the multiple neurites that form after mitosis1,2. It is unresolved whether such spatial restriction makes use of an intrinsic program, like during C. elegans embryo polarization3, or is extrinsic and cue-mediated, as in migratory cells4. Here we show that in hippocampal neurons in vitro, the axon consistently arises from the neurite that develops first after mitosis. Centrosomes, the Golgi apparatus and endosomes cluster together close to the area where the first neurite will form, which is in turn opposite from the plane of the last mitotic division. We show that the polarized activities of these organelles are necessary and sufficient for neuronal polarization: (1) polarized microtubule polymerization and membrane transport precedes first neurite formation, (2) neurons with more than one centrosome sprout more than one axon and (3) suppression of centrosome-mediated functions precludes polarization. We conclude that asymmetric centrosome-mediated dynamics in the early post-mitotic stage instruct neuronal polarity, implying that pre-mitotic mechanisms with a role in division orientation may in turn participate in this event.
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Acknowledgements
We would like to thank E. Cassin and B. Hellias for the hippocampal neurons, L. Ciapponi and the Bloomington Stock Center for fly strains, and C. Gonzalez for advice with the neuroblast division experiment. F.C.dA. is supported by an EMBO long-term fellowship. J.S.D.S. was supported by an FCT/PRAXIS XXI scholarship (Portuguese Ministry of Science and Technology). F.F. was supported by an Alexander von Humboldt scholarship. Part of this work is supported by an EU Contract grant (APOPIS) to C.G.D.Author Contributions F.C.d.A. and G.P. were responsible for all in vitro experiments in mammalian and insect neurons, respectively. J.S.D.S. supervised the hippocampal neuron work. P.G.C. helped with the in situ work.
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Supplementary information
Supplementary Figure S1
The first sprout contains neuronal polarity information. (PDF 229 kb)
Supplementary Figure S2
Organelle polarization marks the site of neuronal polarity. (PDF 496 kb)
Supplementary Figure S3
Drosophila neurons' polarization in vitro and in situ correlates with plane of mitotic division and the localization of the centrosomes. (PDF 592 kb)
Supplementary Figure S4
Centrosomal-mediated polarized microtubule and membrane activities precede morphological polarization in vitro and in situ. (PDF 177 kb)
Supplementary Figure S5
Pharmacological disruption of microtubule polymerization and membrane trafficking prevents morphological polarization. (PDF 99 kb)
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de Anda, F., Pollarolo, G., Da Silva, J. et al. Centrosome localization determines neuronal polarity. Nature 436, 704–708 (2005). https://doi.org/10.1038/nature03811
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DOI: https://doi.org/10.1038/nature03811
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