Skip to main content

Thank you for visiting You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

Directional Notch trafficking in Sara endosomes during asymmetric cell division in the spinal cord


Asymmetric division of neural precursor cells contributes to the generation of a variety of neuronal types. Asymmetric division is mediated by the asymmetric inheritance of fate determinants by the two daughter cells1,2. In vertebrates, asymmetric fate determinants, such as Par3 and Mib, are only now starting to be identified3,4. Here we show that, during mitosis of neural precursors in zebrafish, directional trafficking of Sara endosomes to one of the daughters can function as such a determinant. In asymmetric lineages, where one daughter cell becomes a neuron (n cell) whereas the other divides again to give rise to two neurons (p cell), we found that the daughter that inherits most of the Sara endosomes acquires the p fate. Sara endosomes carry an endocytosed pool of the Notch ligand DeltaD, which is thereby itself distributed asymmetrically. Sara and Notch are both essential for cell fate assignation within asymmetric lineages. Therefore, the Sara endosome system determines the fate decision between neuronal differentiation and mitosis in asymmetric lineages and thereby contributes to controlling the number of neural precursors and differentiated neurons during neurogenesis in a vertebrate.

This is a preview of subscription content, access via your institution

Relevant articles

Open Access articles citing this article.

Access options

Rent or buy this article

Get just this article for as long as you need it


Prices may be subject to local taxes which are calculated during checkout

Figure 1: Neural precursor lineages and asymmetric Sara endosome segregation during neural precursor division.
Figure 2: Targeting of Sara endosomes during neural precursor division forecasts the p fate in n p lineages.
Figure 3: Sara endosomes are required for p cell specification independently of Par3.
Figure 4: DeltaD is a cargo of asymmetric Sara endosomes.


  1. Neumuller, R. A. & Knoblich, J. A. Dividing cellular asymmetry: asymmetric cell division and its implications for stem cells and cancer. Genes Dev. 23, 2675–2699 (2009).

    Article  PubMed  PubMed Central  Google Scholar 

  2. Gonczy, P. Mechanisms of asymmetric cell division: flies and worms pave the way. Nat. Rev. Mol. Cell Biol. 9, 355–366 (2008).

    Article  PubMed  Google Scholar 

  3. Alexandre, P., Reugels, A. M., Barker, D., Blanc, E. & Clarke, J. D. Neurons derive from the more apical daughter in asymmetric divisions in the zebrafish neural tube. Nat. Neurosci. 13, 673–679 (2010).

    Article  CAS  PubMed  Google Scholar 

  4. Dong, Z., Yang, N., Yeo, S. Y., Chitnis, A. & Guo, S. Intralineage directional Notch signaling regulates self-renewal and differentiation of asymmetrically dividing radial glia. Neuron 74, 65–78 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  5. Ando, R., Hama, H., Yamamoto-Hino, M., Mizuno, H. & Miyawaki, A. An optical marker based on the UV-induced green-to-red photoconversion of a fluorescent protein. Proc. Natl Acad. Sci. USA 99, 12651–12656 (2002).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  6. Papan, C. & Campos-Ortega, J. A. Region-specific cell clones in the developing spinal cord of the zebrafish. Dev. Genes Evol. 209, 135–144 (1999).

    Article  CAS  PubMed  Google Scholar 

  7. Coumailleau, F., Furthauer, M., Knoblich, J. A. & Gonzalez-Gaitan, M. Directional delta and Notch trafficking in Sara endosomes during asymmetric cell division. Nature 458, 1051–1055 (2009).

    Article  CAS  PubMed  Google Scholar 

  8. Montagne, C. & Gonzalez-Gaitan, M. Sara endosomes and the asymmetric division of intestinal stem cells. Development 141, 2014–2023 (2014).

    Article  CAS  PubMed  Google Scholar 

  9. Kutateladze, T. G. Phosphatidylinositol 3-phosphate recognition and membrane docking by the FYVE domain. Biochim. Biophys. Acta 1761, 868–877 (2006).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  10. Blatner, N. R. et al. The molecular basis of the differential subcellular localization of FYVE domains. J. Biol. Chem. 279, 53818–53827 (2004).

    Article  CAS  PubMed  Google Scholar 

  11. Hayakawa, A. et al. Structural basis for endosomal targeting by FYVE domains. J. Biol. Chem. 279, 5958–5966 (2004).

    Article  CAS  PubMed  Google Scholar 

  12. Mao, Y. et al. Crystal structure of the VHS and FYVE tandem domains of Hrs, a protein involved in membrane trafficking and signal transduction. Cell 100, 447–456 (2000).

    Article  CAS  PubMed  Google Scholar 

  13. Itoh, F. et al. The FYVE domain in Smad anchor for receptor activation (SARA) is sufficient for localization of SARA in early endosomes and regulates TGF-β/Smad signalling. Genes Cells 7, 321–331 (2002).

    Article  CAS  PubMed  Google Scholar 

  14. Ulrich, F. et al. Wnt11 functions in gastrulation by controlling cell cohesion through Rab5c and E-cadherin. Dev. Cell 9, 555–564 (2005).

    Article  CAS  PubMed  Google Scholar 

  15. Kok, F. O. et al. Reverse genetic screening reveals poor correlation between Morpholino-Induced and mutant phenotypes in Zebrafish. Dev. Cell 32, 97–108 (2014).

    Article  PubMed  PubMed Central  Google Scholar 

  16. Kawaguchi, D., Furutachi, S., Kawai, H., Hozumi, K. & Gotoh, Y. Dll1 maintains quiescence of adult neural stem cells and segregates asymmetrically during mitosis. Nat. Commun. 4, 1880 (2013).

    Article  PubMed  Google Scholar 

  17. Geling, A., Steiner, H., Willem, M., Bally-Cuif, L. & Haass, C. A γ-secretase inhibitor blocks Notch signaling in vivo and causes a severe neurogenic phenotype in zebrafish. EMBO Rep. 3, 688–694 (2002).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  18. Dovey, H. F. et al. Functional γ-secretase inhibitors reduce β-amyloid peptide levels in brain. J. Neurochem. 76, 173–181 (2001).

    Article  CAS  PubMed  Google Scholar 

  19. Appel, B., Givan, L. A. & Eisen, J. S. Delta-Notch signaling and lateral inhibition in zebrafish spinal cord development. BMC Dev. Biol. 1, 13 (2001).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  20. Haddon, C. et al. Multiple delta genes and lateral inhibition in zebrafish primary neurogenesis. Development 125, 359–370 (1998).

    CAS  PubMed  Google Scholar 

  21. Lewis, J. Neurogenic genes and vertebrate neurogenesis. Curr. Opin. Neurobiol. 6, 3–10 (1996).

    Article  CAS  PubMed  Google Scholar 

  22. Itoh, M. et al. Mind bomb is a ubiquitin ligase that is essential for efficient activation of Notch signaling by Delta. Dev. Cell 4, 67–82 (2003).

    Article  CAS  PubMed  Google Scholar 

  23. Scheer, N. & Campos-Ortega, J. A. Use of the Gal4-UAS technique for targeted gene expression in the zebrafish. Mech. Dev. 80, 153–158 (1999).

    Article  CAS  PubMed  Google Scholar 

  24. Scheer, N., Groth, A., Hans, S. & Campos-Ortega, J. A. An instructive function for Notch in promoting gliogenesis in the zebrafish retina. Development 128, 1099–1107 (2001).

    CAS  PubMed  Google Scholar 

  25. Scheer, N., Riedl, I., Warren, J. T., Kuwada, J. Y. & Campos-Ortega, J. A. A quantitative analysis of the kinetics of Gal4 activator and effector gene expression in the zebrafish. Mech. Dev. 112, 9–14 (2002).

    Article  CAS  PubMed  Google Scholar 

  26. Matsuda, M. & Chitnis, A. B. Interaction with Notch determines endocytosis of specific Delta ligands in zebrafish neural tissue. Development 136, 197–206 (2009).

    Article  CAS  PubMed  Google Scholar 

  27. Westerfield, M. The Zebrafish Book: A Guide for The Laboratory Use of Zebrafish (Danio Rerio) (Univ. Oregon Press, 2000).

    Google Scholar 

  28. Kimmel, C. B., Ballard, W. W., Kimmel, S. R., Ullmann, B. & Schilling, T. F. Stages of embryonic development of the zebrafish. Dev. Dyn. 203, 253–310 (1995).

    Article  CAS  PubMed  Google Scholar 

  29. Montero, J. A. et al. Shield formation at the onset of zebrafish gastrulation. Development 132, 1187–1198 (2005).

    Article  CAS  PubMed  Google Scholar 

Download references


We are very grateful to O. Schaad, C. Seum, C. Alliod, E. Derivery and F. Schütz for technical help, A. Reugels and M. Brand for the Par3–GFP and Palmitoylated–RFP constructs, respectively, and A. Oates, C. Gonzalez and J. Bertrand for critical reading of the manuscript. We also thank R. Finazzi, S. Borgers and O. Seum for fish care. C.C. was supported by Fundação para a Ciência e Tecnologia (SFRH/BD/15210/2004) and ONCASYM and the PGDB PhD programme. M.F. was supported by HFSP, CNRS and INSERM. This work was supported by the Département de l’Instruction Publique of the Canton of Geneva, SNSF, the SystemsX epiPhysX grant, ERC advanced grants (Sara and Morphogen), the NCCR Frontiers in Genetics and Chemical Biology programs and the Polish–Swiss research program to M.G-G.

Author information

Authors and Affiliations



S.K., C.C. and I.C. designed and carried out the experiments, generated and interpreted the data and prepared the manuscript. M.F. designed and carried out the iDeltaD antibody uptake, contributed to the Par3 analysis, interpreted the data and contributed to the supervision of the project. M.G-G. supervised the project, interpreted the data and wrote the manuscript.

Corresponding authors

Correspondence to Irinka Castanon or Marcos González-Gaitán.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Integrated supplementary information

Supplementary Figure 1 Neural progenitor lineages at different developmental stages.

ab, Schemes of lineage tracing by Kaede photoconversion of either neural precursor mother cells (a) or daughter cells (b). Single neural precursor cells (a) or one of their daughters (b) expressing Kaede in the neural plate (c), neural rod (d) and neural tube (e) were photoconverted from green to red. When we photoconverted mothers or daughters at plate, rod or tube stage, the second division of the daughters in the case of np and pp lineages occurred late during tube stage. Therefore, the photoconverted mothers in early tube stage are not the daughters of neural precursors that divided at plate or rod stage. ce, Neural precursor mother cell photoconversion at plate stage shows that 75% of the lineages are pp, while 25% corresponds to the nn lineage (c). The frequencies change at rod and tube stages (d,e). At rod stage, 53% corresponds to pp lineage, while 35% and 12% correspond to the np and nn lineages, respectively (d). At tube stage, the frequencies obtained were 18% for the pp lineage, 58% for the np and 24% for the nn lineages (e). From the observed lineages through photoconversions of mother cells at plate (c), rod (d) and tube (e) stages, the expected amount of 1-cell and 2- or more-cell clones after photoconversion of the daughter cells can be calculated (“Expected”). The total expected frequencies correlate with the “Observed” frequencies. It is worth noting that only the sum of all lineages was given for the observed frequency since our assay does not allow distinguish between the three different lineages.

Supplementary Figure 2 sara ubiquitous expression, endosomal localization and lack of correlation between Sara endosome asymmetry and lineage type.

a, sara mRNA expression pattern during embryogenesis. Control: sara sense probe. Scale bar: 250 μm. b, Colocalization between Venus-Sara and CFP-Rab5-positive early endosomes (arrowheads) in the spinal cord. Right panels, magnification of the boxes in the left panels. c,e, Cell frequency distribution of YFPRab5c (c) and YFP-Rab7-positive endosome ratios between the two daughters during neural precursor divisions (e). Both Rab5- and Rab7-positive endosomes segregate symmetrically. d,f, Confocal images of dividing neural precursors showing symmetric distribution of either YFP-Rab5c (d) or YFP-Rab7 endosomes (f). Cell profile counterstained by Gap43-CFP. g, Percentage of cells with an endosomal ratio higher than 1.5 for Venus-Sara-, YFP-Rab5c-, YFP-Rab11a-, and YFP-Rab7-positive endosomes. h, Sara endosome ratio between the daughter cells and the type of lineages generated subsequently (data set from Fig. 2a). Statistical analyses (t-test) comparing the averages of endosome asymmetry in the different types of lineages show that there is not correlation between Sara endosome asymmetry and the type of lineage. Three outliers with abnormally high levels of asymmetry (6, 7 and 12 fold) were excluded from the statistics. i, Correlation between Sara endosome ratio and n vs. p fate in dividing neural precursors expressing Rab5Q81L. While there is no correlation between Sara endosome ratio and n vs. p fate acquisition in wildtype (h), Sara endosome asymmetry is much higher in neural precursors expressing Rab5Q81L and almost all lineages are np, under this condition. Grey area, Sara endosome asymmetry range in wildtype as seen in (h). Dashed lines correspond to the 1.5 fold threshold, which defines “asymmetric dispatch” in this work. j, Confocal image of neural precursors expressing YFP-Rab5Q81L and mRFP-Sara showing Sara endosomes as few large vesicular structures (white arrows). k, Scheme and corresponding confocal images of daughter photoconversion during division of precursor cells expressing YFP-Rab5Q81L/Sara and Kaede. Either the daughter cell that inherits fewer (upper panel) or more YFP-Rab5Q81L/Sara endosomes (lower panel) is photoconverted (middle panels: right panel red channel; left, merge). The fate of the photoconverted daughter is determined 48 h later by the number of cells in the red lineages (red cells; most right panel). l, Percentage of daughters acquiring p vs. n fate among those inheriting Rab5Q81L/Sara or not as in k. Scale bars: 5 μm.

Supplementary Figure 3 Sara endosome segregation correlates with n versus p fate.

ac, Schemes showing the frequency distribution of lineages and the correlation between Sara endosome segregation and mitotic fate at plate (a), rod (b) and tube (c) stages. Frequencies of the nn, np and pp lineages when the neural progenitor mother cell was photoconverted are shown as in Supplementary Fig. 1. The frequency of the different lineages was estimated (“expected”) according to the lineages observed when the neural precursor mother cell was photoconverted. The frequencies observed when either the daughter that inherited more Sara (Sara+) or the daughter that inherited less Sara endosomes (Sara−) was photoconverted do correlate with the expected frequencies.

Supplementary Figure 4 Sara endosomes, Par3 and Mindbomb.

a, Confocal images showing the normal apical localization of Sara endosomes in par3 morphants (cf. Fig. 1k–m). Scale bar: 20 μm. b,c Normal Sara endosome asymmetric segregation during neural precursor division in par3 morphants. Confocal image (b; dotted white line marks cell outline) and frequencies of cells with Sara endosome ratio above 1.5 in wildtype and par3 morphant embryos (c; t-test; p > 0.7). de, Apical localization of Par3-GFP in both wildtype (d) and sara morphant (e). Scale bar: 20 μm. f,g, Asymmetric segregation of Par3-GFP into the two daughter cells during the division of a wild type (f) and sara morphant neural progenitor cell (g). Dotted white line marks cell outline of dividing cell. In a,b and f, cell profile is counterstained by palmitoylated-RFP. h, Cell frequency distribution of apical Par3-GFP ratio between the two daughters during neural precursor divisions (e). Fisher’s Exact Test; p > 0.1. il, Specificity of the Delta antibody uptake assay. i,j, Confocal images showing the spinal cord of embryos injected with either a DeltaD antibody labeled with Zenon 488 (j) or with Zenon 488 alone (i). k,l, Confocal images showing DeltaD staining after the iDeltaD internalization assay in wildtype (k) and mib morphant embryos (l). Note the plasma membrane staining in mib morphants. Scale bar: 20 μm. m, Confocal images showing colocalization between Mib-GFP intracellular vesicular structures and Sara-positive endosomes (white arrowheads) in the spinal cord. Scale bar: 20 μm. n, Confocal images showing that mRFP-Sara endosomes and Mib-GFP segregate preferentially into the same daughter cell (arrowhead). Dotted white line marks cell outline of dividing cell. o, Confocal images showing the asymmetric segregation of Venus-Sara endosomes during neural progenitor in notch1a/notch3 double morphants, and deltad and mib morphants. Dotted white line marks cell outline of dividing cell. p, Venus-Sara endosome ratio between the two daughter cells after neural progenitor division in wildtype and different Notch pathway blockage conditions. Sara endosomes asymmetric segregation (Sara endosome ratio > 1.5) is unaffected in notch1a/notch3 double morphants, and deltad and mib single morphants compared with wild type. Scale bar: 5 μm, unless stated otherwise.

Supplementary information

Supplementary Information

Supplementary Information (PDF 3420 kb)

Segregation of Sara endosomes in a dividing neural progenitor cell.

Sara endosomes (marked by Venus-Sara, depicted in green) are asymmetrically segregated into the two daughter cells. GAP43-CFP (depicted in red) outlines the membrane of the dividing neural precursor cell. (MOV 1753 kb)

Dynamics of Sara endosomes and their DeltaD cargo in dividing neural progenitor cells.

Sara endosomes (marked by mRFP-Sara, depicted in green) transport endogenous internalized DeltaD ligands (iDeltaD, red). GAP43-GFP (depicted in magenta) outlines the membrane of both the dividing neural precursor and neighboring neuro-epithelial cells. (AVI 174 kb)

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Kressmann, S., Campos, C., Castanon, I. et al. Directional Notch trafficking in Sara endosomes during asymmetric cell division in the spinal cord. Nat Cell Biol 17, 333–339 (2015).

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI:

This article is cited by


Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing