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Numb and Numbl are required for maintenance of cadherin-based adhesion and polarity of neural progenitors

Nature Neuroscience volume 10, pages 819–827 (2007)Cite this article

Abstract

The polarity and adhesion of radial glial cells (RGCs), which function as progenitors and migrational guides for neurons, are critical for morphogenesis of the cerebral cortex. These characteristics largely depend on cadherin-based adherens junctions, which anchor apical end-feet of adjacent RGCs to each other at the ventricular surface. Here, we show that mouse numb and numb-like are required for maintaining radial glial adherens junctions. Numb accumulates in the apical end-feet, where it localizes to adherens junction–associated vesicles and interacts with cadherins. Numb and Numbl inactivation in RGCs decreases proper basolateral insertion of cadherins and disrupts adherens junctions and polarity, leading to progenitor dispersion and disorganized cortical lamination. Conversely, overexpression of Numb prolongs RGC polarization, in a cadherin-dependent manner, beyond the normal neurogenic period. Thus, by regulating RGC adhesion and polarity, Numb and Numbl are required for the tissue architecture of neurogenic niches and the cerebral cortex.

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Figure 1: Numb is enriched in apical end-feet of interphase RGCs.
Figure 2: Numb localizes along the basolateral membrane and to vesicular structures near adherens junctions of interphase RGCs.
Figure 3: Numb colocalizes and interacts with cadherin-catenin adhesion complex.
Figure 4: Disruption of neuroepithelial integrity, adherens junctions and actin organization in Numb and Numbl dKO VZ.
Figure 5: Altered subcellular localization of cadherins in Numb and Numbl dKO VZ.
Figure 6: Loss of neuroepithelial integrity and adherens junctions in RGCs expressing Numb and Numbl, Cdh1 or Cdh2 shRNAs.
Figure 7: Numb and Numbl promote RGC polarity in a cadherin-dependent manner.
Figure 8: Effects of manipulating Numb and Numbl expression on the sequential generation of neocortical cell types.

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Acknowledgements

We thank M. Pappy for experimental assistance, R. Flavell, M. Grumet, M. Inagaki, R. Medzhitov, J. Miyazaki, A. Nepveu, C. Gottardi, C. Walsh and W. Zhong for reagents, and M. Caplan, K. Herrup, J. LoTurco, A. Louvi and members of the Sestan lab for helpful discussions and comments. This study was supported by grants from the March of Dimes Birth Defects Foundation (FY05–73) and the US National Institutes of Health (HD045481, AG019394, NS047200). M.R.R. was supported by Autism Speaks. K.Y.K. was supported by the Canadian Institutes of Health Research. Y.N.J. and L.Y.J. are Investigators of the Howard Hughes Medical Institute.

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Authors and Affiliations

  1. Department of Neurobiology, Yale University School of Medicine, 333 Cedar Street, New Haven, 06510, Connecticut, USA

    Mladen-Roko Rašin, Valeswara-Rao Gazula, Joshua J Breunig, Kenneth Y Kwan, Matthew B Johnson, Susan Liu-Chen, Pasko Rakic & Nenad Šestan

  2. Departments of Physiology and Biochemistry, Howard Hughes Medical Institute, University of California San Francisco, 1550 4th Street, San Francisco, 94143, California, USA

    Hua-Shun Li, Lily Yeh Jan & Yuh-Nung Jan

  3. Developmental Neurobiology Program, Institute of Molecular Medicine and Genetics, Medical College of Georgia, 1120 15th Street, Augusta, 30912, Georgia, USA

    Hua-Shun Li

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Supplementary information

Supplementary Fig. 1

Numb and Numbl expression in the developing neocortical wall. (PDF 907 kb)

Supplementary Fig. 2

Numb and Cdh1 immunofluorescent staining of the neocortical wall. (PDF 931 kb)

Supplementary Fig. 3

Serial Numb immunogold electron microscopic analysis of E16.5 mitotic RGCs and surrounding apical end-feet of interphase VZ cells. (PDF 1472 kb)

Supplementary Fig. 4

Numbl interacts with Cdh1 in C6-R radial glial-like cells. (PDF 527 kb)

Supplementary Fig. 5

Disruption of neuroepithelial organization and progenitor dispersion in Numb and Numbl dKO embryos. (PDF 1234 kb)

Supplementary Fig. 6

Disorganization of postnatal neurogenesis sites and ectopic neuronal progenitors in the postnatal neocortex of P21 Numb and Numbl dKO mice. (PDF 1212 kb)

Supplementary Fig. 7

Ectopic immature Dcx+ neurons in the postnatal neocortex of Numb and Numbl dKO mice. (PDF 1041 kb)

Supplementary Fig. 8

Disorganized adherens junctions and RGC alignment in the ventricular zone of E16.5 Numb and Numbl dKO embryos. (PDF 1612 kb)

Supplementary Fig. 9

Knocking down Numb, Nubl, Cdh1 and Cdh2 with shRNAs. (PDF 1249 kb)

Supplementary Fig. 10

Forced expression of all Numb isoforms and Numbl promotes radial polarity after the end of neocortical neurogenesis in vivo. (PDF 976 kb)

Supplementary Fig. 11

Astroglial differentiation does not occur prematurely in late Numb and Numbl dKO embryos. (PDF 739 kb)

Supplementary Table 1

Primary antibodies used in this study. (PDF 1157 kb)

Supplementary Table 2

Plasmids and shRNA sequences used in this study. (PDF 294 kb)

Supplementary Methods (PDF 22 kb)

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Rašin, MR., Gazula, VR., Breunig, J. et al. Numb and Numbl are required for maintenance of cadherin-based adhesion and polarity of neural progenitors. Nat Neurosci 10, 819–827 (2007). https://doi.org/10.1038/nn1924

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