In this region, the Sox2+ NPC compartment displayed a 2- to 3-fold increased expression of N-cadherin and the number of differentiated neurons formed was reduced by ∼38% ( Figures 7N–7Q, 7R–7U, and 7AE). NeuN+ neurons were also interspersed within the VZ comparable to the defects seen in the Foxp4LacZ/LacZ mutant spinal cord and the chick Foxp2/4 double-knockdown experiments ( Figures 2L–2O and 7H, 7L, 7Q, and 7U). Foxp4 mutant forebrains frequently lacked lateral ventricles, with medial and

lateral cortices displaying Docetaxel cell line unusually contiguous contacts along their apical membranes, resulting in convolution and invagination of the neuroepithelium ( Figures 7N–7P, 7R–7T, S8S–S8U, and S8X–S8Z). Sonic hedgehog, whose loss of function is commonly associated with holoprosencephaly, was nevertheless present in all embryos analyzed, and the dorsoventral position of different NPC subtypes was generally intact ( Figures S8U, S8Z, and S8AA–S8AD). This feature of the Foxp4 mutants is notably similar to the phenotype of mice in which AJ components such as Cdc42 have been inactivated ( Cappello et al., 2006 and Chen et al., 2006). Finally, we misexpressed Foxp4 in the developing

cortex and found that it potently suppressed the expression of N-cadherin, Sox2, β-catenin, and other components of AJs, much like the effects seen in the chick spinal cord 3MA (Figures 7V–AC and 7AF). Consequently, the number of Tbr2+ neurons was elevated ∼2-fold and formed ectopic clusters within and adjacent to the VZ (Figures 7Y, 7AC,

and 7AF). Collectively, these results suggest that the suppressive effects of Foxp4 and Foxp2 on NPC adhesion might play a more general role in regulating progenitor maintenance throughout the developing CNS. The polarized organization and proliferation of neuroepithelial progenitors depends on the formation of AJs between NPCs. These contacts act as a self-supporting stem cell niche to maintain cells in an undifferentiated state. Our results identify Foxp4 and Foxp2 as components of a gene regulatory network that balances the assembly and disassembly of AJs to respectively promote NPC proliferation and differentiation. In the normal also course of MN development, Foxp4 levels increase as NPCs shed their adhesive contacts and migrate away from the VZ (Figure 8A). When Foxp proteins are artificially elevated, N-cadherin and Sox2 expression are suppressed, leading to the dissolution of AJs, cytoplasmic distribution of Numb, and ectopic neurogenesis within the VZ (Figure 8B). In contrast, the combined loss of Foxp2 and Foxp4 increases N-cadherin expression and retains NPCs in an undifferentiated, neuroepithelial state (Figure 8C). Together, these findings provide important insights into the developmental programs that influence how NPCs interact with themselves and their environment to regulate the size and shape of the nervous system.