Nuclear factor one B regulates neural stem cell differentiation and axonal projection of corticofugal neurons

Abstract

During development of the cerebral cortex, neural stem cells divide to expand the progenitor pool and generate basal progenitors, outer radial glia and cortical neurons. As these newly born neurons differentiate, they must properly migrate toward their final destination in the cortical plate, project axons to appropriate targets, and develop dendrites. However, a complete understanding of the precise genetic mechanisms regulating these steps is lacking. Here we show that a member of the nuclear factor one (NFI) family of transcription factors, NFIB, is essential for many of these processes in mice. We performed a detailed analysis of NFIB expression during cortical development, and investigated defects in cortical neurogenesis, neuronal migration and differentiation in NfiB −/− brains. We found that NFIB is strongly expressed in radial glia and corticofugal neurons throughout cortical development. However, in NfiB −/− cortices, radial glia failed to generate outer radial glia, subsequently resulting in a loss of late basal progenitors. In addition, corticofugal neurons showed a severe loss of axonal projections, while late-born cortical neurons displayed defects in migration and ectopically expressed the early-born neuronal marker, CTIP2. Furthermore, gene expression analysis, by RNA-sequencing, revealed a misexpression of genes that regulate the cell cycle, neuronal differentiation and migration in NfiB −/− brains. Together these results demonstrate the critical functions of NFIB in regulating cortical development.

Combined allelic dosage of Nfia and Nfib regulates cortical development

“…The individual expression of NFIA and NFIB has been reported in radial glia from E12 in mice (Betancourt et al, 2014;Bunt et al, 2015;Plachez et al, 2008), but their co-expression within these cells has not been determined. To exclude the possibility that knockout of Nfia and Nfib affected different subpopulations of radial glia, we analysed their co-expression at a cellular level in the developing cerebrum.…”

“…To exclude the possibility that knockout of Nfia and Nfib affected different subpopulations of radial glia, we analysed their co-expression at a cellular level in the developing cerebrum. The Nfib locus carries a knock-in β-galactosidase reporter gene substituted into the deleted exon of the Nfib allele as a marker for NFIB expression (Betancourt et al, 2014;Piper et al, 2009;Steele-Perkins et al, 2005). For each stage examined, we validated β-galactosidase and NFIB co-expression (Figure 1) to ensure that we were able to analyse co-expression of NFIA and NFIB using β-galactosidase protein expression as a reporter with fidelity (Figure 2).…”

“…The β-galactosidase gene is knocked into exon 2 of the deleted Nfib allele. Thus, this marker protein acts as a reporter for NFIB expression (Betancourt et al, 2014;Piper et al, 2009;SteelePerkins et al, 2005). At E13 (a), E18 (b) and P40 (c) both proteins were co-expressed throughout the cerebrum.…”

“…Analyses of knockout mouse models suggest that the general underlying cause of these phenotypes is a cellular differentiation defect which affects radial glia, the embryonic neural progenitor cells of the cerebral cortex, which give rise to mature neurons and glia in the cortex (Barry et al, 2008;Betancourt et al, 2014;Gobius et al, 2016;Piper et al, 2009Piper et al, , 2010Piper et al, , 2014. Knockout of either Nfia or Nfib causes the radial glia to remain proliferative and undifferentiated for an extended period of time.…”

“…In the context of the brain, NFIA and NFIB share many common transcriptional downstream targets, including Gfap, Hes1, Hes5, Sox3, Fgf9, Gli3, Id4, Cntn2, Efnb1 and Cdh2 (Barry et al, 2008;Betancourt et al, 2014;Brun et al, 2009;Cebolla and Vallejo, 2006;Harris et al, 2016;Piper et al, 2009Piper et al, , 2010Piper et al, , 2014Wang et al, 2007Wang et al, , 2010. In vitro experiments for targets such as Gfap and Cntn2 have validated their direct regulation by both NFIA and NFIB.…”