Epigenetic factors (EFs) regulate multiple aspects of cerebral cortex development, including proliferation, neuronal differentiation, laminar fate, and regional identity.
More...Epigenetic factors (EFs) regulate multiple aspects of cerebral cortex development, including proliferation, neuronal differentiation, laminar fate, and regional identity. The same neurodevelopmental processes are also regulated by transcription factors (TFs), notably the Pax6→Tbr2→Tbr1 cascade expressed sequentially in radial glial progenitors, intermediate progenitors, and postmitotic projection neurons, respectively. Here, we studied the EF landscape and its regulation in embryonic mouse neocortex. Microarray and in situ hybridization assays revealed that many EF genes are expressed in specific cortical cell types, such as intermediate progenitors, or in rostrocaudal gradients. Furthermore, many EF genes are directly bound and transcriptionally regulated by Pax6, Tbr2, or Tbr1, as determined by chromatin immunoprecipitation-sequencing and gene expression analysis of TF mutant cortices. The results demonstrated that Pax6, Tbr2, and Tbr1 form a direct feedforward genetic cascade, with direct feedback repression. Results also revealed that each TF regulates multiple EF genes that control DNA methylation, histone marks, chromatin remodeling, and noncoding RNA.
Overall design: Tbr1 knockout (KO), Tbr2 conditional knockout (cKO), and Tbr1/2 double KO/cKO (dKO) mouse embryos were produced as described (Bedogni et al., 2010a; Elsen et al., 2013). The Tbr1/2 double mutants were generated by breeding to combine the necessary alleles (Tbr1/;Tbr22F/2F;Nes11Cre). On E14.5, embryos were harvested, and neocortex was immediately dissected and frozen. Genotypes were determined by PCR 216 of tail DNA. Controls were wild type (+/+) for Tbr1 and non-recombined for Tbr2. Each embryonic neocortex was an independent biological replicate. For each genotype, n = 4 samples of neocortex were analyzed by microarray (Affymetrix ST 1.0). The microarray results were analyzed statistically as described (Elsen et al., 2013). In the current paper, we also analyzed previous microarray data from Tbr1 KO (Bedogni et al., 2010a) and Tbr2 cKO (Elsen et al., 2013) neocortices, designated microarray 1 (MA1); the new microarray data were designated microarray 2 (MA2). Tbr1/2 dKO neocortex was analyzed only in MA2.
Less...