EGFP and NKX2-1 protein expression in control brains co-localized in a subset of cells derived from the MGE. EGFP/NKX2-1+ cells were observed in the MGE VZ and SVZ progenitors and a subset of their derived neurons, including the globus pallidus, and striatal interneurons (Xu et al., 2008; Figure 1G solid arrowheads), while interneurons migrating to the cerebral cortex showed little to no NKX2-1 protein expression (Figure 1G open arrowheads). In mutant brains however, most if not all EGFP labeled cells had detectable levels of NKX2-1 protein, with many cells strongly co-expressing NKX2-1 and EGFP in the LGE MZ, and in a region lateral to the globus pallidus (Figure 1G’ solid arrowheads). Thus, Zfhx1b mutants had a defect in their ability to repress Nkx2-1 RNA and protein expression, concomitant with failure of MGE-derived migration to the cerebral cortex. While Zfhx1b was required to repress Nkx2-1 expression, we did not find evidence that Nkx2-1 regulated Zfhx1b expression; this conclusion was based on in situ hybridization analysis of Zfhx1b expression in mice lacking Nkx2-1 in newly born MGE neurons at E15.5 (Nkx2-1 conditional mutant with Dlx5/6-Cre)(Figure S6).
