A targeted NKX2.1 hesc reporter line enables identification of human basal forebrain derivatives

We have used homologous recombination in hESCs to insert sequences encoding GFP into the NKX2.1 locus, a gene required for normal development of the basal forebrain. Generation of NKX2.1-GFP(+) cells was dependent on the concentration, timing and duration of retinoic acid treatment during differentiation. NKX2.1-GFP(+) progenitors expressed genes characteristic of the basal forebrain, including SHH, DLX1, LHX6 and OLIG2. Time course analysis revealed that NKX2.1-GFP(+) cells could up-regulate FOXG1 expression, implying the existence of a novel pathway for the generation of telencephalic neural derivatives. Further maturation of NKX2.1-GFP(+) cells gave rise to GABA-, TH- and SST-expressing neurons as well as to PDGFRalpha(+) oligodendrocyte precursors. These studies highlight the diversity of cell types that can be generated from human NKX2.1(+) progenitors and demonstrate the utility of NKX2.1(GFP/w) hESCs for investigating human forebrain development and neuronal differentiation.