Fig. 1

Klf4 down-regulation enhances neurogenesis in vivo and in vitro. a (Upper) Fixed coronal sections from E11.5 (left) or E14.5 (right) mouse forebrain stained with antibodies against Tuj1 (red), Tbr1 (green), or Map2 (green). Nuclear staining is shown by DAPI (blue). (Lower) Quantification of data shown above (E11.5: Klf4 ^fl/+ (WT), n = 4–8, Klf4 conditional knockout (cKO), n = 3–8, and E14.5: Klf4^fl/+ (WT), n = 3–9, Klf4 cKO, n = 4–6). Scale bars, 50 μm. b (Left) Analysis of Pax6 in sections described in a. Scale bar, 50 μm. (Right) Quantification of data shown on the left (E11.5: WT, n = 5, Klf4 cKO, n = 5, and E14.5: WT, n = 5, Klf4 cKO, n = 3). c (Upper) Immunostaining with Tuj1 or Map2 (both red) antibodies in WT and Klf4 cKO NPCs in undifferentiation and differentiation conditions. Nuclear staining is shown by DAPI (blue). Scale bars, 50 μm. (Lower) Quantification of Tuj1-positive (WT-Un, n = 4, WT-D2, n = 4, WT-D4, n = 6, Klf4 cKO-Un, n = 4, Klf4 cKO-D2, n = 3, Klf4 cKO-D4, n = 8) and Map2-positive (WT-Un, n = 5, WT-D2, n = 5, WT-D4, n = 3, Klf4 cKO-Un, n = 4, Klf4 cKO-D2, n = 4, Klf4 cKO-D4, n = 5) cells in panels above. d Immunostaining with Ki67 (red) and Nestin (green) antibodies in WT and Klf4 cKO NPCs. DAPI (blue). Scale bars, 25 μm. (Right) Quantification of Ki67/Nestin double-positive cells in analysis shown at left (n = 4). e Single WT and Klf4 cKO NPCs were separated by serial dilution and neurosphere formation was induced for 7 days in vitro (DIV). Relative size of primary spheres grown to 7 DIV was quantified by Image J Software. Scale bars, blue (100 pixel), red (200 pixel). f qPCR analysis of indicated mRNAs in WT and Klf4 cKO NPCs. Values correspond to mean ± SD. ANOVA tests were performed to calculate significance (*P < 0.01, **P < 0.001, ***P < 0.0001). See also Supplementary Fig. 1a–f and Supplementary Table 4. Un undifferentiationn condition, D differentiation condition