Figure 4

QKI suppresses differentiation of NPCs both in vitro and in vivo. (A,B) Western blot analysis of the overexpression of the three Qki isoforms (A) and the knockdown of the QK protein with the shQki plasmid (B) after transfection for 48 hours. (C) The QKI protein expression dynamics during the differentiation of the NSCs. (D) For the knockdown of QKI with Qki siRNA, the samples were derived from the same experiment, and the gels/blots were processed in parallel. All analyses were normalized to β-actin as a loading control. Uncropped versions of all western blots are shown in Supplementary Figure 3. (E) Sections of the E16.5 forebrains electroporated at E13.5 with plasmids expressing GFP alone or those co-expressing each isoform of Qki, the sh-Scramble (sh-Scr) and sh-Qki plasmid, the QKI5 plus miR-214, or QKI7 plus miR-214. CP, cortical plate; IZ, intermediate zone; SVZ, subventricular zone; VZ, ventricular zone. (F) Quantitative analysis of the E16.5 dorsal forebrains for the distribution of the GFP⁺ cells in E. The GPF⁺ cells in the VZ plus SVZ, IZ and CP of the cerebral cortex were counted to perform a statistical analysis. Error bars show the standard deviation, and the comparisons were performed by Student’s t-test. the statistically significant P values are shown as *(<0.05), **(<0.01) or ***(<0.001). (G,H) Cultured neural stem cells were transfected with the Qki siRNA or scramble control. Two days after transfection, the cells were fixed and stained with neuron marker MAP2. Scale bar: 100 μm. (H) The ratio of MAP2 positive cells in the Qki siRNA (siQKI) and scramble control (siNC) groups (n = 5, p = 0.017). Error bars show the standard error of mean, and the comparisons were performed using Student’s t-test; the statistically significant P values are shown as *(<0.05), **(<0.01) or ***(<0.001).