Figure 8

Effect of impaired MTH signaling and NICD overexpression on progenitor cell division. (A) At the start of imaging, representative images of the spinal cord of experimental Tg(elav3:LY-mCherry) embryos at 23hpf. Neurons (mCherry) are shown in white. Embryos were injected at the one-cell stage with either CTRLMO or MCT8MO followed by injection at the 16-cell stage in one blastomere with gfp mRNA only or nicd and gfp mRNA. Cells overexpressing NICD (and GFP) are labelled in magenta. Dorsal views of single slices between somite 8-15 are shown, anterior spinal cord up. The scale bar represents 50 µm. (B) Upper panel: Detail of symmetric division originating 2 morphologically similar GFP+ cells. A cell undergoing symmetric mitosis (yellow arrow), and the originating daughter cells (yellow arrowheads); Lower panel: Representative images of asymmetric division. A dividing cell (yellow arrow) originates two daughter cells (yellow arrowheads). Scale bar in B represent 20 µm. In all panels, the spinal cord basal limit is indicated by a dashed yellow line and the pial limit by dotted yellow lines. (C–F) (C) Percentage of analyzed GFP+ cells that underwent division. (D) Distribution of GFP+ dividing cells relative to all GFP+ cells observed in the period from 23-26hpf. χ2 analysis showed differences in the distribution of the number of cells undergoing symmetric or asymmetric divisions amongst experimental groups and CTRLMO (p<0.05). (E) Percentage of a cell undergoing symmetric division. (F) Percentage of cells undergoing asymmetric division. The results in C, D, and F are presented as the mean ± SD; results in D depict the ratio of cell division type in all GFP+ dividing cells analyzed. N =5-6 individuals per group (number of cells evaluated by group: CTRLMO/GFP=56; MCT8MO/GFP=77; CTRLMO/NICD/GFP=93; MCT8MO/NICD/GFP=46); Statistical significance in (C, E, F) was determined by a one-way ANOVA followed by a Holm-Sidak’s multiple comparison post hoc analysis, *p<0.05, **p<0.01, ***p<0.001.