Figure 4: Transition through progenitor cell stages demarcates developing rosettes as VZ and SVZ equivalents.

(a) Differential expression levels for selected genes that are most differentially expressed between HES5+ and HES5− cells in a stage-specific manner. Selected gene members are indicated on the left, developmental stages are indicated on the bottom, and gene categories classified by stage are indicated on the right. Values plotted on the heatmap represent ratios of expression levels relative to ES cells. (b) Relative expression levels (z-scores) based on microarray expression data for the entire differentiation time course for selected germinal zone marker genes. Relative expression levels are shown for HES5+ (top) and HES5− (bottom) samples separately. Genes are ordered from VZ to SVZ and from neurogenic to gliogenic markers. Individual qPCR analyses for all genes tested at all stages are shown in Supplementary Fig. 6c. Note that the apparently high GFAP expression in HES5+ cells at the L-RG stage has in fact low absolute expression values, and only appear high relatively to expression in other stages (all stages per each gene are normalized to 1; that is, highest red intensity). To compare GFAP transcript levels during proliferation and serum induced astrocytic differentiation, see Figs 5d and 3e, respectively. (c) Combined HES5::eGFP reporter expression and immunostainings of neural stem/progenitor markers, RG markers, and proliferation markers throughout the progression period. From top: PAX6 marking the VZ and TBR2 marking the SVZ are shown. Middle: CUX1 marking SVZ is shown. Bottom: the (mainly) SVZ marker POU3F2 is shown. Scale bar: 50 μm (valid for all images in c). (d) High-power magnification of E-RG and M-RG images shown in c. Dashed lines demarcate proposed VZ, SVZ and OSVZ regions, containing apical RG, INPs and basal RG, respectively. Scale bar: 25 μm (valid for all images in d).