Fig. 6: Gene expression signature of transitional RPCs (C3, C4).

a Genes regulating distinct lineages are expressed in a shared transitional state, namely transitional retinal progenitor cells (RPCs), as demonstrated by a dot plot. Often these genes continue to be expressed in the specific cell lineage they regulate, e.g., Atoh7 and Sox11 in retinal ganglion cells (RGCs) (C5), and Otx2 in photoreceptors (C8, C9). The color scale represents natural-log transformed normalized gene expression levels. b Feature heatmap showing the expression of Atoh7, Neurod1, and Otx2 in the wild-type clusters. The dotted line demarcates the transitional RPCs (C3 and C4). Consistent with the dot plot, all three genes are expressed in transitional RPCs, but Atoh7 is expressed in more transitional RPCs than Neurod1 and Otx2. Whereas Atoh7 trails into all three lineages, Neurod1 and Otx2 only continue to be expressed in photoreceptors. c Percentage of cells expressing Atoh7, Neurog2, Neurod1, and Otx2 in the transitional RPCs (C3, C4) and nascent RGCs (C5) in the wild-type (WT) and Atoh7-null (MT) retinas. d Changes in activities of Atoh7, Neurog2, NeuroD1, and Otx2 as cells progress into distinct lineages. For each gene, the gene activity in each cluster is derived by the expression level of that cluster divided by the mean of all clusters. e Additional genes, including many encoding components of the Notch pathway, are enriched in the transitional RPCs (C3, C4). The color scales represent natural-log transformed normalized gene expression levels. f–h Immunofluorescence staining shows substantial co-expression of Atoh7 or Atoh7HA with Olig2 (f), Otx2 (g), and Foxn4 (h) in RPCs. Note the yellow cells are those expressing both markers in each panel. In the case of Otx2, which is expressed in both RPCs and photoreceptors (g), the co-expression only occurs in the RPCs. Each of the co-staining experiments (f–h) was performed three times with sections from different animals. The scale bar is 25 µm. Source data in (c, d) are provided.