Figure 4. Spatial Distribution of Notch and Delta in reactive RG after the injury.

(A–D) t-SNE plots (A and C) and violin plots (B and D) showing deltaA is mainly expressed in pRG-S (cluster 3) and pRG-G2 (cluster 4) RG, whereas the expression of her4.1, a target gene of Notch signaling, is down-regulated in pRG-S (cluster 3) and pRG-G2 RG (cluster 4). (E) Correlation distance matrix showing deltaA exhibits the high correlation with pcna but the low correlation with her4.1 and her4.2 in pRG-S RG (cluster 3, red dots in the t-SNE plot at the top left, each dot represents a single cell). (F–G₃) Tg(Tp1:EGFP) (green) and PCNA (red) immunofluorescences show that PCNA⁺ proliferative RG (open white arrowheads, red cells) and EGFP⁺ RG (green cells) are exclusive in the injured optic tectum (F–F₃). (G–G₃) The representative images of the uninjured optic tectum. (H) Quantification of EGFP⁻/PCNA⁺ and EGFP⁺/PCNA⁺ RG in (F) showing ~82% PCNA⁺ RG are EGFP⁻. Most of the PCNA⁺ proliferative RG have low Notch activity (97/119 cells in 6 sections from 4 fish). (I–I₅) Specific expression of deltaA in PCNA⁺ tectal RG (white arrowheads, white dashed circles) at 3 dpi is confirmed by in situ hybridization. The open white arrowheads indicate a GFP⁻/PCNA⁺/deltaA⁻ non-RG proliferative cells. (J) Quantification of PCNA⁺/deltaA⁺ and PCNA⁺/deltaA⁻ RG in (I) showing ~81% PCNA⁺ RG express deltaA. (60/74 cells in 10 sections from 5 fish). White dashed lines represent the tectal ventricle boundary. RG, radial glia; PGZ, periventricular gray zone; TS, torus semicircularis. Scale bars, 30 μm.