Fig. 2. Transcriptomic changes in murine luminal subpopulations during castration and organ regeneration.

(A) Schematic overview of the castration/regeneration cycle with experimental time-points. (B) Scatterplots of the L1 (x axis) and L2 (y axis) intact signature score (z score) for each cell (dot) assigned to L1 (red) or L2 (blue) at each time point (panel). Dot color intensity is scaled by the strength of their classifier assignment probability for their assigned class (colorbar). (C) Similar transcriptional states of L1 and L2 during castration. PHATE graph of scRNA-seq profiles from luminal cells, colored by time point (left panel) or L1, L2, L3 based on expression profiles in T0 (right panel). L1 cells undergo the most substantial transcriptional changes. On castration Day 28 (dark green, left panel) and regeneration day 1 (light green, left panel) L1 are co-embedded with L2 cells (orange, right panel). (D) Rapid entry of L1 and L2 cells into the cell cycle during regeneration. Each plot shows the distribution of Mki67 mRNA expression (y axis) throughout the C/R cycle (x axis) for L1, L2 and L3 cells. Fraction of cells with Mki67 expression detected is noted on top. *expression is significantly different from intact (T0) (Bonferroni corrected P < 0.05, one sided Wilcoxon rank-sum test) **designates fold change of 1.5 or greater, and AUC of 0.65. (E) IF staining of Ki67 in the anterior lobe at regeneration day 2. Left: Low magnification showing proximal and distal regions. Right: representative higher magnification (20x) of proximal and distal regions. Ki67 (red), Ck8 (Green), Ck5 (white) and DAPI (purple). Scale bars: 200 or 50μm as labeled.