Extended Data Fig. 6: RXRα responds to corpses.
From: Stem cells tightly regulate dead cell clearance to maintain tissue fitness
a, Representative immunofluorescence images for data in Fig. 3a. Nuclear RXRα and RARγ co-occur (open arrowheads) near unengulfed dying cells (closed arrowheads) in mid- late catagen hair follicles (HFs) (white dashed outline). Insets, higher magnification view of yellow boxed regions to left. Scale bar 20 μm. 10-20 HFs per mouse, n = 3 animals per hair cycle stage. b, Representative immunofluorescence images for FACS-quantified data in Fig. 3b. Diptheria toxin A (DTA) expression in Sox9CreER+ mice creates TUNEL+ corpses (white arrowhead) in quiescent HFs (white dashed outline), which induces nuclear RXRα. Scale bar 20 μm. 10-20 HFs analyzed per mouse, n = 2 (Sox9CreER-) n = 4 (Sox9CreER=) animals. c, Quantifications of RXRα intensity (left) and TAM-family expression (right) for FACS-isolated hair follicle stem cells (HFSCs) from K14rtTA+ mice induced to overexpress RXRα (RFP+) at the end of catagen and followed to early anagen (AnaII-III), using the experimental strategy shown in Fig. 2b. n = 3 mice. d, Left, Experimental schematic. Right, FACS-based quantifications of the percentage of total HFSCs which are TAM-family+, lysosomehigh (middle) or contain corpses (right) 4hrs post corpse addition. Inhibition of RXR-family activity by UVI3003 or HX531. n = 6 (Media), n = 9 (Corpses+VEH, Corpses+UVI3003, Corpses+HX531) experimental replicates. e, Bulk RNA-squencing of Rxra wild type (WT) versus Rxra conditional knockout (cKO) HFSCs in media or upon corpse exposure. Left, Heatmap of differentially expressed genes (full list in Supplementary Table 4); Z-scaled expression scores of genes (rows; blue, downregulated; red, upregulated) by experimental replicates (columns; n = 2 media and n = 3 corpse-exposed WT, n = 3 corpse-exposed cKO). Right, Expression of phagocytic genes; TPM, transcript per million. n = 2 media and n = 4 corpse-exposed WT, n = 4 corpse-exposed cKO. f, FACS-based quantification of TAM-family expression levels 4hrs post corpse addition to RXRα wild type (WT) or conditional knockout (cKO) with or without RAR-family inhibition (+AGN [AGN193109] condition). Quantification represents one of two independent experiments. Each dot represents data from one experimental replicate: Media condition (n = 3 WT, n = 3 cKO), for corpses: +VEH (n = 6 WT, n = 6 cKO) and +AGN (n = 3 WT, n = 3 cKO). Quantifications, pairwise independent Student’s t-tests (2-sided), p-values indicated. n.s. not significant (p > 0.05). Data as box-and-whisker plots; box: first-to-third quartiles and median, whiskers 1.5× inter-quartile range. More details on statistics and reproducibility can be found in the Methods section.
