Fig. 5: Identifying AR loss-induced molecular changes in Gli1-lineage stromal cells.

a–b UMAP plots of stromal fibroblast (FB) and smooth muscle (SM) cell clusters from Hi-Myc:R26^mTmG/+:Gli1^CreER/+ and Hi-Myc:R26^mTmG/+:Ar^L/Y:Gli1^CreER/+. c Gene expression levels of mGFP and Ar in UMAP plots separated by indicated genotypes. Color intensity indicates the scaled expression level in each cell. Blue arrow indicates Ar-expressing cells from FB1 in Hi-Myc:R26^mTmG/+:Gli1^CreER/+. d Violin plots visualizing the expression levels of mGFP, Ar, and insulin-like growth factor binding protein 3 (Igfbp3) in stromal clusters of each genotype. e Gene expression levels of Igfbp3 in UMAP plots of stromal FB and SM cells. Color intensity indicates the scaled expression level in each cell. f Violin plots of mGFP⁺ FB cells. g qPCR analysis of Ar and Igfbp3 expression shown as fold change in FACs-sorted mGFP⁺ cells from prostate tissues of indicated genotypes. h Co-localization of IGFBP3 with either AR, mGFP, or Vimentin (Vim) from indicated prostate tissues. Yellow and pink arrows indicate AR⁺ epithelial and stromal cell, respectively. Blue arrows indicate IGFBP3⁺ stromal cells co-expressing with either mGFP or Vim. Scale bars, 20 μm. i Sp1 Chromatin immunoprecipitation qPCR analysis of the Igfbp3 promoter regions, and negative control (Untr4) shown as percent input. j qPCR analysis of cancer-associated fibroblast (CAF) markers shown as fold change in sort-purified mGFP⁺ cells. k Representative images of triple-IF staining for indicated antibodies on prostate tissue sections from indicated genotypes. Blue arrows indicate cells showing double- or triple-positive with the indicated proteins. Scale bars, 20 μm. l Heatmap displaying gene expression of Ar, Vim, and CAF markers across FB clusters of Hi-Myc:R26^mTmG/+:Gli1^CreER/+. m Heatmap depicting expression of genes associated with indicated pathways. n GSEA showing the enrichment of indicated gene signatures in mGFP⁺Ar⁺ FB1 cells from Hi-Myc:R26^mTmG/+:Gli1^CreER/+ versus mGFP⁺Ar^- FB1 cells from Hi-Myc:R26^mTmG/+:Ar^L/Y:Gli1^CreER/+. See also “Methods” section. NES, normalized enrichment score; FDR, false discovery rate. In g, i, and j, data are represented as mean ± SD of three biological replicates. Two-sided Student’s t test, ^**p < 0.01. The exact p values and related source data are provided in the “Source Data file”.