(A) Scatter plot of the intron retention index versus the fold change of gene expression (Prmt5-cKO/control) for total TRA genes expressed in mTEC^hi cells. Pearson’s correlation coefficient (r) and P values are shown. (B) Intron retention index for Prmt5-induced TRA genes (total read count ≥10 in Prmt5-deficient mTEC^hi cells). Each bar indicates an individual Prmt5-induced TRA gene. Genes with increased (fold change >1.5) or decreased (fold change <0.75) intron retention in Prmt5-deficient mTEC^hi cells are shown in red and blue, respectively. (C and D) Heatmaps showing the intron ratio (C) and relative mRNA expression (D) of Prmt5-induced TRA genes (top 30 in B) in mTEC^hi cells from the indicated mice. The data indicate the mean value for the intron ratio and relative RPKM. (E) Graphs show the mean ratio of intron retention of the TRA genes categorized in Figure 3G (total read count >0 in each group). Significance was determined using the FDR of the Benjamini and Hochberg method (^#q < 0.05 and ^###q < 0.001). + indicates the mean. (F) Total mTECs (EpCAM⁺CD45^–UEA-1⁺Ly51^–) isolated from control or Aire-KO mice were subjected to SDS-PAGE followed by immunoblotting with antibodies against Prmt5, SmD3, sDMA, and β-actin. Representative data from 2 independent experiments are shown. Numbers below the bands and beside the arrow indicate the relative intensity of each band. (G) Relative expression levels of Ano9 and Gnb3 in sorted mTEC^hi cells were analyzed by qRT-PCR (control, n = 4; Prmt5-cKO, n = 3). *P < 0.5 and **P < 0.01, by unpaired, 2-tailed Student’s t test. (H) Thymic sections from 4- to 5-week-old control mice, Prmt5-cKO mice, and Aire-KO mice were stained with antibodies against pan-keratin (green) and GNB3 (red). Representative images from 2 independent experiments are shown. Scale bar: 100 μm.