scRNA-seq analysis of splenic DCs sorted from WT and H2-Aa ^cit/cit mice. (A and B) Violin plots showing the expression distribution of DC subset-defining markers (A) or cDC2 subset-defining markers (B) in different clusters from Fig. 4 G. (C) Frequencies of ESAM^hi or Clec12A^hi splenic cDC2. Data points represent individual mice (n = 5 per group). Error bars indicate SD. No significant difference by Student’s t test. (D) KEGG pathway enrichment analysis of genes significantly increased in H2-Aa^cit/cit relative to WT Wdfy4^hi cDC2 (adjusted P ≤ 0.05, n = 203) and Ffar^hi cDC2 (adjusted P ≤ 0.05, n = 261). One-sided hypergeometric test was used to determine statistical significance of enrichment. (E) Volcano plot showing differentially expressed genes in WT versus H2-Aa^cit/cit(cit) Wdfy4^hi cDC2 (n = 570) and Ffar^hi cDC2 (n = 688). Adjusted P ≤ 0.05. Shaded areas contain genes with Log₂Fold change (FC) >0.4 and Log₂FC less than −0.4. Data are representative of one experiment (A, B, D, and E) or two experiments (C). WT littermates were used as controls (A–E).