Fig. 5: scRNA-seq identifies Ankle2 as a critical Zfp335-regulated gene controlling survival of DN4 thymocytes.
From: Loss of Zfp335 triggers cGAS/STING-dependent apoptosis of post-β selection thymocytes
a UMAP projection of WT and ‘true’ Zfp335 mutant DN4 cells colored by genotype. Volcano plot of all differentially expressed Zfp335 target genes (b) or those experimentally shown negatively regulate apoptotic processes (c) between Zfp335 mutant and WT cells. d Violin plots of antiapoptotic Zfp335 target gene expression between Zfp335 mutant and WT DN4 cells. FC indicates log2(Fold Change) between Zfp335cKO and WT, p indicates adjusted p-values. e Differential proportions of Zfp335 mutant cells expressing antiapoptotic genes from c, d compared to WT cells based on dropout-imputation. Representative gating (f) and quantification of apoptosis (g) or DP cell frequency (h) for EV or Ankle2 retrovirus transduced WT (n = 10) or Zfp335cKO (n = 8) DN3 thymocytes from male and female mice cultured on OP9-DL1 cells for 3 days. I Zfp335 ChIP-seq track of Ankle2 locus in WT thymocytes (Zfp335-C or Zfp335-N antibodies, GSE58293). Blue boxes indicate significant binding peaks. Correlation between Ankle2 (j) or Bax (k) and Zfp335 expression in Scid.adh.2c2.SunTag CRISPRi cells expressing nontargeting (open squares, n = 2) or Zfp335-targeting (closed circles, n = 10) gRNAs. Data are compiled from one (a–e), two (j, k), or three (f–h) independent experiments. P-values determined by two-sided Wilcoxon Rank Sum test (b–d), two-way ANOVA with post hoc Tukey’s test for multiple comparisons (g), repeated-measures ANOVA with post hoc Sidak’s test (h), or simple linear regression (j, k). Plots show mean ± SEM. Source data are provided as a Source Data file.
