Fig. 3: hUSI revealed potential senescence regulators in a Perturb-seq dataset.
a, UMAP plot showing the distribution of cells in the Perturb-seq dataset. b, GSEA results for hUSI-identified genes whose perturbations could induce cell into senescence state. Each line corresponds to a different Hallmark gene set, with line color shading indicating NES. c, Boxplot showing the hUSI levels of cells targeted by the top 10 hUSI-identified genes compared to that of the control cells (n = 11,705 cells; ***P < 1 × 10−3, two-tailed Wilcoxon test with Benjamini–Hochberg correction). The box represents the IQR, with its lower and upper edges indicating the 25th and 75th percentiles, respectively. The medium value (50th percentile) is shown within the box, and the whiskers extend to the minimum and maximum values within 1.5 times the IQR of the quartiles. d–g, Senescence validation of NACA knockdown in ARPE19 cells (n = 3 biological replicates; two-tailed t-test). Data are presented as mean ± s.e.m. NACA was knocked down by two siRNAs (si-NACA-1 and si-NACA-2) (d). e, Left: the senescence state is demonstrated by representative images of SA-β-gal staining. Right: the bar chart shows the percentage of SA-β-gal-positive cells (n = 3 biological replicates). Scale bars, 100 μm. The gene expression levels of CDKN1A, CDK1, LMNB1 and MKI67 evaluated by qRT–PCR (f) and the western blot of p21 protein levels before and after NACA knockdown in ARPE19 cells, with GAPDH serving as the internal control (g). h–k and l–o present the senescence validation results for ECT2 and PSMD13 (as those in d–g), respectively. Scale bars, 100 μm. For all statistical tests: **P < 0.01 and ***P < 1 × 10−3. The exact P values are reported in Source Data for Fig. 3. NC, negative control.
