Fig. 2: IκBα loss leads to decreased numbers of LT-HSCs at newborn and fetal stages.

A Scheme of the mouse mating to obtain IκBα WT, HET, and KO hematopoietic tissues. B Box plot with individual values of the frequency of (i) LSK (lin-SCA1+cKIT+) in the liver or bone marrow of newborn (P5/6) both obtained from n = 39 pups in 5 independent experiments and, (ii) LT-HSCs (LSKCD48-CD150+) from the previous gate (LSK) in the liver of newborn (P5/6) IκBα WT (n = 12), HET (n = 20) and KO (n = 7) and in the bone marrow of newborn (P5/6) IκBα WT (n = 11), HET (n = 18) and KO (n = 6). C Box plot with individual values of the frequency of (i) LSK in the fetal liver of E14.5 IκBα WT (n = 7), HET (n = 11) and KO (n = 11), and (ii) LT-HSCs (LSKCD48-CD150+) in E14.5 IκBα WT (n = 7), HET (n = 11) and KO (n = 11), both cased obtained from n = 19 embryos in 3 independent experiments. Statistical test for (B), (C): unpaired two-tailed Dunn’s non-parametric all-pairs comparison test after Kruskal–Wallis test (**p-value < 0.01, *p-value < 0.05, ns p-value > 0.05). Boxplots show the median (center line) first and third quartiles (box limits), and whiskers extend to minimum and maximum values. D Scheme illustrating the two strategies followed to explore the molecular changes in IκBα WT, HET, and KO. E14.5 FL were FACS sorted to obtain 5000 LSK cells for single-cell RNA-sequencing and 500 cells/sample of LT-HSCs for bulk RNA-sequencing. n = 4 embryos per genotype. E Violin plots with individual values depicting the expression levels from scRNAseq data of genes from NF-κB signaling (KEGG ID mmu04064) signature within each annotated cell population. Statistical test: two-tailed Wilcoxon rank sum test (ns p-value > 0.05). F Bar plot depicting significantly enriched MSigDB HALLMARK pathways in IκBα KO against WT/HET genotypes with positive Normalized Enrichment Score (NES), identified by GSEA from the bulk RNA-seq data (Benjamini–Hochberg procedure (FDR) adjusted p-value < 0.05). Bars sorted by NES. G NF-kB signaling signature (KEGG ID mmu04064) GSEA over the bulk RNA-seq data comparing IκBα WT/HET and KO LT-HSCs. Indicated p-value is a nominal p-value obtained from GSEA. Source data are provided as a Source Data file. A–D created with BioRender.com released under a Creative Commons Attribution-NonCommercial-NoDerivs 4.0 International license.