Gene repression upon IR is associated with the loss of H3K9me3 at intronic L1Md loci harboring NF-κB binding sites. (A and C) Experimental design and mRNA expression assessed by qRT-PCR in HSCs 1 mo after TBI. Ct values were normalized to RPL32 and HPRT. Results are expressed as fold change from the mean value of the NIR condition. Each dot represents a pool of three (NIR) or four (IR) mice. Means ± SEM from three to four independent experiments. (B and D) Experimental design and H3K9me3 ChIP-qPCR enrichment 1 mo after TBI. The primer positioning at the intronic L1Md allowing the amplification of a unique and specific product is shown (B). Each dot represents a pool of three (NIR) or four (IR) mice from two to four independent experiments. Results are means ± SEM of the percentage of input normalized to the NIR condition. t test. (E–G) CRISPR/Cas9-induced deletion intronic L1Md in Mecom gene. gRNA positioning around L1Md sequence (E), experimental design (F), and mRNA expression assessed by qRT-PCR in Cas9-gRNA RNP electroporated HSCs 48 h after irradiation in vitro (G). Ct values were normalized to Rpl32. mRNA expression was normalized to mock NIR values. Each dot represents a pool of electroporated HSCs from three to five independent experiments. One-way ANOVA with Sidak’s multiple comparison test. (H and I) De novo motif discovery analysis performed with the BAMMmotif tool on L1Md sequences located in introns of downregulated genes vs. nonderegulated genes (H) or the genes participating vs. not participating to the loss of the LT-HSC signature (I). Enriched motifs were matched to known motifs using the Hocomoco mouse database. *, P < 0.05; **, P < 0.01; ***, P < 0.001.