Fig. 2: Mitochondrial DNA fuses to CRISPR-Cas9-target sites of the nuclear DNA ex vivo and in vivo.
From: Transfer of mitochondrial DNA into the nuclear genome during induced DNA breaks
a Schematic of universal T cell manufactory. Primary T cells isolated from human or mice were activated and edited by CRISPR-Cas9. After edition, T cells underwent ex vivo culture or were infused into recipient mice. b Distribution of mtDNA-nuclear DNA fusions after CRISPR-Cas9 editing for 3, 7, and 14 days in human CAR T cells. Human primary T cells isolated from cord blood were activated for 3 days and subsequently transfected with Cas9/gRNA ribonucleoprotein (RNP) complexes targeting TRAC, TRBC, and PDCD1 loci. Cells were collected after 3, 7, or 14 days and subjected to PEM-seq libraries. Legends are described in Fig. 1c. c Schematic showing the production of mouse TCR-T cells. TCR-T cells post-editing were infused to Rag1−/− recipient mice for 3 weeks, and subsequently isolated for PEM-seq analysis. d Distribution of mtDNA-nuclear DNA fusion junctions in CRISPR-Cas9 treated mouse TCR-T cells before and post-infusion for 3 weeks. Legends are described as depicted in (b). Percentages show the frequency of each mtDNA-nuclear DNA fusion out of Cas9-induced editing events. e Prey lengths of 38 mt-nuclear DNA fusions sharing the same junction from expanded mouse TCR-T cells indicated in (d). f Sequence logo showing the frequency of nucleotides in random molecular barcodes derived from the 38 reads of mt-nuclear DNA fusion at a single junction in expanded TCR-T cells indicated in (d). g Distribution of mtDNA integration in the nuclear DNA post base editor (BE3) treatment in mouse embryos. MT, mtDNA. h Number of mt-nuclear DNA fusion events identified in BE3-treated and -untreated samples. Two-sided t-test.
