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. Author manuscript; available in PMC: 2021 Jul 9.
Published in final edited form as: Nat Genet. 2021 Apr 8;53(5):672–682. doi: 10.1038/s41588-021-00829-8

Extended Data Figure 7 |. Reactivation of L1 retrotransposition impairs myeloid leukemia.

Extended Data Figure 7 |

a, Schematic of 10 sgRNAs against the 5’UTR promoter sequences of the full-length L1Hs. b, CRISPRa-mediated activation of endogenous L1s in MOLM-13 cells using L1-promoter-targeting sgRNAs individually. Relative L1 expression was determined by qRT-PCR in non-transduced cells (control) and cells expressing non-targeting (sgNT) or L1-promoter-targeting sgRNAs (L1-sg1 to L1-sg10). Results are mean ± SD (N = 4 independent experiments) and analyzed by a one-way ANOVA with Dunnett’s test. c, CRISPRa of endogenous L1s in MOLM-13 cells using combinations of multiple L1-promoter-targeting sgRNAs. Results are mean ± SD (N = 4 independent experiments) and analyzed by a one-way ANOVA with Dunnett’s test. d, CRISPRa-mediated activation of endogenous L1s increased the expression of L1-encoded ORF1p in MOLM-13 and OCI-AML3 cells by Western blot. GAPDH was analyzed as a loading control. e, CRISPRa of endogenous L1s increased L1 retrotransposition in MOLM-13 and OCI-AML3 cells. Representative flow cytometry graphs are shown for control (sgNT) or L1 activated (L1-sg278 and L1-sg457) cells harboring the LRE3-EGFP or JM111 control, respectively. f, Activation of L1 retrotransposition impaired MOLM-13 cell growth by the negative-selection competition assay. Treatment with 3TC (10 µM) abrogated L1 retrotransposition-induced cell growth defects. Results are mean ± SD (N = 3 independent experiments) and analyzed by a two-way ANOVA with Dunnett’s test. g, Activation of L1 retrotransposition impaired OCI-AML3 cell growth, whereas 3TC treatment (10 µM) abrogated L1 retrotransposition-induced defects. Results are mean ± SD (N = 3 independent experiments) and analyzed by a two-way ANOVA with Dunnett’s test. h, 3TC treatment had no significant effect on AML cell viability as determined by treating MOLM-13 and OCI-AML3 cells with escalating doses (0 to 640 µM) of 3TC for 5 days. Results are mean ± SD (N = 3 independent experiments) and analyzed by a one-way ANOVA with Dunnett’s test.