Fig 2: MYB::PLEKHO1 can independently overcome G1/S arrest to induce leukemia.(A) V5 and IgG CUT&RUN tracks at the Il3ra, Bcl2, and Myc loci in Cdkn2a-KO MYB::PLEKHO1 leukemia cells. (B) TPM of Il3ra (CD123), Bcl2, Myc, Ncam1 (CD56), Nrp1 (BDCA4), and Irf4 in Hoxb8-FL cell–derived pDCs (n = 8) and Hoxb8-FL cell–derived leukemias of the indicated genotypes (n = 4 per genotype). (C) Growth curves of Hoxb8-FL.MS5 cells transduced with V5-MYB constructs of the indicated genotypes. (D) Western blots showing p16INK4A and p19ARF expression in Hoxb8-FL.MS5 cells of the indicated genotypes. (E) Cell cycle analysis of Hoxb8-FL.MS5 cells. (Left) Representative flow plots of the indicated genotypes. (Right) Quantification of cells in each cell cycle phase as a proportion of total cells (n = 4 per genotype). Significance tests compare proportion of cells in G1. (A and E) Data represent mean ± SEM. Significance determined by 1-way ANOVA with Tukey correction for multiple comparisons. *P <0.05, **P <0.01, ***P < 0.001.

Fig 3: BPDCN cells undergo loss of MYB and cell death upon ATRA treatment.(A) Number of viable GFP–dTom– and GFP+dTom+ spleen cells from leukemic Cdkn2a-KO MYB::PLEKHO1 recipient mice after 2 days of 1 μM ATRA treatment, normalized to 0 μM ATRA (n = 3 from each of 6 recipient mice). (B) Number of viable GFP+dTom+ cells per femur and tibia, and per 1 mL of peripheral blood of Cdkn2a-WT MYB::PLEKHO1 secondary recipient mice, treated daily with 20–40 mg/kg ATRA for 12 days (DMSO n = 5, ATRA n = 3). (C) Number of viable human CD45+ cells per spleen of BPDCN PDX AL04 recipient NSG mice treated daily with 20 mg/kg ATRA for 28 days (DMSO, n = 5; ATRA, n = 5). (D) Number of viable CAL1 and K562 cells after 2 days of ATRA treatment at the indicated dose, normalized to 0 μM ATRA (n = 3 per condition). (E) Annexin V apoptosis analysis of CAL1 and K562 cells after 2 days of ATRA treatment at the indicated dose (n = 3 per condition). Significance tests compare proportion of apoptotic cells (annexin V+DAPI–). (F) Cell cycle analysis of CAL1 and K562 cells after 2 days of ATRA treatment at the indicated dose (n = 3 per condition). Significance tests compare proportion of cells in G1. (G) Median fluorescence intensity for CD11b APC and CD11c APC-Cy7 in CAL1 and K562 cells after 2 days of ATRA treatment at the indicated dose (n = 3 per condition). (H) Western blots showing MYB expression in CAL1 and K562 after 2 days of ATRA treatment at the indicated dose. (I) Western blots showing MYB and MYB::ZFAT expression in BPDCN PDX AL04 and AL05 cells after 2 days of ATRA treatment at the indicated dose. (A–G) Data represent mean ± SEM. Significance determined by 2-tailed t test (A–C, and F) or 1-way ANOVA with Tukey correction for multiple comparisons (D, E, and G). *P <0.05, ***P <0.001.