Deletion of Glis2 interferes with differentiation of MOZ-TIF2 leukemic cells induced by deletion of Ring1A/B. (A) PCR analysis of genomic Ring1B and Glis2 sequences in MOZ-TIF2–transduced Ring1A^−/−;Ring1B^f/f;CreERT2 and Ring1A^−/−;Ring1B^f/f;Glis2^/f/f;CreERT2 BM cells in the presence of either 4-OHT or ethanol. (B) Quantitative RT-PCR analysis of Glis2 expression in Ring1A^−/−;Ring1B^f/f;CreERT2 and Ring1A^−/−;Ring1B^f/f;Glis2^/f/f;CreERT2 BM cells transduced with MOZ-TIF2. RNA was extracted from cells incubated with 4-OHT for 48 hours. Error bars represent SD (n = 4). (C) Serial replating experiments of MOZ-TIF2–transduced Ring1A^−/−;Ring1B^f/f;CreERT2 and Ring1A^−/−;Ring1B^f/f;Glis2^/f/f;CreERT2 BM cells in the presence of either 4-OHT or ethanol. Error bars represent SD (n = 3). (D) Representative colony morphology and May-Grünwald Giemsa staining of cells from first-round colonies formed by MOZ-TIF2–transduced Ring1A^−/−;Ring1B^f/f;CreERT2 and Ring1A^−/−;Ring1B^f/f;Glis2^/f/f;CreERT2 BM cells in the presence of either 4-OHT or ethanol. Scale bar, 50 μm. (E,F) Secondary recipients of AML mice transplanted with MOZ-TIF2–transduced Ring1A^−/−;Ring1B^f/f;Glis2^/f/f;CreERT2 BM cells were treated with tamoxifen or corn oil (vehicle) for 5 consecutive days when the proportion of GFP⁺ MNCs in peripheral blood reached 50%. On the day following the last treatment, BM-MNCs were harvested and subjected to (E) flow cytometric analyses and (F) quantitation of L-GMP cells (mean ± SD, *P < .01, n = 6).