Escape from Notch suppression by homozygous DNMAML-GFP occurs frequently during T-ALL development. (A) Survival curve showing the fraction of LCR (N = 15), LCDDR (N = 8), and LCDD (N = 6) mice that develop T-ALL over time. (B) Flow cytometric analysis of thymic lymphomas of 4 representative LCDDR mice with T-ALL showing loss of expression of DNMAML-GFP. (C) Quantitative real-time polymerase chain reaction analysis for expression of Notch1 target genes (Deltex1, Myc, and Hes1). For the LCDDR1 tumor, messenger RNA (mRNA) of DNMAML-GFP⁺ cells is compared with mRNA of DNMAML-GFP⁻ cells. For the LCDDR2 tumor, mRNA of DNMAML-GFP⁺ cells is compared with mRNA of DNMAML-GFP⁻ cells of sorted ISP cells (CD8⁺TCRβ⁻) or DP cells (CD4⁺CD8⁺). (D) Thymic lymphoma cells from LCDDR mice were enriched for cells expressing DNMAML-GFP and transferred to lethally irradiated recipients. DNMAML-GFP expression was analyzed by flow cytometry in presort T-ALL cells, postsort T-ALL cells, and T-ALL cells in 4 representative secondary recipients. (E-F) DNA sequence analysis of (E) exon 34 (where PEST mutations occur; 4 LCDDR mice and 1 LCDR mouse) or (F) the 5′ region of endogenous Notch1 (where type I rearrangements occur¹⁶ ; 2 LCDDR mice) in thymic lymphomas sorted for DNMAML-GFP⁺ and DNMAML-GFP⁻ subpopulations.