Mice with somatic inactivation of H2ax and p53 in thymocytes die of clonal thymic lymphomas. (A) LHP mice exhibit normal thymocyte development. Shown are representative anti-CD4 and anti-CD8 flow cytometric analysis of total LHP and HP thymocytes, and anti-c-kit and anti-CD25 flow cytometric analysis of lineage-negative LHP and HP thymocytes. The percentage of cells within each quadrant is indicated. (B) H2ax and p53 are deleted in LHP thymocytes. Southern blot analyses of H2ax and p53 status in genomic DNA isolated from thymocytes (Thy) or kidneys (Kid) of LHP or HP mice. Bands corresponding to the floxed (F) and deleted (Δ) alleles are indicated. (C) H2AX expression is lost in LHP thymocytes. Western blot analysis of H2AX and α-tubulin protein levels in total thymocytes of LHP and HP mice. (D) LHP mice die of thymic lymphomas. Kaplan-Meier curves show the percentage of tumor-free survival of LHP (n = 34) and H2ax^−/−p53^−/− (n = 32) mice. *The LHP mice that died of TCR-β⁺ tumors. The H2ax^−/−p53^−/− cohort is a historical control that was characterized previously.¹⁸  Log-rank test: P < .0001. (E) LHP mice develop TCR-β⁻ and TCR-β⁺ thymic lymphomas. Anti-TCR-β and anti-CD4 and anti-CD8 flow cytometric analyses of LHP tumors 111 and 222. The gates used to determine TCR-β, CD4, and CD8 status of tumors are shown. (F) LHP tumors arise from the clonal expansion of single thymocytes. Schematic diagram of the Tcrβ locus. Open boxes represent gene segments; black bars, locations of the 3′Jβ1 and 3′Jβ2 probes; and arrows, HindIII sites. The sizes of the HindIII fragments are shown. Southern blot analyses of Tcrβ rearrangements in HindIII-digested genomic DNA isolated from LHP thymic lymphomas. The locations of bands corresponding to unrearranged germline (GL) Jβ1 and Jβ2 segments are indicated.