53BP1 and p53 synergize to suppress genomic instability and lymphomagenesis

Abstract

p53-binding protein 1 (53BP1) participates in the cellular response to DNA double-stranded breaks where it associates with various DNA repair/cell cycle factors including the H2AX histone variant. Mice deficient for 53BP1 (53BP1(-/-)) are sensitive to ionizing radiation and immunodeficient because of impaired Ig heavy chain class switch recombination. Here we show that, as compared with p53(-/-) mice, 53BP1(-/-)/p53(-/-) animals more rapidly develop tumors, including T cell lymphomas and, at lower frequency, B lineage lymphomas, sarcomas, and teratomas. In addition, T cells from animals deficient for both 53BP1 and p53 (53BP1(-/-)/p53(-/-)) display elevated levels of genomic instability relative to T cells deficient for either 53BP1 or p53 alone. In contrast to p53(-/-) T cell lymphomas, which routinely display aneuploidy but not translocations, 53BP1(-/-)/p53(-/-) thymic lymphomas fall into two distinct cytogenetic categories, with many harboring clonal translocations (40%) and the remainder showing aneuploidy (60%). We propose that 53BP1, in the context of p53 deficiency, suppresses T cell lymphomagenesis through its roles in both cell-cycle checkpoints and double-stranded break repair.

Fig. 1.

Kaplan–Meier survival curve of 53BP1^−/−/p53^−/− double mutant animals and their cohorts. (A) Survival curve of 53BP1^−/−/p53^−/− double mutants relative to cohort genotypes, including 53BP1^+/−/p53^−/− and p53^−/− mice. Several genotypes, including 53BP1^−/−, survived throughout the duration of this study and, as a result, are represented by a single horizontal line. 53BP1^−/−/p53^−/− double mutants have a 50% survival rate compared with their p53 mutant cohorts. Sampling size for each genotype is listed. The differences in the timing of morbidity between 53BP1^+/−/p53^−/− and p53^−/− animals was determined to be statistically insignificant (P = 0.476) by using a one-way ANOVA and a t test with 90% confidence. In contrast, P < 0.001 between the survival curves for 53BP1^−/−/p53^−/− and p53^−/− breedings. (B) Pie chart showing the tumor spectrum observed for 53BP1^−/−/p53^−/− mice (n = 44). T, T lineage lymphomas; B, B lineage lymphomas; S, sarcoma; Te, teratomas; O, other (cause of death undetermined).

Fig. 2.

53BP1^−/−/p53^−/− thymic lymphomas display clonal translocations. SKY analysis of four T cell lymphomas displaying clonal translocations was performed as described in Methods. Chromosome number and translocations, if present, are listed for one tumor. See text for details.

Fig. 3.

Supernumerary chromosomes with little evidence of translocations in 53BP1^−/−/p53^−/− thymic lymphomas. Representative metaphases derived from SKY from a given tumor are shown. n = number of examined metaphase spreads. T2961 possesses 39–60 chromosomes (n = 15). T2963 has 50–54 chromosomes (n = 19). T2945 is essentially euploid (39–42 chromosomes) but has several monosomic chromosomes (6, 10, 17) depending on the cell in addition and trisomy 5 and 11 (n = 8). T3096 was found to have 40–54 chromosomes (n = 20) in addition to nonclonal translocations t(19;14) and t(7;16;4).

Fig. 4.

Increased genomic instability in 53BP1^−/−/p53^−/− thymocytes. (A) Quantification of structural chromosomal aberrations in Con A-stimulated T cells from wild-type (n = 4), p53^−/− (n = 3), 53BP1^−/− (n = 7), and 53BP1^−/−/p53^−/− (n = 3) mice. Of the three p53 mutants analyzed, two were derived from Donehower and colleagues (18, 19), and the third was from Jacks and colleagues (20). Metaphase spreads were hybridized with a telomere-specific Cy3-labeled PNA probe (red) to visualize the four ends of each chromosome and counterstained with DAPI (blue). At least 30 metaphases per culture were analyzed. Aberrations are represented as follows: % aberrations per metaphase = (total number of aberrations/total number of metaphases) × 100. (B) Partial metaphase spreads of 53BP1^−/−/p53^−/− T cells showing three chromosomal breaks (yellow arrows) and a dicentric resulting from rejoining of two broken chromosomes. A normal chromosome with four telomere signals is shown on the left for comparison.