Figure 1.
A 13-gene functional p53 score established from isogenic TP53 CRISPR/Cas9 HMCLs. (A) Principal component analysis of TP53+/+ or TP53−/mut and TP53−/− cells derived from NCI-H929, XG7, JIM3, and NAN3 HMCLs. Principal component analysis was performed on all cells profiled in triplicate wells using RNA-seq (DGE-seq). RNA profiling was performed on TP53+/+ control (bulk) cells and TP53−/− clones. (B) Number of genes significantly differentially expressed between TP53+/+ control cells and TP53−/− clones. The graph represents the number of genes significantly downregulated or upregulated in TP53−/− NCI-H929, XG7, JIM3, and NAN3 clones compared to their respective TP53+/+ or TP53−/mut control cells (false discovery rate < 0.05). Genes were classified according to their unknown or known p53 transactivation: early-direct, late-direct, late-indirect and not regulated by p53.28 Expression profiling was performed by DGE-seq in triplicate wells (see supplemental Table 2 for complete gene listing). (C) Schematic representation of the functional p53 score construction. Sixteen downregulated genes in TP53−/− clones were shared between NCI-H929 and XG7, and 13 known as early-direct p53 regulated genes were selected for establishing the p53 functional score. (D) The p53 score segregates the clones according to their TP53 status. The score was calculated in TP53+/+ or TP53−/mut and TP53−/− NCI-H929, XG7, JIM3, and NAN3 cells under constitutive (left) or 24-hour nutlin3a (2 μM for XG7 cells and 10 μM for all other clones) culture (right). Statistical analyses were performed using the Mann-Whitney U test. Blue, red, and orange represent TP53+/+, TP53−/−, and TP53−/mutated cells, respectively. (E) The p53 score segregates HMCLs according to their p53 status. The score was calculated in 18 HMCLs characterized by DGE-seq. TP53 sequencing was performed on complementary DNA and p53 expression was determined by western blotting (supplemental Table 4). Statistical analyses were performed using the Kruskal-Wallis test with multiple comparisons. (F) The p53 score is functional in 1105 cancer cell lines with different TP53 status. The score was calculated in 1105 cancer cell lines from DepMap and analyzed according to the presence of TP53 deletion and/or mutation (left) and to the number of TP53 deletion or mutation (right). Statistical analyses were performed using the Kruskal-Wallis test with multiple comparisons. ∗∗∗∗P < .0001; ∗∗∗P < .001; ∗∗P < .01; ∗P < .05. ns, not significant.

A 13-gene functional p53 score established from isogenic TP53 CRISPR/Cas9 HMCLs. (A) Principal component analysis of TP53+/+ or TP53−/mut and TP53−/− cells derived from NCI-H929, XG7, JIM3, and NAN3 HMCLs. Principal component analysis was performed on all cells profiled in triplicate wells using RNA-seq (DGE-seq). RNA profiling was performed on TP53+/+ control (bulk) cells and TP53−/− clones. (B) Number of genes significantly differentially expressed between TP53+/+ control cells and TP53−/− clones. The graph represents the number of genes significantly downregulated or upregulated in TP53−/− NCI-H929, XG7, JIM3, and NAN3 clones compared to their respective TP53+/+ or TP53−/mut control cells (false discovery rate < 0.05). Genes were classified according to their unknown or known p53 transactivation: early-direct, late-direct, late-indirect and not regulated by p53.28 Expression profiling was performed by DGE-seq in triplicate wells (see supplemental Table 2 for complete gene listing). (C) Schematic representation of the functional p53 score construction. Sixteen downregulated genes in TP53−/− clones were shared between NCI-H929 and XG7, and 13 known as early-direct p53 regulated genes were selected for establishing the p53 functional score. (D) The p53 score segregates the clones according to their TP53 status. The score was calculated in TP53+/+ or TP53−/mut and TP53−/− NCI-H929, XG7, JIM3, and NAN3 cells under constitutive (left) or 24-hour nutlin3a (2 μM for XG7 cells and 10 μM for all other clones) culture (right). Statistical analyses were performed using the Mann-Whitney U test. Blue, red, and orange represent TP53+/+, TP53−/−, and TP53−/mutated cells, respectively. (E) The p53 score segregates HMCLs according to their p53 status. The score was calculated in 18 HMCLs characterized by DGE-seq. TP53 sequencing was performed on complementary DNA and p53 expression was determined by western blotting (supplemental Table 4). Statistical analyses were performed using the Kruskal-Wallis test with multiple comparisons. (F) The p53 score is functional in 1105 cancer cell lines with different TP53 status. The score was calculated in 1105 cancer cell lines from DepMap and analyzed according to the presence of TP53 deletion and/or mutation (left) and to the number of TP53 deletion or mutation (right). Statistical analyses were performed using the Kruskal-Wallis test with multiple comparisons. ∗∗∗∗P < .0001; ∗∗∗P < .001; ∗∗P < .01; ∗P < .05. ns, not significant.

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