Fig. 4
From: Mutational signatures reveal the role of RAD52 in p53-independent p21-driven genomic instability
Extended p21WAF1/Cip1 over-expression shapes the mutational signature landscape. a Escaped (30 days induced) Saos2- and Li-Fraumeni-p21WAF1/Cip1 Tet-ON cells exhibit specific patterns of single nucleotide substitution (SNS). SNSs with mapping quality above 30 found only in the escaped cells were filtered based on sequencing depth and scored as ESC-specific (see also Additional file 1: Figure S1). Those SNSs were used to calculate the mutational signature of ESC versus OFF cells (for details see “Methods” section and Additional file 1: Figure S1). Heat map shows the number of mutation type at each mutation context, which was corrected for the frequency of each triplet in the human genome (hg19). Histograms present the mutation-type frequency at each mutation context from two biological replicates of escaped Saos2- and Li-Fraumeni-p21WAF1/Cip1 Tet-ON cells, respectively. Both presentations show reproducible patterns of the mutational signatures 6, 15, 3 [8]. b Heat map showing the association of SNSs, nucleotide insertions (INS), and nucleotide deletions (DEL) with the observed chromosomal breakpoints (±50 kb around the breakpoint) versus the remaining genome in escaped Saos2- and Li-Fraumeni-p21WAF1/Cip1 Tet-ON cells. c Real-time RT-PCR assessment of BRCA1 and BRCA2 mRNA expression in induced and non-induced Saos2 and Li-Fraumeni p21WAF1/Cip1 Tet-ON cells (*p < 0.05 (Saos2), *p = 0.05 (Li-Fraumeni), t-test; error bars indicate standard deviation; n = 3 experiments). Loss of heterozygosity at the q arm of chromosome 13 (which hosts the BRCA2 locus (q13.10)) in induced Li-Fraumeni-p21WAF1/Cip1Tet-ON cells [9]. d Immunoblots depict reduced BRCA1 and BRCA2 expression in induced Saos2- and Li-Fraumeni-p21WAF1/Cip1 Tet-ON cells at the indicated time points. α-Tubulin served as loading control. MQ mapping quality, AF allele frequency, DP sequencing depth