Fig. 9
uc.339 expression is directly silenced by TP53. a qRT-PCR for uc.339 in A549 and H460 cells transfected with an anti-TP53 siRNA (si-TP53) (or its anti-scrambled control (si-SCR)), and in A549, H460, and H1299 cells transfected with a plasmid-expressing TP53 (or its empty plasmid counterpart) for 72 h. The levels of uc.339 have been normalized to RNU44 RNA and the results are presented as normalized to si-SCR or empty, respectively. Data are presented as mean ± s.d. of experiments conducted in triplicate. *P < 0.05. b Quantification of TP53 protein (left panel) and of uc.339 expression (right panel) in 22 paired primary NSCLC cancerous tissues (C) and the adjacent non-tumor lung (N). TP53 expression has been quantified with enhanced chemiluminescence (Femto) Kit and normalized to Vinculin, whereas uc.339 expression has been determined by qRT-PCR, normalized to RNU44 and log2 transformed. Data are shown as mean ± s.d., and referred to N mean value. The linear fold-change between C and N was 65.6. Paired t-test P-value <0.0001 (****). c Map showing the location of four identified TP53 consensus sequences (CS #1–4) relative to the transcription starting nucleotide of the uc.339 gene on chromosome 12. d Chromatin Immunoprecipitation showing the binding of TP53 to the four identified TP53 CS. e Luciferase reporter assay in A549, H460, and LoVo cells transfected with a TP53-expressing plasmid (or its empty plasmid counterpart) and a reporter plasmid containing the sequence of CS #4, or a plasmid in which CS #4 has been deleted (del CS #4). Luciferase activity has been normalized to Renilla (RLU), and the results are presented as normalized to empty. Data are shown as mean ± s.d. of experiments conducted in triplicate. *P < 0.05
