Fig 2: Multiple DDR and DNA repair pathway genes targeted by radiation sensitizing miRNAs. (A) Western blot analyses of MAD2L2, WEE1, XPC, KU80, RAD23B, XLF and MCL1 48 h after LNCaP transfection with 20 nM of miR-890 (left) and miR-744–3p (right). Asterisk represents a nonspecific band. ACTB was utilized as a loading reference. (B) Normalized percent protein knock-down (mean ± SE, n = 3) by miR-890 (left) and miR-744–3p (right), relative to control miRNA (from three separate experiments). (C) Luciferase activities following transfection with indicated 3′UTR reporters and miR-890 or control mimetics in 293T cells. (D) Luciferase activities following transfection with indicated 3′UTR reporters and miR-744–3p or control mimetics in 293T cells. Relative luciferase activity (mean ± SE, n =12) is normalized to control miRNA for each reporter. *, P < 0.05 relative to control miRNAs.

Fig 3: DDB2 recruits XPC to telomeric 8-oxoG, while XPA recruitment is transcription-coupled and independent of DDB2.a, c Representative images showing recruitment of GFP-XPC (a) or GFP-XPA (c) to 8-oxoG at telomeres after dye (100 nM, 15 min) plus light (660 nm, 10 min) treatment in U2OS WT and DDB2 KO cells, 30 min post treatment. b, d Percentage telomeres colocalized with GFP-XPC (b) or GFP-XPA (d) after treatment, over a period of 3 h. e, g Representative images of GFP-XPC (e) or GFP-XPA (g) accumulation at damaged telomeres 30 min after dye plus light treatment in cells pretreated with transcription inhibitors a-amanitin and THZ1. f, h Quantification of e (f) and g (h). i, j Colocalization of GFP-XPC with telomeres after dye plus light treatment in U2OS-FAP-TRF1 cells transfected with control or OGG1 siRNA. k, l Colocalization of GFP-XPA with telomeres after dye plus light treatment in U2OS-FAP-TRF1 cells transfected with control or OGG1 siRNA. Data (a–l) represents mean ± SEM from two independent experiments. “n” represents the number of cells scored for each condition. One-way ANOVA (Sidak multiple comparison test) (b, d, f, h) and Student’s two-tailed t-test (j, l): **p < 0.01; ***p < 0.001; ****p < 0.0001. Scale: 5 µm. Source data are provided as a Source Data file. (See also Supplementary Fig. 4).

Fig 4: Effect of TPL and CBP on the mRNA and protein expressions of the NER pathway components. A375 cells were treated with TPL or CBP for 48 h with indicated concentrations. TPL abrogates the (A) mRNA and (B) protein expression levels of the rate-limiting components (ERCC1, ERCC2/XPD, ERCC3/XPB, ERCC4/XPF, ERCC5/XPG, XPA, and XPC) of the NER pathway, while CBP increases the (C) mRNA and (D) protein expression levels. (E) Effect of TPL (30 nM) and CBP (50 μM) alone or in combination on the levels of the NER pathway proteins in melanoma cells.

Fig 5: Schematic diagram demonstrating the inhibitory effects of TPL on CBP induced upregulation of NER pathway. TPL synergistically enhances CBP-induced apoptosis by selectively inhibiting the NER activity and overexpressing p53 in melanoma. The platinum induced DNA damage is identified and primarily repaired by the NER pathway. The damaged DNA can be recognized by XPC, and then XPA and RPA are recruited and act as another damage recognition complex that might participate in lesion verification. The DNA around the lesion site is opened asymmetrically in an ATP-dependent manner, employing the TFIIH complex with its two DNA helicases, ERCC3/XPB and ERCC2/XPD (Evans et al., 1997). TFIIH is involved in both transcription and NER processes. Subsequently, ERCC4/XPF-ERCC1 complex and ERCC5/XPG, two structure-specific endonucleases, incise on the 5′and 3′sides of the DNA adduct, respectively, to remove the damaged single-stranded DNA that contains the lesion. Finally, Pol δ/ε resynthesize the removed oligonucleotides and successfully repair the DNA damage by DNA ligase.