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. 2020 Mar 9;11:1270. doi: 10.1038/s41467-020-15003-7

Fig. 8. RNF113A limits the production of ROS through Noxa1 expression upon DNA damage.

Fig. 8

a RNF113A controls the splicing of the Noxa1 transcript. A shematic representation of the Noxa1 gene is illustrated. Three distinct Noxa1 transcripts are detected by RT-PCR using primers 1 and 2 located within exon 7 and 11, respectively. A first transcript, named “Truncated 2 Noxa1” includes introns 9 and 10 and gives rise to a 283 amino acids long Noxa1 protein. A second transcript referred to as “Truncated 1 Noxa1” has intron 10 and gives rise to a 318 amino acids long protein whereas a third transcript lacking both introns 9 and 10 codes for a 483 amino acids long Noxa1 protein. A quantification of all signals is illustrated on the right. b, c RNF113A or SF3B2 deficiency interferes with Noxa1 protein synthesis. Control (b, c), RNF113A (b)- or SF3B2 (c)-depleted A549 cells treated or not with Cisplatin (25 μM for 24 h) were subjected to WB analyses. d RNF113A limits ROS production upon DNA damage by regulating Noxa1 splicing. Control or RNF113A-depleted A549 cells were treated or not with Cisplatin (25 μM for 24 h) and ROS production was accessed using carboxy-H2DFFDA. Cells were analyzed on FACS Canto II and the data were generated using the FlowJo program. Representative plot from one of multiple experiments shows an increase of ROS production in RNF113A-depleted cells which further increases after Cisplatin treatment (on the top). At the bottom, data are illustrated in the histogram build using % of mean intensity received after analysis data using the BD FACSDIVA software. Data were obtained with three replicates for each experimental condition from three independant experiments (mean ± SD, two-sided T-test (***p < 0.001). e Increased NADPH/NADP ratio upon RNF113A deficiency after treatment with Cisplatin (25 μM for 24 h). The NADPH/NADP ratio in control and RNF113A-depleted cells treated or not with Cisplatin was established (see the “Methods” section for details). Results of two independent experiments are shown (***p < 0.001).