Fig. 6: The function of PARP1 in PD-L1 transcriptional regulation depends on NPM1. | Nature Communications

Fig. 6: The function of PARP1 in PD-L1 transcriptional regulation depends on NPM1.

From: NPM1 upregulates the transcription of PD-L1 and suppresses T cell activity in triple-negative breast cancer

Fig. 6

a PD-L1 promoter activity in MDA-MB-231 cells transfected with control shRNA (sh-PARP1-nc or sh-NPM1-nc), PARP1 shRNA (sh-PARP1), or NPM1 shRNA (sh-NPM1) was measured. b PD-L1 mRNA levels in MDA-MB-231 (top) and HCC1806 (bottom) cells stably transfected with control shRNA, PARP1 shRNA, or NPM1 shRNA. c Protein levels of NPM1, PARP1, and PD-L1 in MDA-MB-231 (left) and HCC1806 (right) cells with stable knockdown of NPM1 or/and PARP1 were detected by Western blot. d Jurkat cells were co-cultured with MDA-MB-231 cells with PARP1 or/and NPM1 stably knocked down, and IL-2 was detected by ELISA. e, f MDA-MB-231 (left) and HCC1806 (right) cells stably transfected with control or NPM1 shRNA were treated with 25 μM olaparib or vehicle (DMSO). PD-L1 mRNA (e), NPM1 and PD-L1 protein expression (f) was detected. g ChIP-qPCR of PD-L1 promoter region was conducted with NPM1, PARP1 or control IgG antibody in MDA-MB-231 cells (left). ChIP-qPCR was conducted with NPM1 or IgG antibody in MDA-MB-231 cells without or with PARP1 stable knockdown (right). h The structures of wildtype NPM1 and the ΔC35 mutant of NPM1. i ChIP-qPCR of PD-L1 promoter was performed with Flag or IgG antibody in MDA-MB-231 cells transiently transfected with Flag-NPM1 or Flag-NPM1-ΔC35. j–l MDA-MB-231 cells were transfected with vector, Flag-NPM1 or Flag-NPM1-ΔC35 plasmid. PD-L1 promoter activity (j), PD-L1 mRNA (k), Flag and PD-L1 protein expression (l) was detected. m HEK293T cells were transfected with Myc-PARP1 alone, or together with Flag-NPM1 WT or Flag-NPM1 ΔC35. The precipitates were then detected with anti-Myc antibody. Data were presented as mean ± s.d. of three independent experiments. g, i were analyzed by two-side Student’s t-test, *P < 0.05, **P < 0.01, ***P < 0.001; ns, not significantly different. j, k were analyzed by one-way ANOVA + two-side Dunnett test, ***P < 0.001, ****P < 0.0001; ns, not significantly different. a, b, d, e were analyzed by ANOVA + two-side Tukey test, **P < 0.01, ***P < 0.001, ****P < 0.0001; ns, not significantly different. Source data are provided as a Source Data file.

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