Fig. 4: PRMT5 inhibition or depletion induces accumulation of DNA damage in ATM-deficient MCL cell lines and restores the expression and tumor-suppressive function of wild-type p53.

a Indicated MCL cell lines were treated with PRMT5 inhibitor GSK3326595 and subjected to IF staining with anti-γH2AX, a marker for DNA damage. Scale bars = 20 μm. b γH2AX foci in (a) were quantified using ImageJ. c Expression of genes (AR, DNAPK, NHEJ1 and RAD51) in DDR pathways analyzed by real-time qPCR in Granta-519 and Z-138 cell lines treated with PRMT5 inhibitor GSK3326595. d Indicated cell lines were treated with PRMT5 inhibitors GSK3326595 (1 µM) and LLY-283 (1 µM) and subjected to RT-PCR analysis of MDM4 mRNA splicing. e Expression of PRMT5 and p53 pathway genes determined by immunoblotting in Granta-519 with or without PRMT5 KO. f Granta-519 cells were stained with anti-γH2AX for 2 h after exposure to 10 Gy γ-irradiation. Scale bars = 20 μm. γH2AX foci were quantitated using ImageJ. g Indicated cell lines were treated as in (d) before they were subjected to western blotting analysis of MDM4 and p53. h Indicated MCL cell lines were treated with PRMT5 inhibitor GSK3326595 or LLY-283 and subjected to apoptosis analysis. i Western blotting analysis of the expression of PRMT5 and p53 in the cell lines treated as in (h). j Expression of p53 target genes (MDM2, p21, and PUMA) analyzed by real-time qPCR in in Z-138, Granta-519 and JVM-2 cell lines treated with LLY-283. k A schematic illustration of the alternative splicing of MDM4 upon PRMT5 depletion stabilizing p53. For all panels: *P < 0.05; **P < 0.01; ***P < 0.001; ****P < 0.0001; ns (not statistically significant), P ≥ 0.05.