Fig. 8

Pharmacological inhibition of PRMT5 impairs ZMYND11-HNRNPA1 interaction and suppresses in vivo metastatic capacity of prostate cancer cells with ZMYND11-low expression. a Schematics of arginine methylation states by PRMT family of enzymes. b Co-immunoprecipitation assay of HEK293T cells expressing T7-tagged HNRNPA1 and the indicated GFP-tagged PRMT family member. c Co-immunoprecipitation assays using 22Rv1 cell protein extracts co-expressing V5-tagged ZMYND11 and T7-tagged HNRNPA1 while having stable expression of control or shRNAs against PRMT5. d Immunoprecipitation-Western blot analysis of an endogenous interaction between ZMYND11 and HNRNPA1 in 22Rv1 cells stably expressing control or ZMYND11-targeting shRNAs. e Immunoprecipitation using V5-tag antibodies from 22Rv1 cells ectopically co-expressing ZMYND11 and HNRNPA1 while treated with PRMT1 inhibitors (AM1 or DCLX069, 50 μM, 48 h) or PRMT5 inhibitors (EPZ015666 or GSK3326595, 10 μM, 48 h). f Co-immunoprecipitation assay of an endogenous interaction between ZMYND11 and HNRNPA1 in 22Rv1 cells treated with the indicated PRMT1/5 inhibitor. g Half-inhibitory concentration (IC50) test of two PRMT5 inhibitors (EPZ015666 or GSK3326595, 10 μM, 48 h) for ZMYND11 knockdown or control cells in 22Rv1. h Schematics of in vivo mouse experiments with PRMT5-Selective Inhibitors. 22Rv1 cells with stable knockdown of ZMYND11 while expressing luciferase were injected into the tail-vein of male NOD-SCID mice. At 5, 6, and 7 weeks after injection, mice were subject to intraperitoneal treatment with the indicated PRMT5 inhibitors. Shown are the representative images at indicated time points (i) and weekly quantification of BLI photon flux of lung metastasis in mice (j). Errors bar, ±SD, n = 5. *p < 0.05, **p < 0.01, ***p < 0.001, Student’s t tests. k FACS-analysis of GFP-positive 22Rv1 cells (CTCs) in the peripheral blood of SCID mice (n = 4). The scatter plot indicated the number of CTCs recovered from each mouse treated with vesicles or PRMT5 inhibitors. Statistical significance was assessed using two-tailed Student’s t test. * p < 0.05. l Organoid images derived from prostatic ZMYND11 knockdown Pten^−/− mice treated with PRMT5 inhibitors (EPZ015666 or GSK3326595, 10 μM); quantitative results are representative of 3 experiments shown at the right. Scale bar: 40 μm. Errors bar, ±SD, n = 5. *p < 0.05, **p < 0.01, Student’s t tests. m A model illustrating how ZMYND11 recognition of arginine methylation constrains HNRNPA1-mediated tumor progression is proposed. Top: In normal cells with higher expression of ZMYND11, the protein can specifically recognize the R194 methylation of HNRNPA1, which is catalyzed by PRMT5. This interaction inhibits HNRNPA1’s involvement in stress granule formation and the alternative splicing of PKM, thereby mitigating tumor aggressiveness. Bottom: In tumor contexts with reduced ZMYND11 expression, HNRNPA1 promotes the formation of stress granules and shifts the PKM isoform balance towards a higher PKM2/PKM1 ratio, contributing to tumor progression. In this scenario, PRMT5 inhibitors emerge as potential therapeutic agents capable of curtailing cancer progression by disrupting the methylation-dependent interaction between ZMYND11 and HNRNPA1, ultimately impairing HNRNPA1’s tumor-promoting activities