Fig. 4

NLRP3 acts as a target of WTAP in high glucose (HG)-induced HK-2 cells. A–G HK-2 cells treated with NG or HG with or without WTAP knockdown or overexpression (n = 3). A m⁶A levels measured by ELISA; B, C RNA immunoprecipitation (RIP) using anti-m⁶A antibody and RT-qPCR analysis of NLRP3 5′UTR and 3′UTR m⁶A levels; D, E relative mRNA and protein expression of WTAP and NLRP3 detected by RT-qPCR and western blot; F, G luciferase activity of NLRP3 5′UTR and 3′UTR; H NLRP3 mRNA level quantified by RT-qPCR in HK-2 cells with or without IGF2BP knockdown (n = 3); I half-life of the NLRP3 transcript measured by RT-qPCR in HK-2 cells with or without IGF2BP1 knockdown (n = 3). J–K The binding of IGF2BP1 to NLRP3 mRNA measured by RIP and RT-qPCR (n = 3). Unpaired Student’s t-test was used for the analysis between two groups, and one-way analysis of variance was used to analyze the data among multiple groups, followed by Tukey’s post hoc test. **P < 0.01, ***P < 0.001 compared with NG + shNC + vector or siNC. ^#P < 0.05, ^##P < 0.01, ^###P < 0.001 compared with HG + shNC + vector