Fig. 4

GPD1 upregulates TRPV2 expression to promote Ca²⁺ influx, leading to apoptosis of bladder tumor cells. A RNA-seq analysis of control 5637 cells (CON), GPD1-OE 5637 cells (GPD1) and K120A GPD1-OE 5637 cells (K120A). Venn diagram showing the overlap of differentially expressed genes between the three groups. B Heatmap showing that the 1270 differentially expressed genes were shared by CON versus GPD1 and K120A versus GPD1. C GSEA of the genes associated with the NOD-like receptor signaling pathway. D RT–qPCR to validate the mRNA levels of differential genes (TRPV2, NLRP1, and ASC) associated with the NOD-like receptor signaling pathway identified by RNA-seq. E Western blotting to detect TRPV2 expression in control 5637 cells (CON), GPD1-OE 5637 cells (GPD1) and K120A GPD1-OE 5637 cells (K120A). F and G Flow cytometry was performed to detect TRPV2 expression on the membranes of 5637 and T24 cells overexpressing GPD1 or K120A GPD1. H and I Flow cytometry was performed to detect TRPV2 expression on the membranes of 5637 and T24 cells treated with G3P/NAD⁺. J and K Flow cytometry was performed to detect intracellular Ca²⁺ levels in 5637 cells and T24 cells overexpressing GPD1 or K120A GPD1. L and M Flow cytometry was performed to detect intracellular Ca²⁺ levels in 5637 cells and T24 cells treated with G3P/NAD⁺. N Western blots show the efficiency of siRNA knockdown of TRPV2 expression in T24 cells. O Flow cytometry was performed to detect intracellular Ca²⁺ levels of 5637 cells transfected with siRNAs in the presence or absence of G3P/NAD⁺. P Flow cytometry was performed to detect intracellular Ca²⁺ levels of T24 cells transfected with siRNAs in the presence of G3P/NAD⁺. Q Flow cytometry analysis of apoptosis in 5637 cells transfected with scramble or siRNAs in the presence or absence of G3P/NAD⁺. R Flow cytometry analysis of apoptosis in T24 cells transfected with scramble or siRNAs in the presence or absence of G3P/NAD⁺