Fig. 4: Pharmacological inhibition of SUMOylation restores adipogenesis in IRX3-KO cells.

To assess whether hyperactive SUMOylation contributes to impaired adipogenesis in IRX3-KO cells, ME3 control and IRX3-KO cells were treated with the SUMO E1 inhibitor ML-792 (0.5 µM) from days \(-\)2 to 9 of adipocyte differentiation. a Fluorescence and brightfield microscopy images of cells stained with Bodipy lipid dye on day 9. Representative fields from biological triplicates (9 fields/well) shown from one of four independent experiments. Scale bar, 400 µm. See Supplementary Fig. S5a for full image sets. b Higher magnification of selected fields from a. Scale bar, 50 µm. c Quantification of Bodipy intensity, lipid droplet number, and average lipid volume per cell. Data from 9 fields per well merged; n \(=\) 3 wells per group. ***p_adj \( < \) 0.001 (IRX3-KO vs. control); ^##p_adj \(=\) 0.002, ^###p_adj \( < \) 0.001 (ML-792 vs. DMSO); ns, not significant; two-way ANOVA with Holm–Sidak correction. d Adipogenic gene expression in ME3 control and IRX3-KO cells treated with ML-792; n \(=\) 3 from one of two experiments. Data normalized to Rps13 and expressed relative to DMSO-treated control. *p_adj \( < \) 0.05, **p_adj \( < \) 0.01, ***p_adj \( < \) 0.001. e Relative effect of ML-792 on gene expression (ML-792/DMSO) in control and IRX3-KO cells. *p_adj \( < \) 0.05, **p_adj \(=\) 0.009; multiple unpaired, two-sided t-tests with Holm–Sidak correction. f Dose-response of ML-792 on lipid accumulation in IRX3-KO cells when added from day \(-\)2 or day 0 through day 9; n \(=\) 4 from one of three experiments. *p_adj \( < \) 0.05, ***p_adj \( < \) 0.001 (days), ^###p_adj \( < \) 0.001 (dose). g Effect of delayed ML-792 applied (blue) or restricted to two-day intervals (red); n = 4. ***p_adj < 0.001; one-way ANOVA with Holm–Sidak correction. h Proposed model: IRX3 suppresses SUMOylation to permit adipogenic differentiation. Bar graphs show means \(\pm \,\)SD. Source data are provided in the Source Data file.