Regulation of adipogenesis via NEDDylation of PPARg and discovery of a novel anti-obesity agent to target this mechanism

Abstract

Neural precursor cell expressed developmentally down-regulated 8 (NEDD8), an ubiquitin-like protein, regulates the activity, stability, and subcellular localization of target proteins. NEDD8 is covalently conjugated to lysine residues of substrates though E1-E2-E3 enzyme cascades, which is called NEDDylation. Here, I characterized the prominent role of NEDDylation for adipogenesis. The depletion of NEDD8 decreased the expression of adipogenic genes, thereby inhibiting adipogenic differentiation. In particular, the expression of peroxisome proliferator-activated receptor gamma (PPARg), a master regulator of adipogenesis, was reduced by knocking down NEDD8, and was enhanced by over-expressing NEDD8 in 3T3-L1 cells. Moreover, PPARg covalently conjugated with NEDD8 during adipogenesis, as shown by Ni2+ pull-down assay with His-NEDD8-overexpressed 3T3-L1 cells. In addition, the stability of PPARg protein was sustained due to decreased ubiquitination during adipogenesis. Notably, mouse double minute 2 (MDM2) was found to be a specific E3 ligase of PPARg NEDDylation. Based on these results, I tested whether a NEDD8-activating enzyme inhibitor, MLN4924, had an anti-obesity effect. The results showed that MLN4924 can inhibit both adipocyte differentiation and fat accumulation in both 3T3-L1 cells and human adipocyte-derived stem cells (H-ADSCs). Consistently, MLN4924 treatment in diet-induced-obesity (DIO) mice model diminished weight gain, accompanied with recovered lipid metabolism to a normal range. Furthermore, MLN4924 treatment in DIO mice recovered glucose tolerance, insulin sensitivity, body weight, leptin level, and the expression of inflammatory cytokine genes. Taken together, these results suggest that NEDD8 leads to crucial post-translational modification to regulate the stability of PPARg proteins in adipogenesis. Accordingly, I report that MLN4924 could be developed as a therapeutic agent for obesity.