The transcription factor PPARG (peroxisome proliferator-activated receptor gamma) is a nuclear receptor involved in lipid metabolism, insulin resistance, and inflammation. It also plays an essential role in placental formation, as demonstrated in murine models. In the human placenta, PPARG is strongly expressed by trophoblasts and directly contributes to their differentiation: mononucleated villous cytotrophoblasts fuse to renew the syncytiotrophoblast, the multinucleated layer that mediates exchanges between the maternal and fetal circulations. Abnormal expression or activity of PPARG is thus associated with placental disorders such as pre-eclampsia and intrauterine growth restriction. Notably, the receptor contains a ligand-binding domain (LBD), whose rare mutations with autosomal dominant transmission cause familial partial lipodystrophy type 3 (FPLD3), characterized by an abnormal distribution of adipose tissue and severe metabolic complications.
The authors sought to assess the impact of three LBD mutations of PPARG on two processes essential to placental development: cell fusion and cell migration. The E352Q mutation, initially described in a case of placental abnormalities that led to neonatal death, was studied in the NIH/3T3 cell line, which lacks endogenous PPARG, as well as in human trophoblasts after silencing of the endogenous gene by RNA interference followed by reconstitution with vectors encoding the wild-type or mutant form. The R262G and L319X mutations were examined in skin fibroblasts derived from patients with FPLD3, compared with control fibroblasts, using wound-healing assays. A structural analysis of the receptor complemented these functional approaches.
The results show that the E352Q mutation significantly reduces the transcriptional activity of PPARG, with or without agonist. Notably, treatment with the agonist GW1929 almost completely restores this activity in vitro, a profile not reported for other FPLD3-associated mutations. This mutation also decreases the fusion of villous cytotrophoblasts into syncytiotrophoblast. In fibroblasts, the R262G and L319X mutations strongly inhibit cell migration, with the effect of L319X being greater than that of R262G. Mutant fibroblasts display a flattened rather than spindle-shaped morphology, along with an increase in focal adhesions labeled by vinculin, a protein described as a suppressor of migration, which is consistent with the observed reduction.
The structural analysis sheds light on these observations: the mutated residues participate in interaction networks that stabilize the ligand-binding pocket and the dimerization surface with RXRα. In conclusion, this work establishes that a single missense or nonsense mutation in the ligand-binding domain of PPARG is sufficient to significantly inhibit cell fusion and migration processes.