Methylmercury drives lipid droplet formation and adipokine expression during the late stages of adipocyte differentiation in 3T3-L1 cells.

Chronic exposure to methylmercury (MeHg) is positively associated with obesity and metabolic syndromes. However, the effect of MeHg on adipogenesis has not been thoroughly investigated. This study investigated the effects of continuous exposure to 0.5 µM MeHg on adipocyte differentiation in 3T3-L1 cells. Oil Red O staining and triglycerides (TG) assays demonstrated that MeHg enhanced the TG content in 3T3-L1 cells. MeHg enhanced the mRNA and protein expression of adipocyte differentiation markers including peroxisome proliferator-activated receptor γ, adiponectin, and fatty acid-binding protein, and their expression levels were prominent during the late stages (days 6-8) after the induction of differentiation. In addition, 0.5 µM MeHg promoted the expression of autophagy-related genes, including light chain 3 B-II and p62, after induction of differentiation. Treatment of 3T3-L1 cells with chloroquine (CQ), an autophagy inhibitor, during the early stages (days 0-2) after induction of differentiation inhibited cellular lipid accumulation in the presence of 0.5 µM MeHg. However, treatment with CQ during the late stages (days 6-8) had little effect on the MeHg-induced increase in TG content and the expression of adipocyte differentiation markers. Although the underlying mechanisms in the late stages remain to be completely elucidated, but the present data suggest that autophagy and other mechanisms play critical roles in adipogenesis during MeHg-induced differentiation. Collectively, our results suggest that continuous exposure to MeHg induces TG accumulation and expression of genes related to adipogenesis, especially during the late stages of 3T3-L1 differentiation, which may contribute to an improved understanding of MeHg-induced adipogenesis.

Protective function of the SQSTM1/p62-NEDD4 complex against methylmercury toxicity.

SQSTM1/p62, hereinafter referred to as p62, is a stress-induced cellular protein that interacts with various signaling proteins as well as ubiquitinated proteins to regulate a variety of cellular functions and cell survival. Methylmercury (MeHg) exposure increases the levels of p62, the latter playing a protective role in MeHg-induced toxicity. However, the underlying mechanism by which p62 alleviates MeHg toxicity remains poorly understood. Herein, we report the interaction of p62 with neural precursor cell expressed developmentally down-regulated protein 4 (NEDD4), a HECT E3 ubiquitin ligase. The region of p62 where NEDD4 binds is located at the proline- and arginine (PR)-rich region (amino acids: 102-119), C-terminal extension of the Phox and Bem1 (PB1) domain. To evaluate the importance of the p62-NEDD4 complex, we examined the compensation of deletion mutant (GFP-Δ102-119 p62) for the lack of endogenous p62 in MEFs. GFP-p62/p62KO cells exhibited significantly higher cell viability than GFP-Δ102-119 p62/p62KO cells after treatment with MeHg. Our findings suggest novel mechanisms to alleviate MeHg toxicity through p62-NEDD4 complex formation.