MDM2 Aggravates Adipose Tissue Dysfunction Through Ubiquitin-Mediated STEAP4 Degradation

Obesity is an important cause of many chronic metabolic diseases such as metabolic syndrome,non-alcoholic fatty liver disease,type 2 diabetes,atherosclerotic heart disease,and cancer,posing a great threat to human health.The strong link between obesity and these chronic metabolic diseases has prompted a greater understanding of the function of adipocytes.Murine double minute 2(MDM2),an E3 ubiquitin ligase,has been highlighted in oncopathology as an upstream regulatory factor of the tumor suppressor p53,but little is known about its regulatory role in adipocyte biology.Here,by isolating adipose precursor cells from C57 mice in vitro and inducing them to differentiate and mature,we found that MDM2 was strongly induced during adipocyte differentiation,which indicated that MDM2 acts as an important part to the development of adipose tissue.On the other hand,the expression of MDM2 in adipose tissue could be regulated by nutritional status,and high-fat diet(HFD)significantly up-regulated the expression level of MDM2 in the adipose tissue.To further explore how MDM2 regulates the biological functions of adipose tissue,we constructed MDM2 adipocyte-specific knock-in(Mdm2-AKI)mice using CRISPR-cas9 technology.Under the condition of normal chow diet(NCD)feeding,Mdm2-AKI mice exhibited a significant increased adipose tissue mass of different parts and a series of metabolic abnormal phenotypes,such as weight gain,insulin resistance,and decreased energy expenditure.Interestingly,chronic HFD feeding resulted in distinct phenotypes of white adipose tissue in different sites of Mdm2-AKI mice.Under HFD-fed condition,epididymal white adipose tissue(eWAT),not inguinal adipose tissue(iWAT),was significantly reduced in weight and developed marked adipose tissue dysfunction,such as senescence,apoptosis,and chronic inflammation,resulting in marked insulin resistance and hepatic steatosis in Mdm2-AKI mice.Mechanistically,we used eWAT from HFD-fed Mdm2-AKI mice and control wild type(WT)mice to analyze the downstream target proteins of MDM2 by label-free quantitative proteome and ubiquitinome.Bioinformatics analysis revealed a wide range of potential roles for MDM2 in lipid and carbohydrate metabolism,and among these potential substrates,we identified the Sixtransmembrane epithelial antigen of prostate 4(STEAP4)as a true substrate of MDM2.We carried out a series of in vitro experiments to confirm that MDM2 could interact with STEAP4 and inhibit the expression of STEAP4 by ubiquitinating the K18 and K161 sites of STEAP4 for degradation.Finally,restoration of STEAP4 in eWAT of Mdm2-AKI mice rescued MDM2-induced adipose dysfunction,insulin resistance,and hepatic steatosis.In conclusion,the MDM2-STEAP4 axis in the adipose tissue was crucial for maintaining healthy adipose tissue function and ameliorating hepatic steatosis.