Lipid Droplets And Mitochondria Interaction Mediated Surface Protein Transfer Regulates Cellular Oxidative Stress

Lipid droplets are spherical organelles composed of a neutral lipid core,a monolayer of phospholipid membrane,and surface proteins.Cellular oxidative stress leads to an increase in the number of lipid droplets,and an increase in lipid droplets can alleviate cellular oxidative stress,but the molecular mechanism by which lipid droplets regulate cellular oxidative stress is unclear.The main cause of oxidative stress in cells is oxidative damage to mitochondria,which will cause mitochondria to release more reactive oxygen species,which will cause mitochondrial damage,endoplasmic reticulum stress,etc.,and cause apoptosis.Studies have shown that lipid droplets and mitochondria can contact each other.We speculate that the interaction between lipid droplets and mitochondria will regulate mitochondrial damage,and then regulate cellular oxidative stress.PLIN5 is a key protein that interacts with lipid droplets and mitochondria.In this study,the regulation of oxidative stress in cells by PLIN5 was examined.The protein profile was used to analyze changes in lipid droplet surface protein components after lipid droplet-mitochondrial contact.Our study identifies the molecular mechanisms of lipid droplet-mitochondrial interactions regulating cellular oxidative stress.The main findings are as follows:1.After the cells were treated with hydrogen peroxide,the number of lipid droplets in the cells increased,and the expression level of PLIN5 increased by WB and q PCR.2.BODIPY and Mito-tracker were used to label lipid droplets and mitochondria,respectively.Lipid droplets and mitochondria increased in hydrogen peroxide-treated cells.In addition,overexpression of PLIN5 promoted lipid droplets to contact mitochondria and interfered with PLIN5 to inhibit lipid droplets from contacting mitochondria.3.Overexpression of PLIN5 can reduce mitochondrial damage and further inhibit apoptosis,while interference with PLIN5 increases mitochondrial damage and increases the rate of apoptosis.4.Four groups of over-expressed PLIN5 group,over-expressed control group,over-expressed PLIN5 + hydrogen peroxide treated group and over-expressed control + hydrogen peroxide treated group were designed to extract lipid droplet proteins,and silver staining detected band differences.The differences in protein components on the surface of lipid droplets were detected by mass spectrometry.It was found that the surface of lipid droplets contained some mitochondrial outer membrane proteins after overexpression of PLIN5,and the protein contents were significantly different,including apoptosis-related proteins BAX,BCL?XL,and AIFM2.5.The lipid droplets were extracted and the mass spectrometry results were verified by WB.The levels of BAX and BCL?XL on the surface of lipid droplets were significantly increased after overexpression of PLIN5(p <0.05).6.After over-expressing BCL?XL in cells,the ROS level of the cells decreased;after over-expression of BAX,the ROS level of the cells increased significantly(p <0.05).Construction of lipid-targeted BAX and BCL?XL vectors(PAT-BAX and PAT-BCL?XL),no significant difference in intracellular ROS levels after overexpression of PAT-BAX and PAT-BCL?XL,and no significant difference in mitochondrial respiratory oxidase expression.The above results indicate that PLIN5 is a key factor in contact between lipid droplets and mitochondria.Oxidative stress up-regulates the expression of PLIN5 and promotes contact between lipid droplets and mitochondria.Lipid droplets and mitochondria contact membrane surface protein exchange,some of the mitochondrial outer membrane proteins associated with apoptosis are transferred to lipid droplets;in addition,these apoptosis-related proteins do not affect oxidative stress and withering on the lipid droplet surface Dead process.Therefore,the contact of PLIN5-mediated lipid droplets with mitochondria reduces mitochondrial damage,inhibits cellular ROS levels and apoptosis,thereby protecting cells and maintaining cell survival.This study preliminary analyzed the molecular mechanism of lipid droplets interacting with mitochondria to regulate oxidative stress in cells,and provided new ideas for lipid droplet biology.