Role Of The Ubiquitin-proteasome System And Autophagy In Regulation Of Insulin Sensitivity In Serum-starved 3T3-L1 Adipocytes

Aim:Insulin resistance is a major hallmark of type 2 diabetes mellitus (T2DM). Recent studies suggest that insulin resistance combined with pancreas beta-cell dysfunction result in impaired glucose tolerance, hyperglycemia, and T2DM. However, current understanding of cellular mechanisms of insulin resistance remains limited. Several factors like oxidative stress, inflammation, mitochondrial dysfunction, endoplasmic reticulum (ER) stress, hypoxia, and lipotoxicity have been proposed as triggers of insulin resistance. The ubiquitin-proteasome system (UPS) and macroautophagy (hereafter referred to as autophagy) are two conserved intracellular proteolytic pathways, responsible for degradation of most cellular proteins in living cells. Currently, both the UPS and autophagy have been suggested to be associated with pathogenesis of insulin resistance and diabetes. However, underlying mechanism remains largely unknown. The purpose of the present study is to investigate the impact of the UPS and autophagy on insulin sensitivity in serum-starved 3T3-L1 adipocytes.Methods:3T3-L1 preadipocytes were converted to 3T3-L1 mature adipocytes by a common method. Quantification of GFP-LC3 puncta formation was measured using immunofluorescence. Peptidase activity was detected by peptidase activity assays. The levels of symbol protein of UPS, autophagy, insulin signaling, and unfolded protein response (UPR) were detected by western blot. Glucose uptake assay was performed to detect insulin sensibility.Results:1) Serum starvation activated the UPS and autophagy and sensitized insulin action. 2) Inhibition of autophagy suppressed serum starvation-enhanced insulin action. Inhibition of the UPS did not significantly affect insulin action.3) Inhibition of the UPS led to compensative activation of autophagy. Autophagy inhibition impaired UPS function. 4) Autophagy inhibition induced ER stress and unfolded protein response. Inability of UPS exacerbated autophagy inhibition-induced detrimental impact on ER stress and UPR.Conclusion:Our results demonstrate that protein quality control maintained by the UPS and autophagy, particularly adaptive activation of autophagy is required for enhancement of insulin action by treatment with serum starvation. This new finding elucidates the functional significance of the link between autophagy, the UPS, and insulin action in adipocytes. Characterization of the mechanism by which autophagy and the UPS regulate insulin signaling should provide important information on our understanding of the pathophysiology of insulin resistance and type 2 diabetes.