Succination promotes ER stress and CHOP stability: Novel role for CHOP in the metabolic function of adipocytes in diabetes

University Of South Carolina At Columbia, Columbia SC

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Abstract

? DESCRIPTION (provided by applicant): Type 2 diabetes mellitus has been diagnosed in ~21 million people in the United States and is closely correlated with obesity, prompting a need for a detailed understanding of adipocyte metabolism in the development of diabetes. The intake of excess nutrients surpasses the energy needs of the cell and leads to increased mitochondrial stress in the adipocyte. We have shown that this is associated with increased levels of the mitochondrial metabolite fumarate. Fumarate can react with cysteine thiol groups to form the chemical modification S- (2-succino)cysteine (2SC), also termed protein succination. Succination is significantly increased in the adipose tissue of type 2 diabetic mouse models (ob/ob and db/db) and in adipocytes matured in high glucose medium, resulting in impaired protein function. Endoplasmic reticulum (ER) stress is well documented in adipose tissue from diabetic mouse models; however, there is limited mechanistic insight regarding the direct cause of ER stress in adipose tissue. The ER chaperone proteins, protein disulfide isomerase (PDI) and ER oxidoreductin 1 (Ero1) contain key cysteine residues essential for proper substrate folding in the ER. In this proposal I will determine the sites of succination on these proteins and investigate if succination inhibits PDI and/or Ero1 enzymatic activity, promoting ER stress in the adipocyte during diabetes. In the presence of prolonged ER stress the unfolded protein response (UPR) will trigger the production of the pro- apoptotic protein C/EBP homologous protein (CHOP). However, CHOP is significantly increased in adipocytes matured in high glucose in the absence of PERK and IRE1 signaling or apoptosis. This suggests the presence of an alternative physiological regulator of CHOP stability under these conditions. Keap1 is a negative regulator of CHOP degradation and has been confirmed to be succinated in cancers where fumarate levels are elevated. In this proposal I will specifically investigate if succinationof Keap1 in the adipocyte contributes to CHOP stability, thereby contributing to altered CHOP function in the adipocyte in diabetes. The results from these studies will demonstrate that protein succination mechanistically link mitochondrial stress to the development of ER stress in the adipocyte and that succination of Keap1 contributes to sustained CHOP stability and adipocyte dysfunction during diabetes.