SAT-548 Quantitative Analysis of the Succinylome in the Thyroid Tissue of High Fat Diet-Induced Hypothyroxinemia in Rats

Hypothyroidism is a common disease, and its molecular mechanism still needs further investigation. Succinylation, as a newly identified protein posttranslational modification, is found to be involved in various metabolisms and cellular processes. In the present study, we performed quantitative analysis of the lysine succinylome in the thyroid of rats with hypothyroxinemia. Hypothyroxinemia was induced in rats through the administration of a high-fat diet and then the thyroid tissues were taken out for further analysis. Affinity enrichment and liquid chromatography-tandem mass spectrometry techniques were applied for the quantitative proteome and succinylome analyses, respectively. Bioinformatic analyses were performed to decipher the differentially expressed proteins and succinylated sites, including the gene ontology (GO) annotation-based classification, Wolfpsort-based subcellular localization prediction, Kyoto Encyclopedia of Genes and Genomes (KEGG)-based functional pathway enrichment, and conserved domains of protein and succinylation sites prediction. Overall, 3869 proteins and 685 lysine succinylation sites on 250 proteins were identified. With a quantification ratio of > 1.5 as the upregulation threshold and < 1/1.5 as the downregulation threshold, we identified 129 differentially expressed proteins and 172 differentially expressed succinylation sites (7 upregulated on 5 proteins and 165 downregulated on 99 proteins). Downregulated expressed proteins were enriched in the thyroid hormone synthesis and thyroid hormone signaling pathways and were mainly localized to the mitochondria and ATPase complex. Decreased succinylated proteins were involved in diverse metabolic pathways and biological processes, including the tricarboxylic acid cycle (TCA cycle), cellular respiration, energy derivation by oxidation of organic compounds and aerobic respiration. Moreover, these decreased succinylated proteins were primarily localized to the mitochondria. To confirm the alterations in mitochondrial respiratory activities, mitochondrial oxygen consumption rates (OCRs) was further employed to verify the LC-MS/MS results. Finally, OCRs related to basal respiration, ATP production and maximal respiration were markedly blunted in the normal thyroid epithelial cells treated with palmitic acid (all p < 0.05), and the changes were reversed when the cells were treated with palmitic acid and desuccinylase together (all p < 0.05). We speculate that the thyroid differentially expressed proteins and changed succinylation levels play a potential role in the mitochondria-mediated energy metabolism in the HFD-induced hypothyroxinemia rat model.