Abstract: FR-PO374
SIRT3 Regulates the Mitochondrial Lysine Acetylation and Metabolic Networks in Proximal Tubular Epithelial Cells in Renal Fibrosis
Session Information
- CKD: Mechanisms - II
November 08, 2019 | Location: Exhibit Hall, Walter E. Washington Convention Center
Abstract Time: 10:00 AM - 12:00 PM
Category: CKD (Non-Dialysis)
- 2103 CKD (Non-Dialysis): Mechanisms
Authors
- Jiang, Lei, Nanjing Medical University, Nanjing, China
- Zhang, Yu, Nanjing Medical University, Nanjing, China
- Luo, Jing, Nanjing Medical University, Nanjing, China
- Cao, Hongdi, Nanjing Medical University, Nanjing, China
- Yang, Junwei, Nanjing Medical University, Nanjing, China
Background
Proximal tubular epithelial cells (PTCs) are high energy demanded relying on mitochondrial oxidative phosphorylation as the main energy source which is disturbed in renal fibrosis. Acetylation is recognized as an important posttranslational modification for mitochondrial function. SIRT3 is the major mitochondrial protein deacetylase and regulates mitochondrial metabolic function. In this study, we investigated whether SIRT3 could mediate the progress of renal interstitial fibrosis by regulating mitochondrial acetylation.
Methods
In vivo, unilateral ureteral obstruction (UUO) or ischemia-reperfusion (I/R) were used to induce renal fibrosis. In vitro, primary tubular epithelial cells were stimulated by TGF-b1. Proteomics and acetylation proteomics were performed on PTCs separated from UUO-operated mice at day 1. Westernblot and immunofluorescence staining were used to detect the SIRT3 expression and the acetylation levels in mitochondria . Immunoprecipitation was used to analyze the acetylation level of the PDHA. Honokiol was used as SIRT3 activator. SIRT3 shRNA adenovirus was used to knockdown SIRT3 expression.
Results
SIRT3 expression was decreased and mitochondrial protein acetylation was increased in tubular epithelial cells in the early phase of renal fibrosis. We identified 1900 unique acK site across 895 proteins between sham and UUO mice. The majority of proteins with 91.84% were hyper-acetylation. The increased acetylized proteins with 26.76% were mitochondrial protein. Notably, all proteins involved in mitochondrial oxidative phosphorylation were acetylated. We found pyruvate dehydrogenase a1(PDHA1) which provides the primary link between glycolysis and the TCA cycle was hyper-acetylation at K63,K149,K267,K277,K385. Immunoprecipitation analysis further confirmed that PDHA1 was acetylated under pro-fibrosis stimulation. And the PDH activity was decreased. Inhibiting SIRT3 could amplify the acetylation of PDHA1, inhibit PDH function and aggravate mitochondrial dysfunction .Activation SIRT3 could inhibit the acetylation of PDHA1, restore the PDH activity, improve the mitochondrial function, and further ameliorate renal fibrosis.
Conclusion
SIRT3 as a regulator of lysine acetylation in mitochondria and present a mechanism for regulating metabolic pathway through PDHA1 in Proximal tubule in renal fibrosis.
Funding
- Government Support - Non-U.S.