Mammalian Sir2 Homolog SIRT3 Regulates Global Mitochondrial Lysine Acetylation

Abstract

Homologs of the Saccharomyces cerevisiae Sir2 protein, sirtuins, promote longevity in many organisms. Studies of the sirtuin SIRT3 have so far been limited to cell culture systems. Here, we investigate the localization and function of SIRT3 in vivo. We show that endogenous mouse SIRT3 is a soluble mitochondrial protein. To address the function and relevance of SIRT3 in the regulation of energy metabolism, we generated and phenotypically characterized SIRT3 knockout mice. SIRT3-deficient animals exhibit striking mitochondrial protein hyperacetylation, suggesting that SIRT3 is a major mitochondrial deacetylase. In contrast, no mitochondrial hyperacetylation was detectable in mice lacking the two other mitochondrial sirtuins, SIRT4 and SIRT5. Surprisingly, despite this biochemical phenotype, SIRT3-deficient mice are metabolically unremarkable under basal conditions and show normal adaptive thermogenesis, a process previously suggested to involve SIRT3. Overall, our results extend the recent finding of lysine acetylation of mitochondrial proteins and demonstrate that SIRT3 has evolved to control reversible lysine acetylation in this organelle.Conserved from bacteria to humans, the sirtuin family of NAD ϩ -dependent protein deacetylase/mono-ADP-ribosyltransferase enzymes controls a variety of cellular processes such as aging, metabolism, and gene silencing (18,24). It has been proposed that sirtuins mediate the longevity-promoting effects of calorie restriction (CR) in yeast, worms, flies, and mice (4,17,22,24). Seven mammalian sirtuins (SIRT1 to -7) are known (11,12,18,24). At least three sirtuins (SIRT3, SIRT4, and SIRT5) localize to mitochondria, suggesting the existence of sirtuin substrates in that organelle (19,26,28,(31)(32)(33). Several lines of evidence link SIRT3 to metabolism: SIRT3 is down-regulated in muscle from diabetic animals (37) and upregulated in white and brown adipose tissue in response to CR (33). Overexpression of SIRT3 in cells affects expression of genes involved in mitochondrial function (33). SIRT3 regulates the acetylation level and activity of acetyl-coenzyme A synthetase 2, a protein that may play a role in energy production in mammals under starvation conditions (20,31). SIRT4 is an ADP-ribosyltransferase that has been implicated in regulating amino acid-stimulated insulin secretion in mice via modification of glutamate dehydrogenase (GDH) (19). No functions have been reported for SIRT5.

Effects of intrinsic aerobic capacity, aging and voluntary running on skeletal muscle sirtuins and heat shock proteins

“…Since the rats utilized in Overmyer's study were three month old male rats, whereas our rats were nine month old females, it is possible that the age and gender difference may contribute to the different results. Nonetheless, SIRT3 has been proposed to be the most prominent deacetylase in the mitochondria (Lombard et al, 2007), and Overmyer et al suggested that the clear difference seen in the acetylation level might be due to SIRT3 activation, but not protein level per se. The higher SIRT3 level and/or activation of HCRs may promote increased deacetylation in mitochondria, thereby increasing the activity of enzymes of oxidative pathways.…”

Sirtuin 3 regulates mouse pancreatic beta cell function and is suppressed in pancreatic islets isolated from human type 2 diabetic patients

“…However, despite a likely role for increased ROS production, it is not possible from the present study to determine whether impaired insulin secretion following SIRT3 knockdown occurred as a result of a primary defect in insulin secretion or as a consequence of increased beta cell apoptosis. The precise mechanisms governing increased ROS production in INS1 cells are unclear; however, in different tissues, SIRT3 has been reported to alter the acetylation status of a number of proteins linked to ROS production [20,23,[40][41][42][43].…”

“…SIRT3 is a key regulator of mitochondrial protein acetylation status [20,37], but the precise regulatory role of this enzyme has not been examined in pancreatic islets or beta cells. We report here that SIRT3 mRNA expression is suppressed in human islets isolated from type 2 diabetes patients (compared with islets from non-diabetic controls), as well as in mouse islets and INS1 cells exposed to chronic inflammatory conditions.…”

“…Recent studies have highlighted a central role for mitochondrial dysfunction in pancreatic beta cell dysfunction [18], while other reports have demonstrated that mitochondrial proteins undergo significant levels of acetylation [19] and that stringent control of relevant acetylase and deacetylase activities is required to maintain appropriate mitochondrial function. To further elucidate the mechanisms of action of NMN, we assessed the role of the major mitochondrial SIRT, SIRT3 [20], in beta cell function. SIRT3 has not been previously reported to have functions in islets or beta cells; however, in other tissues, SIRT3 reportedly provides protection against inflammation [21] and apoptosis [22], in part by lowering reactive oxygen species (ROS) [21,23], thus indicating that SIRT3 may exert a protective role in beta cells.…”

Roles of SIRT3 in heart failure: from bench to bedside

“…Lombard et al (2007) found that no abnormalities were identified in SIRT3-deficient mice under non-stress conditions; however, the hyperacetylation of mitochondrial proteins was observed when stress was applied. Using a comprehensive multi-tissue quantitative acetylproteome analysis approach, Dittenhafer-Reed et al (2015) identified 6286 acetylation sites, of which nearly 4000 were localized to mitochondrial proteins.…”