Belonging to an NAD⁺-dependent enzyme family, sirtuins (SIRTs) have various physiological functions similar to yeast Sir2. SIRTs possess various deacylase activities including deacetylase, desuccinylase, demalonylase, deureristoylase, demyristoylase, and depalmitoylase activities while SIRT4 and SIRT6 also have mono-ADP ribosyl transferase activity (North and Verdin, 2004; Yamamoto et al., 2007; Du et al., 2011; Jiang et al., 2013; Rack et al., 2015). SIRTs mediate cell proliferation, differentiation, apoptosis, inflammation, DNA repair, metabolism, and stress response, as well as carcinogenesis (O'Callaghan and Vassilopoulos, 2017; Preyat and Leo, 2013; Zhu et al., 2019). There are seven subtypes of SIRTs (SIRT1-SIRT7), with conserved functions and structures in mammals. The core domain of each protein comprises two subunits. The large structural domain is mainly composed of conservative Rossmann folds, while the small domain consists of a spiral structure and a zinc finger structure, which is less conserved. Crucially, a gap is formed between these two domains for substrate binding and catalysis (Hoff et al., 2006). This highly conservative core makes it difficult to design subtype-specific activators or inhibitors. The identification of the crystal structure of sirtuin3 (SIRT3) makes structure-based compound design feasible. Mammalian SIRTs have different tissue specificities, subcellular localizations, activities, and functions. SIRT1, SIRT6, and SIRT7 are mainly found in the nucleus, and SIRT2 is present in the cytoplasm. SIRT3, SIRT4, and SIRT5 are predominantly localized to mitochondria (Michishita et al., 2005). However, recent studies have shown that SIRT1 localization depends on the cellular context (Jin et al., 2007; Mattagajasingh et al., 2007; Tanno et al., 2007; Byles et al., 2010). SIRT1 is mainly located in the nucleus of normal cells but is found in the cytoplasm of cancerous cells (Byles et al., 2010). SIRT3 is mainly located in mitochondria, however, its presence in the nucleus and cytoplasm has also reported (Scher et al., 2007; Sundaresan et al., 2008; Iwahara et al., 2012) (Table 1).

In recent years, SIRT3-related research has intensified since its link to a longer lifespan in humans and its critical localization to the mitochondrial compartment have been recognized (Hurst et al., 2002). Age-associated pathologies and longevity are related to SIRT3 in humans. SIRT3 is the only member with strong NAD⁺-dependent deacetylase activity among the mitochondrial SIRTs (Lombard et al., 2007). SIRT3 consists 399 amino acids (Yang et al., 2010), including an N-terminal domain with a 25-amino-acid mitochondrial targeting sequence (MTS) (Onyango et al., 2002), a catalytic region, and a C-terminal domain (Figure 1). SIRT3 translocates to mitochondria with the N-terminal MTS and is cleaved into a 28 kD protein by mitochondrial processing peptidase (Schwer et al., 2002; Iwahara et al., 2012) to induce deacetylase functions (Smith et al., 2008). SIRT3 deacetylates various proteins that not only regulate mtDNA replication, transcription, translation, fatty acid oxidation, and amino acid metabolism, but also modulate enzymes that participate in the tricarboxylic acid (TCA) cycle, and regulate the activity of the electron transport chain (Kumar and Lombard, 2015; Lee, 2019). SIRT3 also plays a unique regulatory role by mediating interactions between mitochondria and intracellular signaling. SIRT3 is involved in the pathogenesis of diverse disorders including cancer, diabetes, neurodegeneration, cardiac hypertrophy, and liver steatosis. Surprisingly, SIRT3 acts as a double-edged sword in cancer. Thus, the development of compounds that alter SIRT3 activity has gained increasing attention from scientists. We will comprehensively review the mitochondrial substrates and functions of SIRT3, highlighting its dual role in various cancers, and summarize the activators and inhibitors of SIRT3 in this article.

SIRT3 is closely involved in the metabolism of amino acids, glycosides, and lipids in mitochondria (Hiromasa et al., 2004; Lombard et al., 2007). As an emerging, pivotal regulator of oxidative stress, SIRT3 regulates reactive oxygen species (ROS) production via deacetylation of key enzymes (Bause and Haigis, 2013). SIRT3 mediates the activities of isocitrate dehydrogenase (IDH) and manganese superoxide dismutase (MnSOD/SOD2) which are two major oxidative stress-responsive proteins, and SIRT3 is recognized as an important ROS scavenger in cells. Recent studies revealed that SIRT3 may regulate antioxidant-related enzymes by deacetylating and activating forkhead transcription factor (FOXO3a) (Sundaresan et al., 2009; Bause and Haigis, 2013; Tseng et al., 2013; Rangarajan et al., 2015). SIRT3 directly deacetylates and activates the enzymes involved in processes regulating mitochondrial energy metabolism, including fatty acid oxidation, the Krebs cycle, and the respiratory chain (Huang et al., 2010; Verdin et al., 2010; Torrens-Mas et al., 2017). The pyruvate dehydrogenase complex (PDC) is deacetylated by SIRT3, which funnels pyruvate to participate in the Krebs cycle, and protein kinase B is activated so that glucose uptake in glycolysis is accelerated (Hiromasa et al., 2004; Xu et al., 2019a). SIRT3 also deacetylates and activates long-chain acyl-CoA dehydrogenase (LCAD) (Hirschey et al., 2010), acetyl-CoA synthetase 2(AceCS2) (Schwer et al., 2006; Hirschey et al., 2010), and 3-hydroxy-3-methylglytaryl-CoA synthetase (HMGCS2) (Hirschey et al., 2010; Shimazu et al., 2010) to regulate lipid metabolism and fatty acid oxidation (Hirschey et al., 2010). Moreover, amino acid metabolism can be mediated by SIRT3 through deacetylation of glutamate dehydrogenase (GDH) (Lombard et al., 2007). SIRT3 deacetylates ornithine transcarbamylase (OTC) to regulate the urea cycle (Hallows et al., 2011).

Furthermore, SIRT3 promotes the progression of the TCA cycle through deacetylation of succinate dehydrogenase (SDH (Cimen et al., 2010)) and IDH (Meng et al., 2019). SIRT3 deacetylates numerous complex I–V subunits to regulate the oxidative respiratory chain (Ahn et al., 2008; Finley et al., 2011a) and ATPase (Wu et al., 2013). In addition, SIRT3 participates in mitochondrial biogenesis and dynamics (Torrens-Mas et al., 2017). SIRT3 also participates in the maintenance of mitochondrial quality. For instance, it was reported that SIRT3 coordinates the mitochondrial unfolded protein response (mtUPR) and upregulates mitochondrial autophagy (Papa and Germain, 2014). The mtUPR promotes a complex transcription program ultimately increasing mitochondrial integrity and fitness in response to oxidative proteotoxic stress. The activation of the SIRT3 axis of the unfolded protein response is linked to metastasis (Kenny et al., 2017). SIRT3 is the key coordinator of the mtUPR induced by mitochondrial protein cytotoxic stress (O'Callaghan and Vassilopoulos, 2017; Xu et al., 2019b; Weng et al., 2020). Moreover, SIRT3 can prevent mitochondrial dysfunction through the regulation of the mitochondrial permeability transition pore (mPTP) (Hafner et al., 2010; Torrens-Mas et al., 2017). Taken together, through reversible acetylation of mitochondrial proteins, SIRT3 plays a key role in mitochondrial metabolism. The main mitochondrial targets of SIRT3 are shown in Figure 2.

SIRT3 mediates numerous biological and pathophysiological activities through the regulation of various biological functions, including repair, energy homeostasis, oxidative stress tolerance, apoptosis, enhanced longevity, anti-inflammatory responses, and other mechanisms (Ansari et al., 2017).

As a major repair enzyme in the base excision repair pathway, 8-oxoguanine DNA glycosylase (OGG1) is a newly discovered target of SIRT3. Cheng et al. found that SIRT3 deacetylates OGG1 to regulate its incision activity and decrease its degradation. Mitochondrial DNA damage repair, mitochondrial integrity, and apoptotic cell death induced by oxidative stress are closely related to the acetylation- and turnover-related regulation of OGG1 by SIRT3 (Cheng et al., 2013). Furthermore, SIRT3 has been reported to protect diverse types of cells from programmed cell death initiated by genotoxicity or oxidative stress by regulating ROS production and the mPTP and preventing Bax translocation to mitochondria (Pellegrini et al., 2012). Another study showed that SIRT3 acts as an anti-apoptotic protein. It was reported that SIRT3 can protect cells from death when mitochondrial NAD⁺ levels are severely decreased, in response to DNA damage (Yang et al., 2007).

SIRT3 is one of the most significant regulators of metabolism (Ansari et al., 2017). Ahn et al. implemented SIRT3 knockout mice to reveal the roles of SIRT3 in physiology. They demonstrated that SIRT3 is crucial to the regulation of basal ATP levels and energy homeostasis (Ahn et al., 2008). SIRT3 can regulate different enzymes including manganese dismutase, SOD2, and catalase, which are important in regulating ROS levels (Merksamer et al., 2013). More about the role of SIRT3 in metabolism is described in Part 2.

Given the high levels of SIRT3 in long-lived individuals, scientists suspect that SIRT3 is potentially associated with longevity. Initially, the discovery of unique single nucleotide polymorphisms (SNPs) linked to centenarians aroused interest in SIRT3 and its role in the human life span (Hurst et al., 2002; Rose et al., 2003; Bellizzi et al., 2005). Males carrying the G477T transversion in exon 3 of SIRT3 live longer than average. However, this nucleotide transition does not change the amino acid sequence and therefore is a silent change (Rose et al., 2003). The second SNP involves the variable number of tandem repeats (VNTR) region within SIRT3 intron 5, and specific VNTR polymorphisms show increased SIRT3 expression and have been linked to increased longevity (Bellizzi et al., 2005). The third SNP, unlike the previous two SNPs, has been associated with an increased risk of age-related metabolic syndrome. Some SNPs induce amino acid substitutions in the conserved deacetylase region, thereby reducing SIRT3 deacetylase activity (Hirschey et al., 2011). Therefore, specific SNPs of SIRT3 are associated with increased human lifespan, but more research is required to determine the mechanisms by which various SNPs affect human lifespan (Kincaid and Bossy-Wetzel, 2013). In addition, it was reported that a product of the FOXO transcription factor family helps to regulate the longevity of nematodes (Shi et al., 2005). Among FOXO family members, FOXO3a is a key substrate for SIRT3. SIRT3 interacts with FOXO3a to activate antioxidant genes, such as MnSOD and catalase, the products of which can reduce the level of ROS and positively affect disorders such as interstitial fibrosis and cardiac hypertrophy (Sundaresan et al., 2009).

SIRT3 not only has NAD⁺ dependent deacetylase activity but also catalyzes the removal of long-chain fatty acyl groups. These include myristoyl, palmitoyl, and others (Feldman et al., 2013). Acylation of long-chain fatty acids can regulate biological factors, including membrane associations, protein-protein interactions, and subcellular location. Gai et al. demonstrated that SIRT3 is an effective demyristoylase and is an expeditious depalmitoylase. Furthermore, they analyzed the crystal structures of SIRT3 in compound material containing a palmitoylated peptide or myristoylated H3K9 (Gai et al., 2016). The palmitoyl and myristoyl groups bound to the C-pocket and an allosteric site, respectively (Gai et al., 2016). However, the effects of demyristoylation, depalmitoylation, and other modifications on the functions of SIRT3 remain unclear.

Cancer is the most common cause of death worldwide. Carcinogenesis is characterized by cell enrichment, excessive proliferation, apoptosis resistance, and metabolic instability. Otto Warburg found that cancer cells have different metabolism patterns than normal tissues in the 1920s. Cancer cells consume many nutrients and rewire their metabolic processes to enable biosynthesis, which allows them to meet their energy and biomass production requirements (Finley and Haigis, 2012). Elevated ROS production, which is frequent in human malignancies, can promote tumorigenesis through a variety of mechanisms (Liou and Storz, 2010). ROS regulate cell proliferation, survival, differentiation, metabolism, and inflammation (Liou and Storz, 2010). Oxidative damage to DNA, lipids, and proteins can disrupt cellular processes, and further promote tumor progression (Finley and Haigis, 2012). Cancer is characterized by the reprogramming of cellular energy metabolism. SIRT3, regarded as a novel and potential therapeutic target, is shown to be involved in most of these cancer pathways. In addition, SIRT3 is abnormally expressed in a variety of cancers as shown in Figure 3. In this review, we emphasize the role of SIRT3 in tumorigenesis and cancer therapy.

The function of SIRT3 in aging, neurological disease, cancer, and stress resistance has drawn much attention. By reversible protein lysine deacetylation, SIRT3 can regulate mitochondrial activity and biosynthetic processes such as the TCA cycle, oxidative stress, glucose, fatty acid metabolism, and apoptosis. The therapeutic potential of SIRT3 in multiple cancers makes it an attractive drug target. The critical biological functions of SIRT3 have sparked research into small molecule modulators that can control its activity (Villalba and Alcaín, 2012). Currently, there is no specific and selective SIRT3 activator or inhibitor due to the conserved molecular structure of the SIRTs family and the mode of SIRT3 activation. In this section, recent advances in the discovery of SIRT3 activators and inhibitors are reviewed.

SIRT3 pathways are involved in pathophysiological processes associated with metabolic disorders. In addition to cancer, SIRT3 also plays important functions in multiple age-associated diseases, including hearing loss, obesity, diabetes, insulin resistance, neurodegeneration, cardiac hypertrophy, liver steatosis, and glucose homeostasis disorders (Giblin et al., 2014).

Since mitochondrial dysfunction is the basis of the pathogenesis of most neurodegenerative diseases, it is not surprising that the mitochondrial deacetylase SIRT3 plays a key role in several brain diseases, such as Parkinson’s disease (PD), Alzheimer’s disease (AD), Huntington’s disease (HD), and stroke (Gomes et al., 2020). Cheng et al. reported that SIRT3 serves as a vital molecule mediating neuroprotective adaptive stress responses. In HD and epilepsy models, hippocampal neurons and striatal vulnerability are increased in SIRT3^−/− mice. In the mouse temporal lobe epilepsy model, deficiency of or reduction in SIRT3 leads to hyperacetylation of some mitochondrial proteins, including SOD2 and cyclophilin D (Cheng et al., 2016). Thus, SIRT3 is required for neuroprotection because it improves mitochondrial function.

Cardiovascular diseases (CVDs) are the leading cause of death globally. Mitochondrial dysfunction plays a crucial role in CVD pathogenesis (Chistiakov et al., 2018). SIRT3 is a major and crucial mitochondrial NAD⁺-dependent deacetylase, that regulates most mitochondrial lysine acetylations. SIRT3 involved in cardiovascular physiology and pathology (Bugger et al., 2016). It has been reported that enhanced expression of SIRT3 protects myocytes from genotoxic and oxidative stress-mediated cell death, in part by blocking the transfer of Bax to mitochondria, as a stress-responsive deacetylase (Sundaresan et al., 2008). SIRT3 regulates the interaction of BaX and Ku70 by deacetylating Ku70. Sundaresan et al. proved that SIRT3 can reduce ROS levels and block cardiac hypertrophy. It was found that SIRT3 activates SOD2 and catalase to block cardiac hypertrophy in primary cardiomyocytes cultures, thereby decreasing ROS levels (Sundaresan et al., 2009). The hearts of SIRT3 knockout mice show accelerated signs of aging, manifested by cardiac hypertrophy and fibrosis. Thus, SIRT3 plays an indispensable role in preventing mitochondrial dysfunction and cardiac hypertrophy during aging (Hafner et al., 2010). In general, SIRT3 largely plays a protective role in myocardia.

Liver diseases have high morbidity, and there are more than 800 million cases (Marcellin and Kutala, 2018). SIRT3 levels in the liver are increased by fasting (Hirschey et al., 2010) and decreased by a high-fat diet (HFD) (Bao et al., 2010; Kendrick et al., 2011). In SIRT3 knockout mice, HFD feeding increases the acetylation of several hepatic proteins. This finding proves that SIRT3 is a key factor regulating hepatic mitochondrial function (Kendrick et al., 2011). Bao et al. found that palmitate enhances ROS and increases hepatocyte cell death in the absence of SIRT3. Restoring the level of SIRT3 and/or treatment with N-acetylcysteine, can ameliorate these adverse effects. SIRT3 ameliorates hepatic lipotoxicity, but paradoxically, downregulation of this adaptive protection occurs upon exposure to high fat in the liver (Bao et al., 2010).

SIRT3 can regulate a variety of mitochondrial functions to regulate fuel utilization. SIRT3 germline knockout mice develop various forms of metabolic dysfunctions at an accelerated rate, including obesity, hepatic steatosis, and insulin resistance (Hirschey et al., 2011). Zhang et al. found that SIRT3 is a positive regulator of macroautophagy and chaperone-mediated autophagy in adipocytes that promotes lipid mobilization via activating AMPK. SIRT3 might be a promising target for the treatment of obesity and related metabolic dysfunction (Zhang et al., 2020). It is crucial to preserve mitochondrial health to prevent insulin resistance and type 2 diabetes mellitus (T2DM) during aging. A major and early characteristic of the pathogenesis of T2DM is insulin resistance in skeletal muscle (DeFronzo and Tripathy, 2009). Jing et al. reported that mitochondrial dysfunction, increased oxidative stress, and JNK activation can be caused by decreased expression of SIRT3, which in turn induces impaired insulin signaling (Jing et al., 2011). The key factors mediating pancreatic β cell impairment in T2DM are chronic inflammation and mitochondrial dysfunction. SIRT3 mediates ROS production and plays a part in anti-inflammatory effects.

SIRT3 is an NAD⁺ -dependent deacetylase mainly located in mitochondria, that can regulate the activity of numerous substrate proteins to affect the fate of mitochondria, the nucleus, and even the entire cell. SIRT3 regulates oxidative stress, amino acid metabolism, fatty acid oxidation, electron transport, and the TCA cycle through deacetylating various substrate proteins (Kumar and Lombard, 2015). SIRT3 can coordinate complete shifts in mitochondrial metabolism and has a potential impact on diseases, especially CVDs and cancer (Chen et al., 2014). Studies using different cancer models to determine the precise role of SIRT3 in tumorigenesis have drawn different conclusions. SIRT3 plays a dual role in cancer, and whether it promotes or suppresses tumors probably depends on the type of cancer and the status of intracellular signaling pathways (Xiong et al., 2016). It appears that SIRT3 may prevent cell death by inhibiting oxidative stress; however, other studies have reported its function in promoting apoptosis (Torrens-Mas et al., 2017). By studying the mechanistic differences in various cancer types, our understanding of the carcinogenic and anticancer effects of SIRT3 may deepen, which can aid the development of novel cancer therapeutic strategies. Further characterization of the mitochondrial substrate protein of SIRT3 will contribute to a deeper understanding of cancer tumorigenesis. Such research may also help develop novel therapeutic strategies and improve patient outcomes. The identification of drugs that alter SIRT3 activity has become an important task. Despite the publication of a few relevant reports, no SIRT3 activator or inhibitor with excellent specificity and selectivity is currently being tested in a clinical trial. The activation pattern of SIRT3 has not been fully elucidated, which limits the development of SIRT3 inhibitors and activators. Therefore, monotherapies and combination therapies targeting SIRT3 hold substantial promise for drug development.