The Sir2 family of proteins were first discovered in Saccharomyces cerevisiae. The sirtuins, class III histone deacetylases (HDAC), are NAD+-dependent protein deacetylases and regulate posttranslational acyl modifications in organisms ranging from bacteria to humans (Haigis and Guarente 2006; Nakagawa and Guarente 2011). The seven sirtuin proteins (SIRT1–SIRT7) in mammals have a conserved core NAD+-binding domain and have diverse substrate proteins, intracellular localizations, and cellular functions (Anderson et al. 2014; Nakagawa and Guarente 2011).
The sirtuin family of proteins shares a conserved catalytic core of approximately 275 amino acids among most organisms. The acetylated peptide and the NAD+ are bound within a cleft that separates a small zinc-binding domain and a large Rossmann-fold domain. Several invariant amino acids are located in the cleft and are involved in substrate binding and catalysis. X-ray crystal structures of several sirtuins are now known and show overall similarity in the structure of core domains. The substrate selectivity among the different human sirtuins is attributed to the differing charge distribution and hydrophobicity in the binding cleft (Avalos et al. 2004). SIRT1 is the largest human sirtuin homologue spanning 747 amino acids, primarily due to its extensive N- and C-terminal extensions.
Sirtuins are physiological modulators regulating functions of critical proteins by posttranslational modifications. The founding member of the sirtuin family is the budding yeast protein silent information regulator 2 (Sir2). Sir2 was originally identified as a component of the chromatin-silencing complex (Sir1-Sir4), which silences transcription at telomeres, mating-type loci and ribosomal DNA (rDNA) repeats. The silencing function of Sir2 was later shown to extend replicative life span in yeast by silencing rDNA repeats and reducing the number of toxic extrachromosomal rDNA circles (Chalkiadaki and Guarente 2015). Sirtuin activity is affected by conditions that alter the cellular energetics, including calorie restriction (CR), fasting, aging, exercise, obesity, and injury (Bordone and Guarente 2005; Guarente 2012). The studies in lower forms of organisms such as yeast, flies, and mice have demonstrated a key role for sirtuins in longevity extension, though some of these reports have been contradicted (Houtkooper et al. 2012). Among the various mammalian homologues of sirtuins, SIRT1 has both nuclear and cytoplasmic localization and has many substrates, and these include transcription factors such FOXO3, NF-κb, and Pgc-1α (Anderson et al. 2014). SIRT2 is localized in the cytoplasm, where it deacetylates and regulates tubulin. SIRT2 is also reported to regulate components of mitotic checkpoints and play a role in glucose homeostasis by deacetylating and activating glucose-6-phosphatase dehydrogenase (G6PD) (Wang et al. 2014). SIRT3, SIRT4, and SIRT5 are known to have mitochondrial localization and drive the fatty acid oxidation, the Krebs cycle, and the urea cycle by deacetylating and activating some of the enzymes in these pathways (Anderson et al. 2014). SIRT4 regulates glutamine catabolism, fatty acid oxidation, and lipid catabolism. SIRT4 also exhibits lipoamidase activity and negatively modulates the pyruvate dehydrogenase (PDH) complex. Other mitochondrial targets of SIRT4 include glutamate dehydrogenase, malonyl CoA decarboxylase, and heat shock protein 60. SIRT5 preferentially targets non-acetyl lysine modifications, including malonylation, succinylation, and glutarylation. SIRT5 deacetylates carbamoyl phosphate synthetase and regulates ammonia detoxification, as evidenced by increased ammonia in the fasting blood of mice genetically deficient in SIRT5. A recent study showed that SIRT5, through demalonylation of glycolytic enzymes, can positively regulate glycolysis, providing a possible link to cancer metabolism. SIRT6 and SIRT7 are nuclear, chromatin-bound proteins. SIRT6-deficient mice exhibited increased glycolysis and tumor growth, corepressed MYC transcriptional activity, and suppressed cancer metabolism. It is also reported that SIRT6-knockout mice exhibit signs of progeria and die prematurely from severe hypoglycemia. Overexpression of SIRT6 in male mice was found to extend life span, possibly by modulating insulin-like growth factor 1 (IGF1) pathway (Kanfi et al. 2012). Among the target proteins identified for SIRT6 are histone H3, C-terminal-binding protein, PARP1, and KRAB-associated protein 1 (KAP1). SIRT7 interacts with histones and RNA polymerase I to regulate rDNA expression. It may also play a role in transcriptional regulation of Myc by modulating HIF-1a and HIF-1b expression. It is also suggested that SIRT7 deacetylates the transcription factor GA-binding protein subunit β1 (GABPβ1) and regulates the expression of nuclear-encoded mitochondrial genes and protects from mitochondrial diseases. SIRT7-deficient mice develop fatty liver disease and die prematurely from cardiomyopathy (Shin et al. 2013).
Sirtuins in Health and Disease
Sirtuins have been shown to regulate life span in different organisms, including yeast, nematodes, fruit flies, and mice, though the results are controversial (Haigis and Guarente 2006; Kaeberlein et al. 1999). Sir2 has been associated with longevity modification in the yeast, and its mammalian analog, Sirt1, has been implied to do the same in mice. As Sirt1, and other sirtuins, have strong associations with metabolic activity and gene silencing, these implications are far from unfounded. Sirtuins have also been observed to prolong life when overexpressed, or in caloric limitation models, which prompted further investigation into the role of this family in aging and disease. Among the mammalian sirtuins, SIRT1 is the most extensively characterized homologue for its role in aging and disease. Caloric restriction (CR) has been shown to extend life span and retard age-related diseases. Many studies suggest a role for SIRT1 in CR-mediated extension of longevity (Hall et al. 2013). Aging has been shown to promote a pseudohypoxic state, decreased NAD+, and decreased SIRT1 activity. It is also suggested that augmentation of NAD+ levels can increase SIRT1 activity and reduce age-associated pseudohypoxic state (Gomes et al. 2013). The scenario has parallels to the metabolic status following injury as hypoxia is one of the features of injury. However, it is still debated whether the decreased mitochondrial functional decline seen with age is due to decreased SIRT1 activity or declined NAD+ regeneration due to declining mitochondrial function with age is responsible for the decreased activity of SIRT1. The same argument applies to conditions following tissue injury. Nevertheless, several studies in experimental animals show that SIRT1 does appear to promote healthy aging by protecting against several age-related pathologies including oxidative stress (Hall et al. 2013). Mice deficient in SIRT6 show severe metabolic defects and have shorter life spans, whereas SIRT6-overexpressing male mice have increased life spans. There are also some experimental evidence linkages between SIRT3 and human aging.
A significant decrease in SIRT1 mRNA and protein presence was observed following ischemia and reperfusion (I/R) injury in the heart. A major cause of cardiac remodeling and heart failure in post-myocardial infarction is I/R injury. Cardiac-specific knockdown of SIRT1 resulted in increased size of myocardial infarction area, and overexpression of SIRT1 resulted in suppression of oxidative stress and apoptosis possibly mediated by FOXO1. Furthermore, ischemic preconditioning induced cytosolic lysine deacetylation and SIRT1 deficiency resulted in lysine acetylation, and the hearts were refractive to preconditioning. It has been observed that long-term caloric restriction induced cardiac protection due to nitric oxide-dependent increase in nuclear SIRT1 levels. Though SIRT1 exerts its beneficial effect through both posttranslational and transcriptional control, during acute injury conditions, the function of SIRT1 in posttranslational modification to restore metabolic homeostasis makes it a relevant target in developing treatment strategies. In out laboratory, when rats lost more than 60% circulating blood volume and were maintained in this state for a few hours, a significant reduction in the activity of SIRT1 was observed in the heart tissues. When SIRT1 was activated by resveratrol or its specific activator SRT1720, survival was improved significantly (Ham and Raju 2016). There was a significant decrease in total ATP and mitochondrial function following the injury, and this was, at least partially, restored with SIRT1 activation. These experiments demonstrate a profound effect of SIRT1 in modulating outcome following injury, possibly by potentiating mitochondrial function (Poulose and Raju 2015).
Sirtuins are considered cell metabolic sensors because their activity may be modulated by cellular energy status as a way to compensate energy peaks or deficits (Guarente 2006). As mentioned before, sirtuins are NAD+-dependent enzymes, and the NAD+/NADH ratio has a bearing on their activity. The mitochondrial resident SIRT3 has been shown to deacetylate and thus activate succinate dehydrogenase, the Complex II of the electron transport chain (Rahman et al. 2014). SIRT4, also inside the matrix of the mitochondria, transfers ADP-ribose to glutamate dehydrogenase inhibiting production of α-ketoglutarate in the TCA cycle (Haigis et al. 2006). SIRT4 is also known to regulate insulin secretion in pancreatic beta cells. In a SIRT3 knockdown model, there were reductions in AMPK phosphorylation implicating the influence of SIRT3 in the regulation of mitochondrial biogenesis as well.
A large amount of data has involved tumorigenesis by modulating cellular stress responses and DNA repair (Chalkiadaki and Guarente 2015). Several studies support a role for SIRT1 as a tumor suppressor. SIRT1 was shown to have a feedback regulation on Myc-dependent transformation by deacetylating this proto-oncogene, though there are contradicting reports recently. Studies showed that overexpression of SIRT1 in vivo protects against metabolic syndrome-associated liver cancer by reducing DNA damage and inflammation. A moderate increase of SIRT1 protected mice from spontaneous and aging-associated cancers. Additionally SIRT1-mediated repression of HIF-1a activity was demonstrated, inhibiting growth and angiogenesis of tumors. Interestingly, other investigators show evidence for a tumor-promoting function of SIRT1 (Chalkiadaki and Guarente 2015). However, the precise molecular mechanism through which SIRT1 exerts this tumor-promoting function remains unclear. The roles of other sirtuins in cancer development are still emerging; it is too early to conclude whether sirtuins have a contributory role or inhibitory role in tumorigenesis.
Inflammation plays a central role in the pathogenesis of many diseases and health conditions, including cancer, diabetes, cardiac disease, aging, and injury. The available data suggest an anti-inflammatory role for sirtuins, especially SIRT1 and SIRT3. Several studies have shown SIRT1 to be protective against inflammation, a function that may counteract the effects of inflammatory factors such as NF-κB and TNF-α. The transcription factor NF-κb is deacetylated and inactivated by SIRT1. NF-κb directs the transcription of a number genes in inflammation. Resveratrol, an activator of SIRT1, has been shown to inhibit inflammatory responses in vitro and in vivo models. SIRT2 also is reported to be a negative regulator of NF-κB gene expression and therefore potentially of inflammation as well. SIRT6 binds many of the same promoters in the mouse genome as NF-κB subunit RelA, and it deacetylates Lys-9 of histone H3, leading to RelA destabilization and cessation of gene expression.
Sirtuins have been explored in a number of neurodegenerative diseases such as Alzheimer’s disease, Parkinson’s disease, Huntington’s disease, amyotrophic lateral sclerosis, and spinal and bulbar muscular atrophy. In primary neurons, SIRT1 promotes neurite outgrowth and increases cell survival and decreases mTOR signaling (Sebastian et al. 2012).
Sirtuin family of proteins are evolutionarily conserved from bacteria to humans and function as critical modulators of physiological functions. Sirtuins are enzymes that function either as mono-ADP-ribosyltransferase or deacylase, including deacetylase, desuccinylase, demalonylase, demyristoylase, and depalmitoylase. Through posttranslational modification of key proteins involved in metabolic pathways, they control cellular homeostasis by non-transcriptional and transcriptional regulation. The ability of sirtuins to deacetylate and control functions of proteins allows cellular metabolic machinery to utilize this family of proteins to respond to sudden metabolic demands of the cells, rather than relying on transcriptional machinery. These proteins are also critical in metabolic adaptations to caloric restriction, fasting, aging, chronic metabolic diseases, as well as injury. SIRT1 is the most studied sirtuin, and it has been shown to exert profound effects on mitochondrial function, thereby controlling cellular energetics. The biology of sirtuins and their role in health and disease are still emerging.
The work was supported (RR) by a grant (R01GM101927) from the National Institutes of Health, Bethesda, MD, USA.
- Gomes AP, Price NL, Ling AJ, Moslehi JJ, Montgomery MK, Rajman L, White JP, Teodoro JS, Wrann CD, Hubbard BP, Mercken EM, Palmeira CM, de Cabo R, Rolo AP, Turner N, Bell EL, Sinclair DA. Declining NAD(+) induces a pseudohypoxic state disrupting nuclear-mitochondrial communication during aging. Cell. 2013;155:1624–38.PubMedPubMedCentralCrossRefGoogle Scholar
- Haigis MC, Mostoslavsky R, Haigis KM, Fahie K, Christodoulou DC, Murphy AJ, Valenzuela DM, Yancopoulos GD, Karow M, Blander G, Wolberger C, Prolla TA, Weindruch R, Alt FW, Guarente L. SIRT4 inhibits glutamate dehydrogenase and opposes the effects of calorie restriction in pancreatic beta cells. Cell. 2006;126:941–54.PubMedCrossRefGoogle Scholar
- Ham PB 3rd, Raju R. Mitochondrial function in hypoxic ischemic injury and influence of aging. Prog Neurobiol. 2016. http://dx.doi.org/10.1016/j.pneurobio.2016.06.006.Google Scholar
- Shin J, He M, Liu Y, Paredes S, Villanova L, Brown K, Qiu X, Nabavi N, Mohrin M, Wojnoonski K, Li P, Cheng HL, Murphy AJ, Valenzuela DM, Luo H, Kapahi P, Krauss R, Mostoslavsky R, Yancopoulos GD, Alt FW, Chua KF, Chen D. SIRT7 represses Myc activity to suppress ER stress and prevent fatty liver disease. Cell Rep. 2013;5:654–65.PubMedPubMedCentralCrossRefGoogle Scholar
- Wang YP, Zhou LS, Zhao YZ, Wang SW, Chen LL, Liu LX, Ling ZQ, Hu FJ, Sun YP, Zhang JY, Yang C, Yang Y, Xiong Y, Guan KL, Ye D. Regulation of G6PD acetylation by SIRT2 and KAT9 modulates NADPH homeostasis and cell survival during oxidative stress. EMBO J. 2014;33:1304–20.PubMedPubMedCentralGoogle Scholar