烟酰胺腺嘌呤二核苷酸和Sirtuins在衰老和疾病中的作用

烟酰胺腺嘌呤二核苷酸(nicotinamide adenine dinucleotide, NAD⁺)作为氧化还原反应的重要辅酶,是能量代谢的核心。NAD⁺也是非氧化还原NAD⁺依赖性酶的共底物,包括沉默信息调节因子(Sirtuins)、聚ADP-核糖聚合酶(poly ADP-ribose polymerases, PARPs)、CD38/CD157胞外酶等。NAD⁺已成为细胞信号转导和细胞存活的关键调节剂。最近的研究表明,Sirtuins催化多种NAD⁺依赖性反应,包括去乙酰化、脱酰基化和ADP-核糖基化。Sirtuins催化活性取决于NAD⁺水平的高低。因此,Sirtuins是细胞代谢和氧化还原状态关键传感器。哺乳动物中已经鉴定并表征了7个Sirtuins家族成员(SIRT1-7),其参与炎症、细胞生长、生理节律、能量代谢、神经元功能、应激反应和健康衰老等多种生理过程。本文归纳了NAD⁺的生理浓度及状态、NAD⁺     

Mammalian sirtuins and energy metabolism

Sirtuins are highly conserved NAD+-dependent protein deacetylases and/or ADP-ribosyltransferases that can extend the lifespan of several lower model organisms including yeast, worms and flies. The seven mammalian sirtuins, SIRT1 to SIRT7, have emerged as key metabolic sensors that directly link environmental signals to mammalian metabolic homeostasis and stress response. Recent studies have shed light on the critical roles of sirtuins in mammalian energy metabolism in response to nutrient signals. This review focuses on the involvement of two nuclear sirtuins, SIRT1 and SIRT6, and three mitochondrial sirtuins, SIRT3, SIRT4, and SIRT5, in regulation of diverse metabolic processes.     

Comparative interactomes of SIRT6 and SIRT7: Implication of functional links to aging

Sirtuins are NAD⁺‐dependent deacetylases that regulate a range of cellular processes. Although diverse functions of sirtuins have been proposed, those functions of SIRT6 and SIRT7 that are mediated by their interacting proteins remain elusive. In the present study, we identified SIRT6‐ and SIRT7‐interacting proteins, and compared their interactomes to investigate functional links. Our interactomes revealed 136 interacting proteins for SIRT6 and 233 for SIRT7 while confirming seven and 111 proteins identified previously for SIRT6 and SIRT7, respectively. Comparison of SIRT6 and SIRT7 interactomes under the same experimental conditions disclosed 111 shared proteins, implying related functional links. The interaction networks of interactomes indicated biological processes associated with DNA repair, chromatin assembly, and aging. Interactions of two highly acetylated proteins, nucleophosmin (NPM1) and nucleolin, with SIRT6 and SIRT7 were confirmed by co‐immunoprecipitation. NPM1 was found to be deacetylated by both SIRT6 and SIRT7. In senescent cells, the acetylation level of NPM1 was increased in conjunction with decreased levels of SIRT6 and SIRT7, suggesting that the acetylation of NPM1 could be regulated by SIRT6 and SIRT7 in the aging process. Our comparative interactomic study of SIRT6 and SIRT7 implies important functional links to aging by their associations with interacting proteins. All MS data have been deposited in the ProteomeXchange with identifiers PXD000159 and PXD000850 ( http://proteomecentral.proteomexchange.org/dataset/PXD000159 , http://proteomecentral.proteomexchange.org/dataset/PXD000850 ).     

SIRT3 and cancer: tumor promoter or suppressor?

Sirtuins (SIRT1-7), the mammalian homologues of the Sir2 gene in yeast, have emerging roles in age-related diseases, such as cardiac hypertrophy, diabetes, obesity, and cancer. However, the role of several sirtuin family members, including SIRT1 and SIRT3, in cancer has been controversial. The aim of this review is to explore and discuss the seemingly dichotomous role of SIRT3 in cancer biology with particular emphasis on its potential role as a tumor promoter and tumor suppressor. This review will also discuss the potential role of SIRT3 as a novel therapeutic target to treat cancer.     

Will the real aging Sirtuin please stand up?

Initial studies linking Sirtuins to longevity in yeast initiated what is now a rich vein of aging research that is full of promise and fraught with controversy. Missing was a demonstration that enhanced Sirtuin expression extends lifespan in mammals. Now Kanfi et al. provide the evidence but with an interesting plot twist – the lesser known SIRT6 is the longevity factor.While Sirtuins, a highly conserved class of protein deacetylases, have been linked to longevity in a variety of model organisms and are among the most studied proteins in the context of aging, their role remains controversial. Increased expression of the founding member of the class, yeast SIR2, was reported to extend replicative lifespan over a decade ago and this was followed by reports that overexpression of SIR2 orthologs leads to enhanced longevity in worms and flies ^{1, 2}. More recent reports have called the worm and fly data into question ^{3, 4} and while increased Sir2 activity may enhance longevity under certain conditions in all three organisms, mechanistic studies have not converged on a specific activity coupled to aging.Seven mammalian Sir2 homologs (SIRT1-SIRT7) have been identified. A myriad of activities in a range of mammalian tissues have been attributed to SIRT1, a protein most structurally related to yeast Sir2, often linked to protection from the onset of chronic disease states. Yet transgenic mice with increased SIRT1 expression are not long-lived ⁵, further adding to what is becoming a great science mystery. Until recently, while SIRT1 was being intensely studied, the other Sirtuins were mostly waiting in the wings. In the last few years, however, many of them have been linked to interesting biological functions and, with the recent report by Kanfi et al. ⁶, SIRT6 assumes a central role in aging.The primary finding of Kanfi et al. is that transgenic mice overexpressing SIRT6 have enhanced longevity, but only in males ⁶. While the longevity enhancement is modest (15% increase in median lifespan), the data is rigorous. Similar effects were seen in two different founder transgenic lines, large numbers of mice were included in the study and a mixed genetic background was selected for the study with equal contribution of two long-lived inbred backgrounds. The authors also attempted to address whether the longevity benefit results from retarded aging, as opposed to protection from cancer, a common cause of mortality in mice. While a challenging issue to resolve in a single study, tantalizing evidence is provided that SIRT6 is indeed a modulator of aging. For instance, if longevity is enhanced simply by eliminating the most common type of cancer, lung in these mice, then a post-mortem analysis would yield a skewing of the tumor spectrum. Yet no significant differences were seen in the incidence of different tumors. Moreover, other age-related pathologies were evident in equal proportion. These findings are consistent with a model whereby the aging process is delayed, with animals developing and succumbing to the same diseases later in life (Figure 1). Coupled with earlier findings that SIRT6 overexpression in mice (1) leads to protection from diet-induced obesity and (2) suppresses expression of NF-κB-dependent inflammatory markers, and moreover that SIRT6 expression is induced by caloric restriction in rats, it is fair to say that the deacetylase has certainly entered the realm of top-tier candidate modulatory factors of aging.Open in a separate windowFigure 1SIRT6 expression and longevity. Decreased SIRT6 expression leads to genome instability, as well as progeroid phenotypes that resemble accelerated aging. In contrast, increased SIRT6 expression alters cell and physiological phenotypes in a manner that promotes enhanced longevity.Long debated is whether mouse or human models of progeria, diseases with a rapid onset of a subset of aging pathologies, are truly accelerated aging syndromes ⁷. One argument against this hypothesis was that the genetic interventions giving rise to progeroid syndromes are non-overlapping with those linked to enhanced longevity. This disconnect is starting to dissolve as genes that go both ways are being identified. p53 is arguably the first example; constitutive p53 activation induces progeria phenotypes ⁸, whereas enhanced p53 activity under normal regulatory control leads to lifespan extension ⁹. With the study by Kanfi et al. ⁶, SIRT6 emerges as a new member of this short list of genes since it was previously shown that SIRT6^−/− mice have metabolic defects and rapidly present with abnormalities often associated with aging, such as kyphosis and loss of subcutaneous fat ¹⁰. Thus, reduced SIRT6 expression may accelerate aspects of aging whereas enhanced SIRT6 expression may delay them.A primary question is why increased SIRT6 expression enhances longevity, and several models are possible in addition to regulation of NF-κB. First are the metabolic consequences of enhanced SIRT6 activity, which are further elaborated in Kanfi et al. by examination of metabolic changes during aging of mice on a normal ad lib diet ⁶. First, Sirt6-transgenic mice are able to maintain healthy glucose metabolism later in life than controls, a feature often associated with longevity. However, both males and females maintain better metabolic function, therefore, this finding, while it may be linked, is separable from the dimorphic effects on lifespan. A further clue comes from the finding that IGF-1 levels are reduced in serum of male but not female overexpressing SIRT6 and that this is coupled to a corresponding increase in IGFBP-1 levels. Interestingly, these changes place the males in line with the females with regard to both factors. The downstream consequences of reduced IGF-1 levels are most apparent in white adipose tissue, which show significant reductions in AKT phosphorylation again only in males. Metabolic changes in fat tissue are increasingly linked to lifespan. These findings raise the intriguing possibility that the enhanced longevity in male Sirt6-transgenic mice occur because they have metabolic profiles more like that of females.Whole-genome microarray analysis of liver tissue from mice overexpressing SIRT6 raises another intriguing possible mechanism linked to metabolism. First, the biggest gene expression changes were apparent in male transgenic livers and, importantly, many of the observed differences overlap with those of calorie-restricted mice. Since those mice also have reduced serum IGF-1 and increased IGFBP-1 levels ¹¹, gene expression changes could be cause or consequence of changes to the endocrine axis. Moreover, these findings support the hypothesis that SIRT6 overexpression phenocopies aspects of calorie restriction in males. Another possibility that Sirt6-transgenic mice eat less, mimicking calorie restriction in a more direct fashion, can be largely discounted since an early study reported equal food consumption ¹².While SIRT1 is relatively promiscuous in its choice of substrate for deacetylation, finding targets for SIRT6 proved a more challenging endeavor. Recently, however, the identification of histone H3K9 and K56 may explain roles identified for SIRT6 in the DNA damage response and in telomere maintenance ^{13, 14}. Cells from SIRT6^−/− mice experience a dramatic elevation in genome instability indicative of defective double strand break (DSB) repair ¹⁰ and by contrast, enhanced SIRT6 activity stimulates DSB repair, particularly in response to oxidative stress ¹⁵. Deacetylation of H3K56 is linked to enhanced DNA repair, as is mono-ADP ribosylation of PARP, an alternate NAD⁺-dependent enzymatic activity of SIRT6. SIRT6 also associates with telomere heterochromatin, where it deacetylates both H3K9 and H3K56 ^{13, 16}. Reduced SIRT6 expression in primary fibroblasts leads to a range of replication-dependent defects in telomere maintenance, promoting premature senescence. Functions of SIRT6 either at sites of DSBs or at telomeres could underlie both progeroid and/or enhanced longevity phenotypes.While SIRT1 resembles yeast Sir2 from a structural perspective, it may be SIRT6 that is a closer functional ortholog. In their respective organisms, expression levels of each correlate with longevity. Moreover, both are most closely linked with histone deacetylation and have roles in genome maintenance both at telomeres and in response to DNA damage. Both also may link genome maintenance to nutrient and stress conditions in the cell. A challenge moving ahead will be to link SIRT6 enzymatic functions and cellular properties to the metabolic and longevity phenotypes apparent in the transgenic and knockout mice.The studies of Kanfi et al. place SIRT6 in a group of genes that when genetically modified can lead to enhanced mammalian longevity. However, they do not rule out roles for other Sirtuins in aging. For instance, while whole body overexpression of SIRT1 appears not to enhance lifespan, given the range of its functions in different tissues, more informative may be the lifespan phenotypes of mice overexpressing SIRT1 in specific tissues. In addition, little is known about other Sirtuins and given interesting metabolic roles especially for those localized to the mitochondria, it would not be surprising if they affect aging as well ². The role of Sirtuins in aging remains controversial and undoubtedly twists and turns are yet to come, but the report by Kanfi et al. makes the case for Sirtuins and aging far more compelling, at least in the case of SIRT6.     

Mechanism of Inhibition of the Human Sirtuin Enzyme SIRT3 by Nicotinamide: Computational and Experimental Studies

Sirtuins are key regulators of many cellular functions including cell growth, apoptosis, metabolism, and genetic control of age-related diseases. Sirtuins are themselves regulated by their cofactor nicotinamide adenine dinucleotide (NAD⁺) as well as their reaction product nicotinamide (NAM), the physiological concentrations of which vary during the process of aging. Nicotinamide inhibits sirtuins through the so-called base exchange pathway, wherein rebinding of the reaction product to the enzyme accelerates the reverse reaction. We investigated the mechanism of nicotinamide inhibition of human SIRT3, the major mitochondrial sirtuin deacetylase, in vitro and in silico using experimental kinetic analysis and Molecular Mechanics-Poisson Boltzmann/Generalized Born Surface Area (MM-PB(GB)SA) binding affinity calculations with molecular dynamics sampling. Through experimental kinetic studies, we demonstrate that NAM inhibition of SIRT3 involves apparent competition between the inhibitor and the enzyme cofactor NAD⁺, contrary to the traditional characterization of base exchange as noncompetitive inhibition. We report a model for base exchange inhibition that relates such kinetic properties to physicochemical properties, including the free energies of enzyme-ligand binding, and estimate the latter through the first reported computational binding affinity calculations for SIRT3:NAD⁺, SIRT3:NAM, and analogous complexes for Sir2. The computational results support our kinetic model, establishing foundations for quantitative modeling of NAD⁺/NAM regulation of mammalian sirtuins during aging and the computational design of sirtuin activators that operate through alleviation of base exchange inhibition.     

Over expression of wild type or a catalytically dead mutant of Sirtuin 6 does not influence NFκB responses

SIRT6 is involved in inflammation, aging and metabolism potentially by modulating the functions of both NFκB and HIF1α. Since it is possible to make small molecule activators and inhibitors of Sirtuins we wished to establish biochemical and cellular assays both to assist in drug discovery efforts and to validate whether SIRT6 represents a valid drug target for these indications. We confirmed in cellular assays that SIRT6 can deacetylate acetylated-histone H3 lysine 9 (H3K9Ac), however this deacetylase activity is unusually low in biochemical assays. In an effort to develop alternative assay formats we observed that SIRT6 overexpression had no influence on TNFα induced nuclear translocation of NFκB, nor did it have an effect on nuclear mobility of RelA/p65. In an effort to identify a gene expression profile that could be used to identify a SIRT6 readout we conducted genome-wide expression studies. We observed that overexpression of SIRT6 had little influence on NFκB-dependent genes, but overexpression of the catalytically inactive mutant affected gene expression in developmental pathways.     

Seven sirtuins for seven deadly diseases ofaging

Sirtuins are a class of NAD⁺-dependent deacetylases having beneficial health effects. This extensive review describes the numerous intracellular actions of the seven mammalian sirtuins, their protein targets, intracellular localization, the pathways they modulate, and their role in common diseases of aging. Selective pharmacological targeting of sirtuins is of current interest in helping to alleviate global disease burden. Since all sirtuins are activated by NAD⁺, strategies that boost NAD⁺ in cells are of interest. While most is known about SIRT1, the functions of the six other sirtuins are now emerging. Best known is the involvement of sirtuins in helping cells adapt energy output to match energy requirements. SIRT1 and some of the other sirtuins enhance fat metabolism and modulate mitochondrial respiration to optimize energy harvesting. The AMP kinase/SIRT1–PGC-1α–PPAR axis and mitochondrial sirtuins appear pivotal to maintaining mitochondrial function. Downregulation with aging explains much of the pathophysiology that accumulates with aging. Posttranslational modifications of sirtuins and their substrates affect specificity. Although SIRT1 activation seems not to affect life span, activation of some of the other sirtuins might. Since sirtuins are crucial to pathways that counter the decline in health that accompanies aging, pharmacological agents that boost sirtuin activity have clinical potential in treatment of diabetes, cardiovascular disease, dementia, osteoporosis, arthritis, and other conditions. In cancer, however, SIRT1 inhibitors could have therapeutic value. Nutraceuticals such as resveratrol have a multiplicity of actions besides sirtuin activation. Their net health benefit and relative safety may have originated from the ability of animals to survive environmental changes by utilizing these stress resistance chemicals in the diet during evolution. Each sirtuin forms a key hub to the intracellular pathways affected.     

The Sirtuin family: therapeutic targets to treat diseases of aging

Sirtuins have emerged as therapeutic targets to treat age-related diseases. There are seven human Sirtuins (SIRT1-7) that display diversity in cellular localization and function. Growing evidence suggests that small-molecule activators of SIRT1 may counteract age-related afflictions such as type 2 diabetes. Alternatively, inhibitors of SIRT2 may be useful in the treatment of neurodegenerative diseases such as Parkinson's disease. Recent discoveries of small-molecule and protein modulators of Sirtuin deacetylation activity have provided enormous insight into the biological and molecular functions of Sirtuins and have validated their potential as therapeutics.     

Brain Activation of SIRT1: Role in Neuropathology

Sirtuins (SIRTs) are a family of regulatory proteins of genetic information with a high degree of conservation among species. The SIRTs are heavily involved in several physiological functions including control of gene expression, metabolism, and aging. SIRT1 has been the most studied sirtuin and plays important role in the prevention and progression of neurodegenerative diseases acting in different pathways of proteins involved in brain function. SIRT1 activation regulates important genes that also exert neuroprotective actions such as p53, nuclear factor kappa B, peroxisome proliferator-activated receptor-gamma (PPARγ), PPARγ coactivator-1α, liver X receptor, and forkhead box O. It is well established in literature that growing population aging, oxidative stress, inflammation, and genetic factors are important conditions to development of neurodegenerative disorders. However, the exact pathophysiological mechanisms leading to these diseases remain obscure. The sirtuins show strong potential to become valuable predictive and prognostic markers for diseases and as therapeutic targets for the treatment of a variety of neurodegenerative disorders. In this context, the aim of the current review is to present an actual view of the potential role of SIRT1 in modulating the interaction between target genes and neurodegenerative diseases on the brain.     

Food for thought: linking caloric intake to behavior via sirtuin activity

Sirtuins are thought to form crucial links between energy levels and cellular metabolism. Libert et al. now provide evidence that SIRT1 activity in the brain modifies mammalian emotional behavior via monoamine signaling and that changes in this pathway might contribute to human affective disorders.     

The flavoring agent dihydrocoumarin reverses epigenetic silencing and inhibits sirtuin deacetylases

Sirtuins are a family of phylogenetically conserved nicotinamide adenine dinucleotide-dependent deacetylases that have a firmly established role in aging. Using a simple Saccharomyces cerevisiae yeast heterochromatic derepression assay, we tested a number of environmental chemicals to address the possibility that humans are exposed to sirtuin inhibitors. Here we show that dihydrocoumarin (DHC), a compound found in Melilotus officinalis (sweet clover) that is commonly added to food and cosmetics, disrupted heterochromatic silencing and inhibited yeast Sir2p as well as human SIRT1 deacetylase activity. DHC exposure in the human TK6 lymphoblastoid cell line also caused concentration-dependent increases in p53 acetylation and cytotoxicity. Flow cytometric analysis to detect annexin V binding to phosphatidylserine demonstrated that DHC increased apoptosis more than 3-fold over controls. Thus, DHC inhibits both yeast Sir2p and human SIRT1 deacetylases and increases p53 acetylation and apoptosis, a phenotype associated with senescence and aging. These findings demonstrate that humans are potentially exposed to epigenetic toxicants that inhibit sirtuin deacetylases.     

SIRT6 deficiency causes ovarian hypoplasia by affecting Plod1-related collagen formation

SIRT6 is a key member of the mammalian sirtuin family of conserved nicotinamide adenine dinucleotide (NAD⁺)-dependent deacetylases. Previous studies have shown that SIRT6 can regulate metabolism, DNA damage repair and aging. Ovarian aging process usually share similar mechanisms with general aging, which is characterized by decreases in both numbers of ovarian follicles and the quality of oocytes. It is reported that the expression level of SIRT6 was significantly decreased in the ovaries of aged mice, and the level of SIRT6 was positively correlated with ovarian reserve, indicating that SIRT6 may be potential markers of ovarian aging. However, its biological roles in follicular development are still unclear. Here, we explored the effect of SIRT6 on follicular development and found that ovarian development was interrupted in SIRT6 knockout (KO) mice, leading to disruptions of puberty and the estrus cycle, significant decreases in numbers of secondary and antral follicles, and decreased collagen in the ovarian stroma. Plod1, a lysyl hydroxylase that is vital for collagen crosslinking and deposition, was decreased at both the mRNA and protein levels in SIRT6-deficient ovaries and granulosa cells (GCs). Additionally, we found abnormal estrogen levels in both SIRT6 KO mice and SIRT6 KD GCs, accompanied by decreases in the levels of the estrogen biosynthesis genes Cyp11a1, Cyp19a1, Mgarp, and increases in the levels of TNF-α and NF-κB. These results confirmed the effect of SIRT6 on follicular development and revealed a possible molecular mechanism for SIRT6 involvement in follicular development via effects on estrogen biosynthesis and collagen formation.     

SIRT6在缺血再灌注损伤中的作用机制及研究进展

缺血性损伤后恢复血液供应会导致缺血再灌注（ischemia reperfusion, IR）损伤,这会导致组织损伤进一步加剧。IR损伤伴随着一系列机制,包括谷氨酸兴奋性毒性、钙超载、氧化应激、炎症和细胞凋亡,最终导致细胞死亡。IR损伤过程均由Sirtuins家族调控,在Sirtuins家族中,特异性定位于细胞核中的SIRT6可以促进对DNA损伤和氧化应激的抵抗,抑制基因组的不稳定性,在代谢稳态中发挥作用,同时SIRT6在人重要脏器中处于高度表达状态。但SIRT6在IR损伤中研究较少,结合国内外最新的研究进展,对SIRT6在IR损伤中的作用进行了回顾性的总结和分析,希望对国内外学者对于SIRT6在IR损伤中的研究提供一些参考依据。     

Function of SIRT1 in physiology

Sirtuins were originally defined as a family of oxidized nicotinamide adenine nucleotide (NAD⁺)-dependent enzymes that deacetylate lysine residues on various proteins. The sirtuins are remarkably conserved throughout evolution from archae to eukaryotes. They were named after their homology to the Saccharomyces cerevisiae gene silent information regulator 2 (Sir2). The mammalian sirtuins, SIRT1-7, are implicated in a variety of cellular functions ranging from gene silencing, control of the cell cycle and apoptosis, and energy homeostasis. As SIRT1 is a nuclear protein and is the mammalian homolog most highly related to Sir2, it has been the focus of a large number of recent studies. Here we review some of the current data related to SIRT1 and discuss its mode of action and biological role in cellular and organismal models. Published in Russian in Biokhimiya, 2009, Vol. 74, No. 7, pp. 869–876.