线粒体代谢、去乙酰化酶(SIRTUINS)与衰老

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Mitochondrial Metabolism, Sirtuins, and Aging

Abstract

摘要

The sirtuins are a family of proteins that act predominantly as nicotinamide adenine dinucleotide (NAD)-dependent deacetylases. In mammals seven sirtuin family members exist, including three members, Sirt3, Sirt4, and Sirt5, that localize exclusively within the mitochondria. Although originally linked to life-span regulation in simple organisms, this family of proteins appears to have various and diverse functions in higher organisms. One particular property that is reviewed here is the regulation of mitochondrial number, turnover, and activity by various mitochondrial and nonmitochondrial sirtuins. An emerging consensus from these recent studies is that sirtuins may act as metabolic sensors, using intracellular metabolites such as NAD and short-chain carbon fragments such as acetyl coenzyme A to modulate mitochondrial function to match nutrient supply.

去乙酰化酶是一类蛋白质,主要作用是烟酰胺腺嘌呤二核苷酸依赖的去乙酰化酶。在哺乳动物中存在7个 sirtuin 家族成员,包括3个成员,Sirt3,Sirt4和 Sirt5,他们专门定位于线粒体内。虽然最初与简单生物的寿命调控有关,但这个蛋白家族似乎在高等生物中具有多种多样的功能。这里综述的一个特性是调节各种线粒体和非线粒体去乙酰化酶的线粒体数量、周转率和活性。最近这些研究中出现的一个共识是去乙酰化酶可以作为代谢传感器,使用胞内代谢物如 NAD 和短链碳片段如乙酰辅酶A 来调节线粒体功能以匹配营养供应。Previous Section 上一节Next Section 下一节

THE SIRTUIN FAMILY OF PROTEINS

蛋白质 SIRTUIN 家族

Originally characterized in yeast as regulators of life span (Kaeberlein et al. 1999), the sirtuins are an evolutionarily conserved family of proteins that appear to exert a wide range of biological functions (Finkel et al. 2009). In mammals seven sirtuin family members exist. Sirt1 appears to be the closest mammalian homolog to yeast Sir2, the first member of the sirtuins linked to aging. Because of this homology, initial studies of mammalian sirtuins focused predominantly on the biology of Sirt1. Insights from these studies have implicated Sirt1 in the regulation of a variety of metabolic phenotypes including insulin secretion (Moynihan et al. 2005Bordone et al. 2006), lipid mobilization from adipocytes (Picard et al. 2004), and regulation of glucose tolerance (Rodgers et al. 2005). With that said, the role of sirtuins in mammalian aging remains an open question, and even some of the earlier work in lower organisms involving Sir2 and life span has recently been questioned (Burnett et al. 2011).

Sirtuins 最初在酵母中被定性为寿命调节因子(Kaeberlein 等人,1999年) ,sirtuins 是一个进化上保守的蛋白家族,似乎具有广泛的生物学功能(Finkel 等人,2009年)。在哺乳动物中有七个去乙酰化酶家族成员。Sirt1似乎是与酵母 Sir2最接近的哺乳动物同源基因,sirtuins 是与衰老有关的第一种 sirtuins。由于这种同源性,哺乳动物去乙酰化酶的初步研究主要集中在 Sirt1的生物学上。从这些研究中得出的结论认为,Sirt1参与了多种代谢表型的调节,包括胰岛素分泌(Moynihan 等人,2005年; Bordone 等人,2006年) ,脂肪细胞的脂肪动员(Picard 等人,2004年) ,以及葡萄糖耐量的调节(Rodgers 等人,2005年)。尽管如此,去乙酰化酶在哺乳动物衰老中的作用仍然是一个悬而未决的问题,甚至早期在低等生物中涉及去乙酰化酶2和寿命的一些工作最近也受到了质疑(Burnett 等人,2011年)。

Although there has been considerable attention directed toward Sirt1 biology, there is also a growing interest in understanding the function of the related family members. It is clear that each of the mammalian sirtuins has a distinct subcellular localization. Sirt1, Sirt6, and Sirt7 are nuclear proteins, although a fraction of Sirt1 can be found in the cytosol. Sirt2, on the other hand, is predominantly cytosolic, although again it can be found in the nucleus in certain situations (North and Verdin 2007). Finally, three sirtuins—Sirt3, Sirt4, and Sirt5—appear to be found exclusively in the mitochondria. From a human genetics point of view, the strongest association between aging and sirtuins is the association between polymorphisms in the mitochondrial Sirt3 and longevity (Rose et al. 2003).

尽管 Sirt1生物学已经引起了相当大的关注,但是对于理解相关家族成员的功能也有了越来越大的兴趣。很明显,每一种哺乳动物去乙酰化酶都有一个不同的亚细胞定位。Sirt1、 Sirt6和 Sirt7是核蛋白,虽然 Sirt1的一小部分可以在胞浆中找到。另一方面,Sirt2主要是细胞溶质,尽管在某些情况下它也可以在细胞核中找到(North 和 Verdin 2007)。最后,在线粒体中发现了 sirt3、 Sirt4和 sirt5三种去乙酰化酶。从人类遗传学的角度来看,衰老和去乙酰化酶之间最强的联系是线粒体 Sirt3基因多态性和长寿之间的联系(Rose 等人,2003年)。

The ability of sirtuins to influence metabolism and potentially life span is believed to revolve around the ability of sirtuin family members to function as protein deacetylases. In addition to this enzymatic function, Sirt4 can further act to ADP ribosylate target proteins. Unlike other protein deacetylases, sirtuins require nicotinamide adenine dinucleotide (NAD) as a cofactor in the deacetylation reaction (Imai et al. 2000). The link among NAD, NADH, and sirtuin activity has led many to believe that this family of proteins acts in some fashion as a sensor of energetic status. This may particularly be true in the mitochondria, where levels of NAD and NADH are high and where a disproportionate fraction of proteins appear to be acetylated (Kim et al. 2006).

Sirtuin 影响新陈代谢和潜在寿命的能力被认为是围绕 sirtuin 家族成员作为蛋白质去乙酰化酶的功能。除此之外,Sirt4还可以进一步作用于 ADP 核糖基化靶蛋白。与其他蛋白质去乙酰化酶不同,去乙酰化酶需要烟酰胺腺嘌呤二核苷酸(NAD)作为去乙酰化反应的辅助因子。NAD、 NADH 和 sirtuin 活性之间的联系使许多人相信这个蛋白家族以某种方式作为能量状态的传感器。在线粒体尤其如此,那里 NAD 和 NADH 水平很高,而且不成比例的蛋白质似乎被乙酰化(Kim 等人,2006年)。

In this review we will examine the link between sirtuin function and mitochondrial metabolism. While we highlight the role of those sirtuin family members that are uniquely mitochondrial in their localization, we will also discuss other sirtuins that can influence mitochondrial function, biogenesis, and turnover. Finally, although we discuss each sirtuin individually, there is a growing realization that one sirtuin family member can affect the function of other family members. A recent example is Sirt1-dependent regulation of Sirt6 expression in the control of hepatic metabolism (Kim et al. 2010b).

在这篇综述中,我们将检查 sirtuin 功能和线粒体代谢之间的联系。当我们强调这些 sirtuin 家族成员的作用,是独特的线粒体在其定位,我们也将讨论其他 sirtuin,可以影响线粒体功能,生物发生和周转。最后,虽然我们分别讨论每个去乙酰化酶,但是人们越来越意识到一个去乙酰化酶家庭成员可以影响其他家庭成员的功能。最近的一个例子是 sirt1依赖的 Sirt6表达调控肝脏代谢(Kim 等人,2010b)。

Sirt1 AND MITOCHONDRIAL FUNCTION

Sirt1与线粒体功能

Evidence suggests that mitochondrial biogenesis is regulated at least in part by proliferator-activated receptor coactivator-1α (PGC-1α), a transcriptional coactivator of peroxisome proliferator-activated receptor-γ (PPARγ) as well as other transcription factors (Fernandez-Marcos and Auwerx 2011). It was therefore of considerable interest when it was shown that PGC-1α was in fact a deacetylation target of Sirt1 and that acetylation regulated PGC-1α activity (Nemoto et al. 2005Rodgers et al. 2005). There are at least 13 lysine residues on PGC-1α that appear to be reversibly acetylated (Rodgers et al. 2005). Site-directed mutants that lack all 13 of these sites alter the ability of PGC-1α to regulate gene expression, although it remains unclear if all, or only a subset, of PGC-1α’s acetylation sites are truly regulatory in nature. Sirt1 appears to be the predominant in vitro and in vivo regulator of PGC-1α deacetylation. For instance, in vitro knockdown of Sirt1 in hepatic cells leads to increased PGC-1α acetylation with a corresponding reduction in a set of genes that are the rate-limiting enzymes responsible for hepatic gluconeogenesis (Rodgers et al. 2005). Similarly, both overexpression and knockdown studies support a role for Sirt1 in regulating PGC-1α activity through reversible deacetylation, which in turn has dramatic effects on in vivo hepatic glucose and lipid metabolism (Rodgers and Puigserver 2007Erion et al. 2009). A similar relationship appears to exist in skeletal muscle. In particular, in skeletal muscle, fasting was shown to lead to a Sirt1-dependent deacetylation of PGC-1α, and this deacetylation appeared to be required for PGC-1α-dependent gene expression, including gene products required for effective mitochondrial biogenesis (Gerhart-Hines et al. 2007). Together these studies link Sirt1 and PGC-1α activities in metabolically active tissues such as the liver and skeletal muscle (Fig. 1).


有证据表明,线粒体生物发生至少部分受到增殖物激活受体辅激活因子 -1(pgc-1)、过氧化物酶体增殖物激活受体转录辅激活因子(ppar)以及其他转录因子的调控(Fernandez-Marcos and Auwerx 2011)。因此,当发现 pgc-1实际上是 Sirt1的去乙酰化靶点,而乙酰化调节 pgc-1活性时,引起了相当大的兴趣(Nemoto 等人,2005年; Rodgers 等人,2005年)。Pgc-1上至少有13个赖氨酸残基似乎是可逆乙酰化的(Rodgers 等人,2005年)。缺乏所有13个位点的定点突变改变了 pgc-1调节基因表达的能力,尽管目前尚不清楚 pgc-1乙酰化位点的全部或仅仅一个子集是否真正具有调节性质。Sirt1似乎是 pgc-1脱乙酰化的主要体外和体内调节因子。例如,在体外击倒肝细胞中的 Sirt1导致 pgc-1乙酰化增加,相应的一组基因(肝糖异生的限速酶)减少(Rodgers 等人,2005年)。同样,过度表达和击倒研究都支持 Sirt1通过可逆的去乙酰化来调节 pgc-1的活性,而可逆的去乙酰化又对体内的葡萄糖和脂质代谢产生巨大的影响。类似的关系似乎存在于骨骼肌中。特别是在骨骼肌中,禁食会导致 pgc-1的 sirt1依赖性脱乙酰化,而这种脱乙酰化似乎是 pgc-1依赖性基因表达所必需的,包括有效的线粒体生物合成所需的基因产物(Gerhart-Hines 等人,2007年)。这些研究联系了 Sirt1和 pgc-1活性在代谢活跃的组织,如肝脏和骨骼肌(图1)。

Figure 1. 图1

Regulation of PGC-1α acetylation and activity by Sirt1. The transcriptional coactivator and regulator of mitochondrial biogenesis PGC-1α is, at least in part, regulated by lysine acetylation. In the setting of low nutrient availability, the intracellular and particularly the nuclear levels of NAD are believed to increase and lead to activation of Sirt1 enzymatic activity. This leads to PGC-1α deacetylation, resulting in increased PGC-1α activity and hence, ultimately, in an increase in mitochondrial number.

Sirt1对 pgc-1乙酰化及活性的调控。线粒体生物发生的转录辅激活因子和调控因子 pgc-1至少部分受赖氨酸乙酰化调控。在营养物质利用率低的情况下,细胞内 NAD 水平尤其是细胞核 NAD 水平升高,导致 Sirt1酶活性的激活。这导致 pgc-1脱乙酰化,导致 pgc-1活性增加,因此,最终,线粒体数量增加。

Regulation of PGC-1α acetylation and activity by Sirt1. The transcriptional coactivator and regulator of mitochondrial biogenesis PGC-1α is, at least in part, regulated by lysine acetylation. In the setting of low nutrient availability, the intracellular and particularly the nuclear levels of NAD are believed to increase and lead to activation of Sirt1 enzymatic activity. This leads to PGC-1α deacetylation, resulting in increased PGC-1α activity and hence, ultimately, in an increase in mitochondrial number.

Sirt1对 pgc-1乙酰化及活性的调控。线粒体生物发生的转录辅激活因子和调控因子 pgc-1至少部分受赖氨酸乙酰化调控。在营养物质利用率低的情况下,细胞内 NAD 水平尤其是细胞核 NAD 水平升高,导致 Sirt1酶活性的激活。这导致 pgc-1脱乙酰化,导致 pgc-1活性增加,因此,最终,线粒体数量增加。

The above studies suggest that mitochondrial biogenesis might be regulated by tissue energetic status and that the sirtuins would represent important energy sensors in this homeostatic loop. Indeed, the notion that PGC-1α acetylation and function, and by extension mitochondrial activity, are regulated in a nutrient-dependent fashion by Sirt1 is appealing. Nonetheless, the concept that Sirt1 is in turn responding to nutrient-sensitive changes in basal NAD levels, although often invoked, has until recently had little experimental support. The difficulty in proving the supposition is that measurement and manipulation of NAD levels in various subcellular compartments is experimentally challenging. One recent approach is to carefully examine mice with a deletion of a major NAD-consuming enzyme, poly(ADP-ribose) polymerase-1 (PARP-1). These mice appear to have elevated NAD levels along with increased Sirt1 activity (Bai et al. 2011). Furthermore, consistent with increased NAD levels leading to increased PGC-1α activity, PARP-1−/− mice have increased mitochondrial content (Bai et al. 2011). Another recent report suggested that adiponectin, a secreted adipokine, could regulate intracellular NAD levels (Iwabu et al. 2010). Again, in these studies the addition of adiponectin to cells appeared to increase mitochondrial content through a Sirt1- and PGC-1α-dependent pathway. These results might be particularly important because metabolic diseases are often associated with low adiponectin levels (Hotta et al. 2000), as well as with mitochondrial dysfunction (Petersen et al. 2004), although the link between these two observations was previously unknown. Interestingly, these observations would also seem to support the growing link between genetic variants of Sirt1 and a person’s risk for developing obesity (Peeters et al. 2008Zillikens et al. 2009).

上述研究表明,线粒体的生物发生可能受组织能量状态的调节,而去乙酰化酶可能是这个稳态环路中的重要能量传感器。事实上,pgc-1的乙酰化和功能,以及由此引申出的线粒体活性,是由 Sirt1以一种营养依赖的方式调节的观点是很吸引人的。尽管如此,Sirt1对营养敏感的基础 NAD 水平变化反过来作出反应的概念,尽管经常被提及,但直到最近几乎没有实验支持。难以证明的假设是,测量和操纵各种亚细胞室 NAD 水平是实验性的挑战。最近的一种方法是仔细检查缺失一种主要的 nad- 消耗酶,多聚腺苷二磷酸核糖聚合酶 -1(PARP-1)的小鼠。这些小鼠的 NAD 水平似乎随着 Sirt1活性的增加而升高(Bai 等人,2011年)。此外,与 NAD 水平升高导致 pgc-1活性升高一致,PARP-1-/-小鼠的线粒体含量增加(Bai 等人,2011年)。另一个最近的报告表明,脂联素,一种分泌的脂肪因子,可以调节细胞内 NAD 水平(Iwabu 等人,2010年)。在这些研究中,向细胞中添加脂联素似乎通过 Sirt1和 pgc-1依赖途径增加了线粒体的含量。这些结果可能特别重要,因为代谢性疾病通常与低脂联素水平有关(Hotta 等人,2000年) ,以及与线粒体功能障碍有关(Petersen 等人,2004年) ,尽管这两个观察结果之间的联系以前未知。有趣的是,这些观察结果似乎也支持了 Sirt1基因变异与一个人患肥胖症风险之间日益增长的联系(Peeters 等人2008; Zillikens 等人2009)。

The generation of new mitochondria through a Sirt1/PGC-1α-regulated pathway is complemented by another important connection between Sirt1 and the mitochondria. In particular, it would appear that Sirt1 is an important regulator of removing damaged mitochondria through the process of autophagy (Lee et al. 2008). The field of autophagy represents a rapidly expanding area of study, and a full review of the subject is not possible. Suffice it to say that autophagy is an evolutionarily conserved process present in organisms ranging from yeast to mammals. One of the major intracellular roles of autophagy is the removal of damaged organelles such as mitochondria. Evidence suggests that Sirt1 can stimulate autophagy and that Sirt1−/− tissues appear to accumulate abnormal-appearing mitochondria, consistent with what is seen in autophagy-deficient tissues (Lee et al. 2008). A cytoplasm-restricted mutant of Sirt1 can still stimulate autophagy, suggesting that this activity represents an extranuclear function of the protein (Morselli et al. 2011). The molecular basis for how Sirt1 stimulates autophagy is not clear. There is evidence that key molecules required for autophagy including Atg5 and Atg7 are direct targets of sirtuin-dependent deacetylation (Lee et al. 2008). In addition, the FoxO family of transcription factors, known targets of Sirt1 deacetylation as well as regulators of autophagy, has also been implicated (Hariharan et al. 2010Kume et al. 2010). Nonetheless, although details remain to be elucidated, the notion that Sirt1 can regulate both the creation of new mitochondria as well as the removal of old mitochondria suggests a role for sirtuins in overall mitochondrial flux and in the maintenance of what may be viewed as “youthful” mitochondria in the cell.

通过 Sirt1/PGC-1调节通路产生新的线粒体,同时还有 Sirt1和线粒体之间的另一个重要联系。特别是,似乎 Sirt1是通过自噬过程移除受损线粒体的重要调节因子(Lee 等人,2008年)。自噬领域代表了一个迅速扩展的研究领域,对这个课题进行全面的回顾是不可能的。只要说自噬是一个进化上保守的过程,存在于从酵母到哺乳动物的生物体中就足够了。细胞内自噬的主要作用之一是去除受损的细胞器,如线粒体。有证据表明,Sirt1可以刺激自噬,而且 Sirt1-/-组织似乎积累了异常的线粒体,这与自噬缺陷组织中的现象一致(Lee 等人,2008年)。Sirt1的细胞质限制突变体仍然可以刺激自噬,这表明这种活性代表了蛋白质的核外功能(Morselli 等人,2011年)。Sirt1如何刺激自噬的分子基础尚不清楚。有证据表明,包括 Atg5和 Atg7在内的自噬所需的关键分子是 sirtuin 依赖性脱乙酰化的直接靶标(Lee 等人,2008)。此外,转录因子的 FoxO 家族,Sirt1去乙酰化的已知目标以及自噬的调节因子,也有牵连(Hariharan 等人,2010; Kume 等人,2010)。尽管如此,尽管细节仍有待阐明,但是 Sirt1可以同时调节新线粒体的产生和旧线粒体的去除这一概念表明去乙酰化酶在整个线粒体通量和维持细胞中可能被视为“年轻的”线粒体中发挥了作用。

A final area that we wish to discuss in which the biology of Sirt1 and the mitochondria intersect is the secretion of insulin by the pancreatic β cell. Previous studies have suggested that mitochondrial uncoupling protein-2 (UCP2) was an important negative regulator of insulin secretion by the β cell (Zhang et al. 2001). Indeed, UCP2 appears to be up-regulated in obese animals and UCP2-deficient mice have increased glucose-stimulated insulin release (Zhang et al. 2001). Subsequent studies identified Sirt1 as an important repressor of UCP2 expression in the pancreas (Moynihan et al. 2005Bordone et al. 2006). It is believed that starvation may induce UCP2 expression in the β cell via a reduction in tissue NAD levels (Bordone et al. 2006). There is also evidence that a recently described extracellular, circulating form of the enzyme nicotinamide phosphoribosyltransferase (Nampt) may be an important in vivo mediator of insulin secretion (Revollo et al. 2007). The Nampt enzyme is a key enzyme in NAD biosynthesis and is responsible for the conversion of nicotinamide to nicotinamide mononucleotide, a metabolite that can in turn be converted directly to NAD. Again, these observations highlight the potential function of Sirt1 in linking metabolites such as NAD to the maintenance of overall metabolic homeostasis.

我们希望讨论的最后一个领域是 Sirt1和线粒体的生物学相交的胰腺细胞的胰岛素分泌。以往的研究表明,线粒体解偶联蛋白 -2(UCP2)是一个重要的负调节胰岛素分泌的细胞(张等。2001年)。事实上,UCP2在肥胖动物中似乎是上调的,而 UCP2缺陷的小鼠增加了葡萄糖刺激的胰岛素释放(Zhang 等人,2001年)。随后的研究确定 Sirt1是胰腺 UCP2表达的重要抑制因子(Moynihan 等人,2005; Bordone 等人,2006)。据认为,饥饿可能通过降低组织 NAD 水平而诱导细胞内 UCP2的表达(Bordone 等人,2006年)。还有证据表明,最近描述的一种细胞外循环形式的烟酰胺磷酸核糖转移酶(Nampt)可能是体内胰岛素分泌的重要介质(Revollo 等人,2007年)。Nampt 酶是 NAD 生物合成的关键酶,负责将烟酰胺转化为烟酰胺单核苷酸,这种代谢物可以直接转化为 NAD。再次,这些观察突出了 Sirt1在连接代谢物如 NAD 和维持整体代谢稳态方面的潜在功能。Previous Section 上一节Next Section 下一节

Sirt3 AND MITOCHONDRIAL FUNCTION

Sirt3与线粒体功能

The studies we have reviewed indicating that the predominantly nuclear Sirt1 was an important regulator of mitochondrial function spurred interest in the biology of those sirtuin family members that directly localize to this organelle. Comparative analysis of total liver mitochondrial protein acetylation following distinct genetic knockout of each of the three mitochondrial-enriched sirtuins—Sirt3, Sirt4, and Sirt5—showed that Sirt3 is the major mitochondrial deacetylase (Lombard et al. 2007). At the same time, the dynamic flux in mitochondrial protein acetylation in response to changes in caloric load as illustrated by feeding and fasting (Kim et al. 2006), caloric restriction (Schwer et al. 2009), and caloric excess (Hirschey et al. 2011Kendrick et al. 2011) suggest that, as is the case with Sirt1, Sirt3 may possess a nutrient-sensing regulatory role governing mitochondrial protein function.


我们回顾的研究表明,主要核的 Sirt1是线粒体功能的重要调节因子,激发了对那些直接定位于这一细胞器的 sirtuin 家族成员的生物学兴趣。3种线粒体丰富的 sirtuins ー Sirt3、 Sirt4和 sirt5ー被不同的基因剔除后,肝脏线粒体总蛋白乙酰化的比较分析表明,Sirt3是主要的线粒体去乙酰化酶(Lombard et al. 2007)。同时,线粒体蛋白质乙酰化的动态通量响应热量负荷的变化,如喂食和禁食(Kim 等人,2006年) ,热量限制(Schwer 等人,2009年) ,和热量过剩(Hirschey 等人,2011年; Kendrick 等人,2011年)表明,Sirt3可能具有营养敏感的调节作用控制线粒体蛋白质功能。

Over the last few years, many laboratories have identified and functionally characterized mitochondrial targets of Sirt3 deacetylation and shown that Sirt3 does indeed modulate numerous mitochondrial pathways via protein lysine-residue deacetylation. As mitochondria are most readily identified as the “powerhouse” of the cell, we initially review the role of Sirt3 in mitochondrial bioenergetics. Sirt3 deacetylates and activates multiple steps in substrate catabolism, starting with the oxidation of fatty acids. As an example, Sirt3 has been shown under nutrient-restricted conditions to deacetylate and hence activate long-chain acyl coenzyme A (acyl-CoA) dehydrogenase to increase fatty acid β-oxidation (Hirschey et al. 2010Hallows et al. 2011). Additional substrate catabolic pathway targets include glutamate dehydrogenase, which facilitates the oxidative deamination of glutamate to α-ketoglutarate, and the citric acid cycle enzyme isocitrate dehydrogenase 2 (IDH2) (Lombard et al. 2007Schlicker et al. 2008). In keeping with its role as the final common denominator in mitochondrial energy production, numerous proteins in the electron transport chain have been shown to be directly deacetylated by Sirt3. Sirt3 activation has been shown to increase oxidative phosphorylation (Ahn et al. 2008Bao et al. 2010aCimen et al. 2010) and to deacetylate and activate enzymes in complexes I and II of the electron transport chain (Ahn et al. 2008Cimen et al. 2010Finley et al. 2011Kendrick et al. 2011), with additional deacetylation of proteins in complex V (Schlicker et al. 2008Bao et al. 2010b).

在过去的几年中,许多实验室已经鉴定和功能特征的线粒体目标 Sirt3脱乙酰基,并表明 Sirt3确实调节许多线粒体途径通过蛋白赖氨酸残基脱乙酰基。由于线粒体最容易被确定为细胞的“发电站” ,我们首先回顾了 Sirt3在线粒体生物能学中的作用。从脂肪酸的氧化开始,Sirt3使底物分解代谢中的多个步骤脱乙酰化并激活。例如,Sirt3已经被证明在营养限制条件下去乙酰化,因此激活长链酰基辅酶 a (酰基辅酶 a)脱氢酶增加脂肪酸氧化(Hirschey 等人,2010; Hallows 等人,2011)。其他底物分解代谢途径靶点包括促进谷氨酸氧化脱氨基的谷氨酸脱氢酶和三羧酸循环异柠檬酸脱氢酶2(IDH2)(Lombard et al. 2007; Schlicker et al. 2008)。为了保持其作为线粒体能量产生的最终公分母的角色,电子传递链中的许多蛋白质已被证明可以被 Sirt3直接去乙酰化。3的激活已被证明可以增加复合物 v 中的氧化磷酸化,并在复合物 i 和 II 中脱乙酰化和激活酶(Ahn et al. 2008; Cimen et al. 2010; Finley et al. 2011; Kendrick et al. 2011) ,以及复合物 v 中其他蛋白质的脱乙酰化作用(Schlicker et al. 2008; Bao et al. 2010b)。

An additional aspect of mitochondrial bioenergetics operational under caloric restriction or fasting conditions is the conversion of acetate to acetyl-CoA, a necessary step required for energy production in extrahepatic tissues and for the generation of ketones in the liver. Again, Sirt3 appears to have an important regulatory role in these pathways, as both acetyl-CoA synthetase 2 (Hallows et al. 2006Schwer et al. 2006) and 3-hydroxy-3-methylglutaryl-CoA synthase 2 (Shimazu et al. 2010) are targets of Sirt3 deacetylation. A final metabolic pathway that is modulated by Sirt3 is the urea cycle, responsible for the detoxification of ammonia during amino acid metabolism. Here, too, Sirt3 deacetylates and activates a key enzyme, ornithine transcarbamoylase (Hallows et al. 2011).

线粒体生物能学在限制热量或禁食条件下运作的另一个方面是将醋酸盐转化为乙酰辅酶 a,这是肝外组织产生能量和在肝脏中生成酮所需的一个必要步骤。再次,Sirt3似乎在这些途径中有一个重要的调节作用,因为乙酰辅酶 a 合成酶2(Hallows et al. 2006; Schwer et al. 2006)和羟甲基戊二酸单酰辅酶A 合酶2(Shimazu et al. 2010)都是 Sirt3去乙酰化的目标。最后一个受 Sirt3调控的代谢途径是尿素循环,负责氨基酸代谢过程中的氨解毒。在这里,同样,Sirt3去乙酰化并激活一种关键酶,鸟氨酸转氨酶(Hallows et al. 2011)。

Sirt3 is also emerging as a regulatory protein in the modulation of additional mitochondrial programs, including the deacetylation and inhibition of the mitochondrial ribosomal protein L10 (MRPL10) (Yang et al. 2009). This results in an NAD-dependent inhibition of mitochondrial protein synthesis, which might function as an energy-sparing response under nutrient-restricted conditions. Another mitochondrial protein that is inhibited by Sirt3 deacetylation is the mitochondrial matrix peptidyl–prolyl isomerase cyclophilin D (Ppif) (Hafner et al. 2010Shulga and Pastorino 2010Shulga et al. 2010). Our understanding of the function of cyclophilin D has expanded in recent years to include a role not only in increasing susceptibility to mitochondrial permeability transition (Baines et al. 2005Nakagawa et al. 2005), but also in regulating mitochondrial calcium efflux with the concordant regulation of Ca2+-dependent mitochondrial enzyme activities (Elrod et al. 2010). An additional, albeit indirect, effect of Sirt3-dependent inactivation of cyclophilin D is the dissociation of hexokinase II from the mitochondria, which plays a role in the promotion of oxidative phosphorylation by Sirt3 (Shulga et al. 2010).

在调节额外的线粒体程序中,Sirt3也逐渐成为一种调节蛋白,包括去乙酰化和抑制线粒体核糖体蛋白质 L10(MRPL10)(Yang et al. 2009)。这导致依赖于 nad- 的线粒体蛋白质合成的抑制,这可能作为一个能量节省反应在营养限制条件下。另一种被 Sirt3去乙酰化抑制的线粒体蛋白是线粒体基质肽基脯氨酰异构酶亲环素 d (Ppif)(Hafner 等人2010; Shulga and Pastorino 2010; Shulga 等人2010)。近年来,我们对亲环素 d 功能的理解有所扩展,不仅包括增加线粒体通透性转换的易感性(Baines 等人,2005年; Nakagawa 等人,2005年) ,而且还包括调节线粒体钙外流与 Ca2 + 依赖性线粒体酶活性的一致性调节(Elrod 等人,2010年)。依赖于 Sirt3的亲环素 d 失活的另一个间接影响是己糖激酶 II 与线粒体的解离,它在 Sirt3促进氧化磷酸化中起作用(Shulga et al. 2010)。

The mitochondrial sirtuins also appear to play an important role in the control of reactive oxygen species. This regulatory role may be particularly relevant to modulating the development of age-associated degenerative conditions. At the direct substrate level, the reactive oxygen species scavenging enzyme MnSOD is activated by Sirt3, and numerous lysine residues have been implicated in mediating this induction of enzyme activity (Qiu et al. 2010Tao et al. 2010Chen et al. 2011). Via a more indirect mechanism, the caloric restriction-associated activation of IDH2 has been shown to increase NADH, which is proposed to facilitate the increase in reduced glutathione levels found in association with Sirt3-mediated activation of IDH2 (Someya et al. 2010). The emerging role of Sirt3 in modulating various pathways in metabolism and stress modulatory programs is shown in Figure 2.

线粒体去乙酰化酶似乎也在控制活性氧类中起着重要作用。这种调节作用可能特别相关的调节发展年龄相关的退行性条件。在直接底物水平上,活性氧类清除酶 MnSOD 被 Sirt3激活,许多赖氨酸残基参与了这种酶活性的诱导(邱等人,2010; 陶等人,2010; 陈等人,2011)。通过一个更间接的机制,卡路里限制相关的活化 IDH2已被证明增加 NADH,这被提议促进与 sirt3介导的活化 IDH2相关的谷胱甘肽水平的增加(Someya 等人2010)。Sirt3在调节代谢和应激调节程序的各种途径中的新兴作用如图2所示。

Figure 2. 图2

A role for Sirt3 in metabolic adaptation and stress defense in the setting of low nutrients. The sirtuin family member Sirt3 is the predominant mitochondrial deacetylase. Multiple targets of Sirt3 have been described. These targets involve enzymes linked to substrate utilization as well as core components of the electron transport chain. In addition, Sirt3 has been linked to a wide array of other stress-related programs including the maintenance of redox homeostasis. See text for additional details.

Sirt3在低营养条件下代谢适应和逆境防御中的作用。Sirtuin 家族成员 Sirt3是主要的线粒体脱乙酰酶。描述了 Sirt3的多个靶点。这些目标包括与底物利用有关的酶以及电子传递链的核心成分。此外,Sirt3已经与一系列其他与压力相关的项目联系在一起,包括维持氧化还原体内平衡。详细信息请参阅文本。

Finally, a recently defined target of Sirt3, namely, mitochondrial aldehyde dehydrogenase 2 (ALDH2), has uncovered an additional biological role for the acetylation of lysine residues (Lu et al. 2011). In contrast to the prior targets of Sirt3 discussed above, the change in acetylation status of ALDH2 did not alter the activity of this dehydrogenase. However, because the reactive metabolite of the analgesic agent acetaminophen disrupts proteins by binding to lysine residues, the role of Sirt3 in acetaminophen metabolite binding to ALDH2 was explored (Lu et al. 2011). This analysis confirmed that lysine-residue acetylation can indeed function as an allosteric inhibitor of xenobiotic-reactive metabolite binding to mitochondrial proteins by exposing lysine residues to other posttranslational modifiers, and that in this instance Sirt3 activation has detrimental consequences (Silberman and Mostoslavsky 2011). These results might help explain the clinical observation that acetaminophen liver injury is exacerbated by fasting (Lu et al. 2011)

最后,最近定义的 Sirt3靶点,即线粒体 aldehyde 2(ALDH2) ,揭示了赖氨酸残基乙酰化的额外生物学作用(Lu 等人,2011年)。与前面讨论的 Sirt3的目标相比,ALDH2乙酰化状态的改变并没有改变该脱氢酶的活性。然而,由于止痛剂 acetaminophen 的活性代谢产物通过与赖氨酸残基结合破坏蛋白质,Sirt3在 acetaminophen 代谢产物与 ALDH2结合中的作用被探索(Lu 等人,2011)。这项分析证实,赖氨酸残基乙酰化确实可以作为异生物活性代谢物与线粒体蛋白质结合的变构抑制剂,将赖氨酸残基暴露给其他翻译后修饰物,在这种情况下,Sirt3的激活具有有害后果(Silberman 和 Mostoslavsky,2011年)。这些结果可能有助于解释临床观察,对乙酰氨基酚肝损伤加重的禁食(卢等人2011年)

The investigations of the targets of Sirt3 collectively show that in response to caloric restriction or fasting, Sirt3 deacetylates a vast array of mitochondrial proteins, with a resulting panoply of effects including activation, inhibition, and allosteric modification of protein functioning. The physiological impact of these effects is beginning to be explored in a range of age-related or nutrient-sensitive diseases. On balance, recent observations suggest that Sirt3 activation is generally ameliorative with respect to the development of age-associated diseases such as cardiac dysfunction (Sundaresan et al. 2009Hafner et al. 2010), hearing loss (Someya et al. 2010), metabolic syndromes and diabetes (Hirschey et al. 2011Jing et al. 2011Kendrick et al. 2011), and cancer (Kim et al. 2010a).

对 Sirt3基因靶点的研究表明,Sirt3基因在应对限制热量摄入或禁食时,能够去乙酰化大量的线粒体蛋白质,从而产生包括激活、抑制和蛋白质功能变构修饰在内的一系列效应。在一系列与年龄有关或对营养敏感的疾病中,这些效应的生理影响正在开始探索。总的来说,最近的观察表明,Sirt3的激活通常可以改善年龄相关疾病的发展,如心脏功能障碍(Sundaresan 等人,2009年; Hafner 等人,2010年) ,听力损失(Someya 等人,2010年) ,代谢综合征和糖尿病(Hirschey 等人,2011年; Jing 等人,Kendrick 等人,2011年) ,以及癌症(Kim 等人,2010a 年)。Previous Section 上一节Next Section 下一节

Sirt5 AND MITOCHONDRIAL FUNCTION

Sirt5与线粒体功能

The identification of targets of Sirt5 in the mitochondria has been more limited, although the urea cycle enzyme carbamoyl phosphate synthetase 1 (CPS-1) has been identified as a Sirt5 deacetylation target (Nakagawa et al. 2009). The activation of CPS-1 catalyzes ammonia to urea and would be expected to have ameliorative effects via the elimination of oxidative stress-promoting ammonium.

虽然已经证实氨甲酰磷酸合成酶1(CPS-1)是 Sirt5去乙酰化的靶标,但是在线粒体中对 Sirt5靶标的鉴定还是很有限的。CPS-1催化氨转化为尿素,可能通过消除氧化应激促进铵的作用而起到改善作用。

Recent work shows that in addition to acetyl groups, additional short-chain carbon fragments covalently attached to specific amino acids may be targets of Sirt5. These include the removal of succinyl and malonyl groups from lysine residues (Du et al. 2011Peng et al. 2011). Succinyl-CoA and malonyl-CoA are metabolic intermediates of the tricarboxylic acid (TCA) cycle (Peng et al. 2011Zhang et al. 2011). To date, very few specific protein targets of Sirt5-mediated desuccinylation or demalonylation have been identified. However, the levels of hepatic protein succinylation and malonylation are highly enriched in Sirt5 knockout mice (Du et al. 2011Peng et al. 2011). Although these novel posttranslational modifications attributed to Sirt5 need to be fleshed out, they are compatible with the concept that the sirtuin proteins target lysine-residue modifications to alter metabolic functioning in response to changes in levels of various intracellular nutrients.

最近的研究表明,除了乙酰基外,另外一些与特定氨基酸共价结合的短链碳片段可能是 Sirt5的靶标。其中包括从赖氨酸残基中去除琥珀酰和丙二酰基(Du et al. 2011; Peng et al. 2011)。琥珀酰辅酶 a 和丙二酰辅酶 a 是三元羧酸(TCA)循环的代谢中间产物(Peng et al. 2011; Zhang et al. 2011)。到目前为止,很少有蛋白质目标的 sirt5介导脱 uccinylation 或 dealonylation 已经被确定。然而,在 Sirt5基因敲除小鼠中,肝蛋白琥珀酸化和丙二酰化的水平高度富集(杜等,2011; 彭等,2011)。虽然这些新颖的翻译后修饰属于 Sirt5需要充实,他们符合的概念,sirtuin 蛋白的目标是赖氨酸残基修饰改变代谢功能,以响应各种细胞内营养素水平的变化。Previous Section 上一节Next Section 下一节

CONCLUSIONS

结论

Although originally described in yeast, the mammalian sirtuins represent an intriguing family of proteins that appear to function as sensors and regulators of metabolic status. Included among this family’s diverse function is the coordinated control of mitochondrial activity. This regulation includes the creation and targeted destruction of mitochondria by Sirt1, as well as the regulation of substrate utilization and oxidative phosphorylation by Sirt3. Table 1 shows the metabolic substrates of Sirt1, −3, and −5 discussed in this review and summarizes the metabolic effects of these deacetylation reactions.

虽然最初在酵母中被描述,哺乳动物的去乙酰化酶代表了一个有趣的蛋白家族,似乎作为传感器和代谢状态调节器的功能。这个家族的多种功能包括线粒体活动的协调控制。这个调节包括 Sirt1对线粒体的创造和定向破坏,以及 Sirt3对底物利用和氧化磷酸化的调节。表1显示了代谢底物的 Sirt1,-3,-5在这篇综述中讨论,并总结了这些脱乙酰化反应的代谢效应。

Table 1. 表一

List of described sirtuin substrates, their subcellular localization, and the effect of deacetylation on protein function

所描述的 sirtuin 底物清单,它们的亚细胞定位,以及脱乙酰化对蛋白质功能的影响

In addition to these biochemical properties, there are tantalizing clues that sirtuins may play an important physiological role in overall metabolic homeostasis and perhaps in modulating age-related metabolic pathologies. Finally, although the predominant function of the sirtuin family revolves around NAD-dependent lysine deacetylation, other less-characterized enzymatic activities including ADP ribosylation, as well as other lysine modifications (e.g., demalonylation and desuccinylation), are just beginning to be explored. Although considerable gaps exist in our understanding, further dissection of sirtuin biology promises to provide important insight into how metabolic supply is coupled to mitochondrial activity.

除了这些生化特性,还有一些诱人的线索表明去乙酰化酶可能在整个代谢稳态中发挥重要的生理作用,也许还可以调节与年龄有关的代谢病理。最后,虽然 sirtuin 家族的主要功能是依赖于 nad- 的赖氨酸脱乙酰化,但其他一些不太特征化的酶活性,包括 ADP 核糖基化,以及其他赖氨酸修饰(如脱孤蛋白修饰和脱琥珀酸修饰) ,才刚刚开始探索。虽然在我们的理解中存在着相当大的差距,但是对去乙酰化酶生物学的进一步解剖有望提供重要的见解,了解代谢供应是如何与线粒体活性相结合的。

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