Sirtuin Regulation in CR - Part 1

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The below is part 1 of a paper that is pdf-availed. Sirtuin Regulation in Calorie Restriction.Qiu X, Brown KV, Moran Y, Chen D.Biochim Biophys Acta. 2010 Aug;1804(8):1576-83. Epub 2009 Sep 24.PMID: 19782772 The beneficial effects of calorie restriction diet in extending lifespan and preventing diseases have long been recognized. Recent genetic and molecular studies in model organisms began to uncover the molecular regulation of calorie restriction response, with the gene SIR2 playing an essential role. This article summarizes the latest development on how mammalian SIR2 homologs coordinately regulate the calorie restriction response. Introduction Metabolic syndrome, cancer, neurodegenerative diseases, and

immunedysfunction are the major threats to human health. These seemingly unrelateddiseases share one common theme: their incidence increases with age. It isintriguing that interventions slowing the aging process may have the potential toprevent a wide range of diseases associated with aging. This notion is supportedby the beneficial effects of calorie restriction (CR) diet. 30-40% reduction of foodintake increases lifespan up to 50% in laboratory rodents. Indeed, this dietaryintervention of aging ameliorates many late-onset diseases [1]. An important question is whether the multifaceted effects of CR are mediated byregulated biological pathways that are amenable to studies and can beharnessed for disease prevention. Recent genetic and molecular studies inmodel organisms began to uncover the molecular regulation of CR response,with the gene SIR2 playing an essential role. SIR2 is highly conserved

from yeastto mammals [2]. SIR2-like genes are collectively known as ‘sirtuins' (reviewed in[3] and several articles in this issue of BBA). In this review, we will discuss theregulation of sirtuins during CR, the effects of such regulation on aging anddiseases of aging, and the implications on drug development. Sirtuins as mediators of calorie restriction response The lifespan of the budding yeast Saccharomyces cerevisiae can be measuredby the number of daughter cells that one mother cell can produce (replicativelifespan) [4]. SIR2 promotes yeast replicative lifespan by inhibiting the generationof extrachromosomal ribosomal DNA circles (ERCs), maintaining intact telomericchromatin, and ensuring appropriate distribution of oxidatively damaged proteins[5-8] . It was shown that the replicative lifespan of S. cerevisiae can be extendedby reducing the available sugar in the medium [9].

Lifespan extension by thisactivity of SIR2, and does not extend the lifespan when SIR2 is deleted [10]. However, SIR2-independent lifespan extension by CR has been observed in ayeast strain in which the SIR2 paralog HST2 is also capable of inhibiting ERCgeneration [11]. Lifespan extension by CR is abolished when both SIR2 andHST2 are deleted. These findings have interesting implications in lifespanregulation in higher organisms, which have multiple members of the SIR2 family.For instance, there are 4 SIR2 homologs in C. elegans and 7 in mammals [2].Multiple members of SIR2-like proteins may regulate lifespan cooperatively inresponse to diet. Evidence is emerging for elaborate cooperation between 7mammalian sirtuins in regulating CR response (see below). Another model of yeast aging has been developed, in which lifespan is measuredby the length of time that cells survive in a nondividing

state when exposed to ahypocaloric environment (chronological lifespan) [12]. In contrast to its effect onreplicative aging, SIR2 is not required for chronological life extension by CR [13,14]. These studies generate another layer of complexity in lifespan regulation bySIR2, which may have interesting implications in higher organisms. Whilechronological aging may be a good model for postmitotic cells, replicative agingmay more closely mirror mitotic cells in mammals [15]. It has been proposed thatnutrient-responsive signaling pathways regulate chronological aging andreplicative aging through different downstream effectors: increased stressresistance for chronological aging and decreased ribosome biogenesis forreplicative aging (Figure 1). It is logical that in mammals, aging in mitotic cellsand postmitotic cells are regulated through different mechanisms. In mitotic cells,such as stem cells, interventions

that increase their proliferation cause aging andthe exhaustion of the stem cell pool [16, 17]. Yet, this does not apply to postmiticcells, such as neurons, where increased ability to combat cell death and toincrease stress resistance plays a larger role [18]. Figure 1. CR regulation in yeast. Mild CR suppresses the ERCs and preventsreplicative aging dependent on SIR2. Severe CR suppresses ribosomebiogenesis and prevents replicative aging independent of SIR2. Chronologicalaging can be prevented by increased stress resistance and genomic stabilityinduced by severe CR and SIR2 deficiency. The hypothesis that yeast chronologic aging (which is not delayed by SIR2)represents a model for aging in post-mitotic cells, whereas yeast replicative aging(which is delayed by SIR2) models aging in mitotic cells may explain somepuzzling observations in SIR2 regulation of aging. Overexpression of the

SIR2homolog Sir-2.1 in Caenorhabditis elegans extends lifespan by up to 50% [19].However, resveratrol, a compound produced by plants under stress and anactivator of SIR2, increases lifespan by inhibiting Sir-2.1-mediated repression ofER stress genes [20]. One possibility is that under normal conditions, mitotic cellsmay play a larger role in lifespan determination than post-mitotic cells in worms.Thus, overexpression of Sir-2.1 extends lifespan. However, under stressconditions, post-mitotic cells may be more sensitive to stress and become thedetermining factor in lifespan. Therefore, Sir-2.1 inactivation leads to lifespanextension. The CR condition used in these studies (switching from minimum medium plus2% glucose to water) is drastically different from the one in which SIR2-dependent lifespan extension by CR is observed (switching from 2% glucosemedium to 0.5% glucose medium) [10, 21, 22].

Therefore, it is likely that differentpathways are activated in mild versus severe CR conditions (Figure 1). Mild CRincreases oxygen consumption and respiration, and leads to a reduction inNADH, an electron donor for respiration and a competitive inhibitor of SIR2 [21].In addition, the NAD salvage pathway is upregulated to synthesize NAD fromnicotinamide (NAM) and ADP-ribose, resulting in decreased levels of NAM,another inhibitor of SIR2 [23]. As a result, the silencing activity of SIR2 isactivated by mild CR. However, under severe CR condition, it is likely thatdownregulation of SIR2 contributes to increased chronological lifespan [24].sir2D mutants show increased expression of many genes involved in DNA repairand stress resistance, which are also activated by severe CR. Furthermore, SIR2deficiency, like severe CR, results in increased resistance to heat and oxidativedamage and elevated genomic stability.

Thus, the role of SIR2 in regulating CRresponse in yeast is context-dependent. For an extensive discussion on CRresponse in yeast, please refer to [25, 26]. Which CR condition, mild or severe, is more relevant to the one applied tomammals (30-40% reduction in calorie intake), which exhibits the maximumlifespan extension? One possibility is that when mammals are calorie restricted,tissues with different energy demands experience varying degrees of CR. Whenfood is scarce, mammals may have to redistribute the limited resource tomaintain survival and shut down unnecessary energy expenditures, such asgrowth, synthesis and reproduction. Thus, tissues necessary for basic survival,such as the muscle, the heart, and certain brain regions are protected fromstarvation and experience mild CR. However, tissues for synthesis, such as theliver and the pancreas, and the reproductive system are likely to

experiencesevere CR. Although in the circulation, levels of glucose are roughly the samethroughout the body, tissues may experience varied degrees of CR due todifferent capacities for glucose uptake. As an example, hepatocytes andpancreatic ß cells use glucose transporter GLUT2, which has a high Km andtherefore a low affinity for glucose. Thus, hepatocytes and ß cells mayexperience more severe CR compared to tissues like muscle and adipose tissue,which use glucose transporter GLUT4. Another possibility is that mild and severe CR conditions may also occur atdifferent time points during the day. CR animals are normally fed once a day oftheir daily quota or once every two days of double of their daily quota. After longtermCR, animals are trained to gorge their entire food supply within half an hourof feeding. During the course of each feeding cycle, CR animals may experiencea fed state, a mild

CR state and then a severe CR state. Thus, CR in mammalsmay resemble both mild and severe CR in yeast depending on tissue type andtime of day. Despite the lack of evidence for ERCs in organisms other than yeast, the activityof Sir2 homologs seem to affect lifespan in flies, worms, and mammals. InDrosophila melanogaster, increased expression of the Sir2 homolog dSir2extends lifespan [27], and dSir2 deficiency blocks the lifespan-extending effect ofCR. A non-sirtuin deacetylase, Rpd3, which is considered specific for histones,has also been suggested to mediate the effect of CR in Drosophila [28] and mayinteract with dSir2. Decreasing rpd3 expression results in elevated transcriptionlevels of dSir2, and mutations in dSir2 blocks the lifespan-extending effect ofrpd3 mutations. These observations suggest that dSir2 may act downstream ofrpd3 in mediating the effects of CR in Drosophila

[27]. It has been suggested that Sir-2.1 extends lifespan in C. elegans via activation ofthe transcription factor DAF-16 and probably also through additional targets [29].Nevertheless, there are contradictions regarding the role of Sir-2.1 in mediatingCR effects in C. elegans: whereas some studies support this role [30], othersdismiss it [31-33]. These differences may originate from the different approachesto limit nematode feeding, as Wang and Tissenbaum (2006) relied on a mutant(eat-2) with defects in the nematode's pharynx that resulted in lower feeding rate,while in the other studies it was the amount of food that was limited. Since thereare 4 SIR2 homologs in nematode, it would be interesting to see whether theother SIR2 homologs have functions redundant to Sir-2.1 and whether deletion ofall 4 genes would completely abolish CR-induced lifespan extension. The ability of sirtuins to

extend lifespan in yeast, flies, and nematodes viadifferent pathways is intriguing from the evolutionary perspective. This begs aquestion: do mammalian sirtuins extend lifespan and mediate CR response? Inmammals, there are 7 sirtuins, SIRT1-7, localized in various cellularcompartments [34]. The lifespans of sirtuin knockout or transgenic mice are yetto be determined. However, it is intriguing that polymorphisms of certain sirtuinshave been linked to human longevity [35], although this study requires furtherconfirmation [36]. In addition, emerging genetic evidence suggests thatmammalian sirtuins are required for the CR response. SIRT1 may be required forincreased physical activity of CR mice, as SIRT1 knockout mice do not haveincrease activity [37]. It is likely that SIRT1 in the hypothalamus is necessary tosense hunger and trigger the foraging response. In addition, genetic, molecularand pharmacological

studies discussed below are all consistent with the role ofmammalian sirtuins in regulating the CR response. SIRT1 and metabolic regulation It has been thought that CR extends lifespan by decreasing metabolism and theassociated production of damaging reactive oxygen species. However, thistraditional view has been challenged by recent findings indicating that CR in factincreases metabolism. CR yeast switches metabolism from fermentation towardthe mitochondrial tricarboxylic acid cycle and increased respiration [22]. Similarly,increased oxygen consumption has been observed in CR worms [38]. Evidencefor changes in oxygen consumption in CR mice is controversial, as the miceexperience drastic alterations in body weight and composition [39]. However, CRmice do experience a metabolic switch that mirrors the one observed in CRyeast. When energy is abundant, instead of storing excess energy in the

form ofethanol, animals store it as fat. During CR, animals turn on fatty acid oxidationand switch fuel usage from glucose to fatty acids. Since fatty acids are morereduced than glucose, fatty acid oxidation will consume more oxygen per carbonthan glycolysis. At the cellular level, increased mitochondrial biogenesis in CRtissues suggests a tissue-specific increase in metabolic rate of CR animals [40]. How do sirtuins regulate these metabolic changes? SIRT1, a nuclear sirtuin,coordinates the metabolic switch in multiple tissues by regulating differentmolecular targets (Figure 2). In the white adipose tissue (WAT), SIRT1 negativelyregulates the nuclear receptor PPARg, a key regulator of adipocyte differentiationand fat storage [41]. Thus, activation of SIRT1 suppresses fat storage andpromotes the mobilization of fat from the WAT to be utilized by other tissues. Inthe muscle, SIRT1 activates

PGC-1-alpha, a master regulator of mitochondrialbiogenesis and fatty acid oxidation [42]. Consistent with the role of SIRT1 inpromoting fat mobilization and fatty acid oxidation, SIRT1 activity seems to beupregulated in the muscle and the WAT of CR mice. Both SIRT1 expression andNAD level are increased in these two tissues [43]. Figure 2. Tissue-specific regulation of SIRT1 by CR. SIRT1 is downregulatedin the liver and pancreas of CR mice, resulting in decreased gluconeogeneis andinsulin secretion. It is upregulated in the muscle and the WAT, leading toincreased mitochondrial biogenesis, fatty acid oxidation and insulin sensitivity.These metabolic changes cause a cellular energy-deficit state and activation ofAMPK. Activated AMPK further amplifies these metabolic changes. SIRT1activators mimic the CR response by activating PGC-1-alpha in the muscle. ...

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