doi: 10.1161/01.RES.0000148666.20729.1c
© 2004 American Heart Association, Inc.
Editorial |
Sirviving Cardiac Stress
Cardioprotection Mediated by a Longevity Gene
From the Department of Medicine, Johns Hopkins University, Baltimore, Md.
Correspondence to Michael T. Crow, PhD, Department of Medicine, Division of Pulmonary and Critical Care Medicine, Johns Hopkins Asthma and Allergy Center, 5A.58, 5501 Hopkins Bayview Circle, Baltimore, MD 21224. E-mail mcrow1{at}jhmi.edu
Key Words: cardioprotection • cardiac stress • insulin signaling
Over the last decade, a number of genetic modifiers of yeast and animal lifespan have been discovered and studied in some detail. Many of these genes lie on or impinge on the evolutionarily conserved insulin signaling pathway.1 In fact, one of the first genes discovered that was linked to lifespan regulation in Caenorhabditis elegans was DAF2, the product of which is highly related to mammalian insulin/insulin-like growth factor (IGF) receptors.2 Many of the intermediates activated by insulin intracellular signaling, such as phosphatidylinositol 3' kinase (PI3K) and AKT/PKB, mediate cytoprotection associated not only with the insulin receptor but other receptor-mediated signaling pathways as well.3 Preventing the cell death that occurs in response to stress would seem to be a reasonable component of an overall cellular strategy for extending lifespan because the cumulative effects of cell loss have been implicated in various age-related disorders, including cardiovascular disease. Increased activity of prosurvival molecules through increased growth factor/insulin signaling would seem to be one way to achieve extension. Yet the lifespan extending-mutation in the C elegans DAF2 gene caused decreased, rather than increased, signaling. Although it seems counterintuitive that decreased prosurvival signaling is capable of extending lifespan, the notion is supported by the remarkable success that caloric restriction has had in increasing lifespan across the phylogenic spectrum.4 Here too lifespan extension is associated with reduced glucose, insulin, and IGF1 plasma levels, as well as reduced insulin signaling.5 Clearly, either the role of cytoprotection in lifespan extension has been overstated or the complexities of insulin signaling and its downstream effectors underestimated.
In this issue of Circulation Research, Alcendor et al6 report on the expression of the longevity factor encoded by SIRT1 (Sir2
) and its potential role as an endogenous inhibitor of stress-mediated cell death in isolated neonatal cardiomyocytes. This study is newsworthy not because it introduces yet another prosurvival factor that is capable of preventing cell death in cultured cardiomyocytes, but because it is a prosurvival factor that is transcriptional regulator linked to lifespan extension, and as such, may confer survival that is fundamentally different from that provided by overexpressing constitutively activated signaling intermediates, such as Akt/PKB and PI3-Kinase. SIRT1 is the mammalian homologue of the yeast SIR (silent information regulator) 2 gene. SIR2 encodes a histone deacetylase (HDAC) that was originally identified for its role in chromatin reorganization and silencing. It is highly conserved from archaebacteria to humans. In mammals, there are seven SIR2 orthologs (SIRT1–7), which collectively are known as the Sirtuins.7 Of the genes that encode active enzymes, one is expressed in the nucleus and PML bodies (SIRT1), one in the cytoplasm (SIRT2), and one in the mitochondria (SIRT3). Another active ortholog has been reported (SIRT5) but its intracellular localization is unknown at this time.8
The connection of SIR2 to lifespan extension is compelling. In both yeast and C elegans, extra copies of SIR2 are sufficient to increase lifespan, whereas sirtuin activators mimic the effects of caloric restriction in yeast, worms, and flies.9–11 Deletion of SIR2 in C elegans prevents the increase of longevity associated with the insulin receptor mutant DAF2,12 whereas in yeast, deletion of SIR2 prevents lifespan extension caused by caloric restriction and by sublethal levels of stress.13–15 Finally, Sir2 protein levels increase in rodents under caloric restriction and in human cells treated with serum from these animals.16
Among HDACs, Sir2 is unique in that its deacetylase activity is NAD-dependent (see Figure, A). Sir2 requires oxidized nicotinamide adenine dinucleotide (NAD) as a cofactor and is negatively regulated by either NADH or the product of the deacetylase reaction, nicotinamide (NAM), through competitive and noncompetitive mechanisms, respectively. Genetic manipulations in yeast that decrease the NAD/NADH ratio (deletion of NADH-dehydrogenase (NADH-DH) or prevent entry of NAM through the salvage pathway (deletion of PCN1, the gene encoding nicotinamidase [NAMase]) inhibit Sir2 activity. Whereas NAD salvage pathways in mammals do differ,17 the concept is the same, namely that genetic manipulations to increase NAD will increase Sir2 activity.
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The dependency of Sir2 on NAD potentially links the activation of Sir2 to metabolic changes that occur during glucose (caloric) restriction in yeast. These include an obligatory switch from fermentation to respiration, which causes an increase in the NAD/NADH ratio.15 In animals, both insulin and IGF1 plasma levels markedly decrease in response to caloric restriction,16 but whether changes in NAD/NADH also occur is controversial. It is important, however, to remember that Sir2 is negatively regulated by NAM and that changes in the level of this intermediate may also regulate the activity of Sir2 independently of NAD and NADH levels.
In the Alcendor article,6 the authors show that rat Sir2 is expressed in the nucleus and that its protein levels fall slightly during serum deprivation, and actually slightly increase in response to hydrogen peroxide. However, these slight changes in expression may mask much larger changes in activity if there are concurrent alterations in NAD metabolism. Interestingly, the most robust change in Sir2 protein levels was seen in dog hearts in which the aorta was banded and the heart then paced to induce left ventricular hypertrophy (LVH) that decompensates into heart failure. No change in Sir2 levels were observed in dogs that were exposed to aortic banding alone. These animals developed stable LVH, suggesting that the changes in Sir2 seen in banded/paced animals were associated with maladaptive remodeling of the heart, a process that involves increased cell death. Perhaps, as the authors suggest, the large upregulation in Sir2 expression represents a failed attempt to prevent cell death.
In isolated neonatal cardiomyocytes, inhibitors of Sir2 activity cause a moderate increase in basal cell death and an upregulation in the expression of the hypertrophy-associated gene, atrial naturietic factor (ANF), even though cell size actually decreases. An acetylase-dead (dominant-negative) mutant of Sir2 (dnSir2) also increased basal cell death as well as slightly potentiating hydrogen peroxide-induced cell death. Likewise, increasing Sir2 levels by adenoviral-mediated gene transfer protected myocytes from serum starvation-associated cell death, while also increasing overall cell size. The cell death that accompanied decreased Sir2 activity is, for the most part, apoptotic, but where in the cell death signaling pathway Sir2 intervenes was not addressed. A factor that was not considered in this study was the potential role of other Sirtuins that might be expressed in the myocytes, namely those encoded by SIRT2 in the cytosol and SIRT3 in the mitochondria. The deacetylase activity of these molecules would be affected by the inhibitors that were used in the study and, although dnSir2 could affect the cytoplasmic Sirtuin encoded by SIRT2, a systematic targeted knockdown of these Sirtuins will be necessary to clarify the role of all Sirtuins in the cell.
Nonetheless, the results from the article support the contention that Sir2 is an endogenous suppressor of cell death in, at least, neonatal cardiomyocytes. Given the low level of unstimulated cell death and the modest potentiation of cell death caused by the inhibitors, it appears, however, to be a weak suppressor. Then again, the stimuli that were used were limited. Other death-inducing stimuli that mimic the stresses that the heart experiences under pathological conditions, such as hypoxia, acidification, or exposure to reactive oxygen species (ROS) were not examined.
The question then is how does Sir2 mediate cytoprotection? B in the Figure is a partial list of the targets susceptible to deacetylation by Sir2 and the effects that deacetylation has on their function. Alcendor et al6 focused on p53 as a target in their study. They show that it is deacetylated in the nucleus and that this is likely to be mediated by Sir2, because NAM increased its acetylation. Inhibiting Sir2 also increased the ability of p53 to transactivate genes, as measured with a p53 response element linked to a minimal promoter driving the luciferase gene. To assess the functional significance of these changes to cell death caused by Sir2a inhibition by NAM, the authors show that transactivation-defective p53 mutant, which acts as a dominant negative for wild type p53-mediated gene transcription (dnp53) blocks the increase in cell death caused by NAM. However, dnp53 had no statistically significant effect in preventing the cell death caused by serum deprivation, even though overexpression of Sir2 did, suggesting that not all Sir2a-mediated effects are p53-mediated.
In C elegans, the ability of SIR2 to extend lifespan is dependent on the gene DAF16.10 The mammalian homolog of DAF16 is the transcription factor family known as FOXO (forkhead box-containing protein, O subfamily). The four known members of this subfamily, FOXO1, FOXO3, FOXO4, and FOXO6 control the transcription of genes that are involved in cell growth, cell death, responses to stress and DNA damage, differentiation, and the regulation of metabolism.18 In the presence of serum or growth factors, FOXO is rendered transcriptionally inactive by nuclear exclusion resulting from Akt/PKB-mediated phosphorylation and cytoplasmic sequestration (see Figure, C). ROS can also promote FOXO nuclear exclusion through p66shc-mediated activation of Akt/PKB.19 On the other hand, reduced signaling activity caused by serum-deprivation or mutations that affect upstream signaling pathways (eg, the DAF2 mutant in C elegans) shifts the distribution of FOXO toward the nucleus, where it is now active. In the nucleus, FOXO can be acetylated, the extent of which is controlled in part by the opposing effects of the Sir2 deacetylase. The functional consequences of partial deacetylation by Sir2 are somewhat controversial. One group has reported that FoxO deacetylation decreases but does not abolish both target gene expression across the board including both antistress and prodeath genes,20 whereas another proposes that the effects of deacetylation are target gene specific, with the antistress gene expression increased, and prodeath gene expression decreased.21 This would tip the balance between FOXO-driven antistress and prodeath genes in favor of cytoprotection.
In the FOXO pathway, then, there are two ways to prevent cell death: one in which FoxO is rendered transcriptionally inactive through Akt/PKB-mediated nuclear exclusion, and another in which FoxO is only partially active as a transcription factor. The latter is dependent on Sir2 activity and represents a gain-of-function for FoxO. It is this gain-of-function that appears to be necessary for lifespan extension and that, at least for the moment, makes understanding the effects of Sir2 in myocytes intriguing. Whether the gain-of-function involves an indiscriminant reduction in transcriptional activity or selective transcription of certain FoxO targets or both remains to be resolved. But it is important to remember that FoxOs are not just transcriptional activators, but transcriptional suppressors as well. Therefore, lowering the overall activity of FoxO could produce not only a quantitative but qualitative change in gene expression as well. Secondly, the number of identified FoxO targets continues to increase and the relevant targets for lifespan extension and cytoprotection may not just be prodeath genes, such as FasL and BIM, but other genes linked to differentiation or metabolism or genes yet to be discovered. Whether the regulation of FoxO genes is relevant to cytoprotection of cardiac myocytes in response to serum deprivation was not addressed by Alcendor et al,6 but it is an attractive area for future investigation, as are other Sir2 targets, such as Ku70 (Figure, B).
In summary, the literature on cytoprotection in the heart is replete with studies showing that overexpression of constitutively active forms of PI3K or Akt/PKB can be protective, whereas inhibition of these same kinases blocks the cytoprotective effects caused by increased signaling through insulin and growth factor-stimulated pathways. Yet recent studies on the genetics of aging and the molecular mechanisms that underlie the remarkable lifespan extension caused by caloric restriction indicate that reduced activity through the insulin signaling pathway is not only cytoprotective but leads to additional benefits culminating in lifespan extension. What these additional benefits are is unknown but they are likely to affect the long-term sensitivity of the heart to stress. The current study by Alcendor et al6 is likely to spark interest among heart researchers to address not only the issues raised in this commentary but to understand how important Sir2 is to maintain myocyte viability and function.
Footnotes
The opinions expressed in this editorial are not necessarily those of the editors or of the American Heart Association.
References
1. Barbieri M, Bonafe M, Franceschi C, Paolisso G. Insulin/IGF-1 signaling pathway: an evolutionarily conserved mechanism of longevity from yeast to humans. Am J Physiol Endocrinol Metab. 2003; 238: E1064–E1074.
2. Kimura KD, Tissenbaum HA, Liu Y, Ruvkun G. Daf-2, an insulin receptor-like gene that regulates longevity and diapause in Caenorhabditis elegans. Science. 1997; 277: 942–946.
3. Downward J. PI3-kinase, Akt, and cell survival. Semin Cell Dev Biol. 2004; 15: 177–182.[CrossRef][Medline] [Order article via Infotrieve]
4. Masoro EJ. Dietary restriction: current status. Aging Clin Exp Res. 2001; 13: 261–262.
5. deCabo R, Furer-Galban S, Anson RM, Gilman C, Gorospe M, Lane MA. An in vitro model of caloric restriction. Exp Ger. 2003; 38: 631–639.
6. Alcendor RR, Kirshenbaum LA, Imai S-i, Sadoshima J. Silent information regulator 2, a longevity factor and a class III histone deacetylase, is an essential endogenous inhibitor of apoptosis in cardiac myocytes. Circ Res. 2004: 95: 971–980.
7. North BJ, Verdin E. Sirtuins: Sir2-related NAD-dependent protein deacetylases. Genome Biol. 2004; 5: 224.[CrossRef][Medline] [Order article via Infotrieve]
8. Blander G, Guarente L. The Sir2 family of protein deacetylases. Annu Rev Biochem. 2004; 73: 417–435.[CrossRef][Medline] [Order article via Infotrieve]
9. Kaeberlin M, McVey M, Guarente L. The SIR2/3/4 complex and SIR2 alone promote longevity in Saccharomyces cerevisiae by two different mechanisms. Genes Dev. 1999; 13: 2570–2580.
10. Tissenbaum HA, Guarente L. Increased dosage of a sir-2 gene extends lifespan in Caenorhabditis elegans. Nature. 2001; 420: 227–230.
11. Howitz KT, Bitterman KJ, Cohen HY, Lamming DW, Lavu S, Wood JG, Zipkin RE, Chung P, Kisielewski A, Zhang LL, Scherer B, Sinclair DA. Small molecule activators of sirtuins extend Saccharomyces cerevisiae lifespan. Nature. 2003; 425: 191–196.[CrossRef][Medline] [Order article via Infotrieve]
12. Hekimi S, Guarente L. Genetics and the specificity of the aging process. Science. 2003; 299: 1351–1354.
13. Lin SJ, Defossez PA, Guarente L. Requirement of NAD and Sir2 for lifespan extension by calorie retraction in Saccharomyces cerevisiae. Science. 2000; 289: 2126–2128.
14. Anderson RM, Bitterman KJ, Wood JG, Medvedik O, Sinclair DA. Nicotinamide and PNC1govern lifespan extension by calorie restriction in Saccharomyces cerevisiae. Nature. 2003; 423: 181–185.[CrossRef][Medline] [Order article via Infotrieve]
15. Lin S-J, Kaeberlein M, Andalis AA, Sturtz LA, Defossez P-A, Cullota VC, Fink GR, Guarente L. Calorie restriction extends Saccharomyces cerevisiae lifespan by increasing respiration. Nature. 2002; 418: 344–348.[CrossRef][Medline] [Order article via Infotrieve]
16. Cohen HY, Miller C, Bitterman KJ, Walt NR, Hekking B, Kessler B, Howitz KT, Gorospe M, de Cabo R, Sinclair DA. Calorie restriction promotes mammalian cell survival by inducing the SIRT1 deacetylase. Science. 2004; 305: 390–392.
17. Revollo JR, Grimm AA, Imai SI. The NAD biosynthesis pathway mediated by nicotinamide phosphoribosyltransferase regulates Sir2 activity in mammalian cells. J Biol Chem. In press.
18. Accili D, and Arden KC. FoxOs at the crossroads of cellular metabolism, differentiation, and transformation. Cell. 2004; 117: 421–426.[CrossRef][Medline] [Order article via Infotrieve]
19. Nemoto S, and Finkel T. Redox regulation of Forkhead proteins through a p66shc-dependent signaling pathway. Science. 2002; 285: 2450–2452.
20. Motta MC, Divecha N, Lemieux M, Kamel C, Chen D, Gu W, Bultsma Y, McBurney M, Guarente L. Mammalian SIRT1 represses forkhead transcription factors. Cell. 2004; 116: 551–563.[CrossRef][Medline] [Order article via Infotrieve]
21. Brunet A, Sweeney LB, Sturgill JF, Chua KF, Greer PL, Lin Y, Tran H, Ross SE, Mostoslavsky R, Cohen HY, Hu LS, Cheng H-L, Jedrychowski MP, Gygi SP, Sinclair DA, Alt FW, Greenberg ME. Stress-dependent regulation of FOXO transcription factors by the SIRT1 deacetylase. Science. 2004; 303: 2011–2015.
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