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Sir2 Links Chromatin Silencing, Metabolism, and Aging

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Sir2 Links Chromatin Silencing, Metabolism, and Aging Downloaded from genesdev.cshlp.org on October 3, 2021 - Published by Cold Spring Harbor Laboratory Press PERSPECTIVE Sir2 links chromatin silencing, metabolism, and aging 1Leonard Guarente Department of Biology, Massachusetts Institute of Technology (MIT), Cambridge, Massachusetts 02139 USA Aging is manifested by a progressive decline in vitality at mating type loci (Rine and Herskowitz 1987) and telo- over time leading to death. Studies in budding yeast al- meres (Gottschling et al. 1990), and SIR2, but not SIR3 or low aging to be followed in individual pedigrees of cells, SIR4, is required for silencing in the rDNA (Bryk et al. that is, those of mother cells, consequent to many 1997; Smith and Boeke 1997). Silencing causes a more rounds of cell division (Mortimer and Johnston 1959). closed, inaccessible regional chromatin structure, as as- These studies have led to the general conclusion that the sayed by various probes of DNA accessibility (Loo and silencing protein Sir2 is a limiting component of longev- Rine 1994; Bi and Broach 1997). Even though expression ity; deletions of SIR2 shorten life span and an extra copy of marker genes inserted into the rDNA is repressed, of this gene increases life span (Kaeberlein et al. 1999). silencing of rDNA transcription itself may be more mod- Recent studies have spurred interest in Sir2 as a candi- est, as continued ribosome synthesis is essential for date longevity factor in a broad spectrum of eukaryotic growth. The Sir proteins may also function in DNA re- organisms. SIR2 gene homologs have been found in a pair by nonhomologous end-joining (NHEJ) (Tsukamoto very wide range of organisms ranging from bacteria to et al. 1997; Boulton and Jackson 1998). In this regard, the humans (Brachmann et al. 1995). Moreover, a biochemi- Sir2/3/4 proteins and Ku relocalize from telomeres to cal activity of Sir2 likely responsible for chromatin si- sites of DNA breaks to aid in their repair by NHEJ (Fig. lencing, nicotinamide–adenine dinucleotide (NAD)-de- 1A) (Martin et al. 1999; Mills et al. 1999). A primary role pendent histone deacetylase, has recently been discov- of the Sir complex at telomeres therefore may be to pro- ered and shown to be broadly conserved (Imai et al. vide a reservoir of factors that can be mobilized for the 2000). In this review, I will briefly discuss silencing as it immediate repair of DNA damage. pertains to SIR2 and its relationship to aging. I will then The function of Sir2 in promoting longevity in yeast trace the studies that led to the discovery of the NAD- mother cells appears to relate to silencing in the rDNA. dependent histone deacetylase. I will next speculate how The stability of the 100–200 tandem copies of rDNA on the regulation of Sir2 by NAD could represent the link chromosome XII requires SIR2, as the frequency of re- between caloric intake and the pace of aging, which is combination at that locus increases about 10-fold in sir2 widely observed in many organisms (Weindruch et al. mutants (Gottlieb and Esposito 1989). One of the prod- 1986). Finally, I will present a speculative model of how ucts of rDNA recombination is extrachromosomal a gradual disruption in chromatin silencing may occur rDNA circles (ERCs) (Fig. 1B), which, once formed, rep- and how such a change may cause aging. licate and segregate preferentially to mother cell nuclei (Sinclair and Guarente 1997). ERCs thus accumulate in mother cells as they grow older and ultimately trigger Maintenance of chromatin silencing and genome senescence. At least one function of Sir2 in yeast longev- stability by Sir2 ity, therefore, is to forestall the appearance of the first rDNA circle in mother cells by creating a silenced chro- Silencing of genomic DNA was first observed by repres- matin structure. sion of genes near certain translocation breakpoints in Silencing requires particular lysines in the extended Drosophila (for review, see Wakimoto 1998). Studies in amino-terminal tail of histones H3 and H4 (Thompson et Drosophila and yeast have led to the identification of al. 1994; Hecht et al. 1995; Braunstein et al. 1996). These factors that act in trans to mediate silencing. Among and other lysines of the tail are acetylated in active chro- these are the proteins encoded by the yeast SIR genes, matin but deacetylated in silenced chromatin (Braun- which are responsible for silencing at repeated DNA se- stein et al. 1993, 1996). The deacetylated histones evi- quences in yeast: mating type loci, telomeres, and the dently can fold into a more compact, closed nucleosomal rDNA. SIR2, SIR3, and SIR4 are all required for silencing structure (Luger et al. 1997). These considerations led to the suggestion that Sir2 could be a histone deacetylase. Further evidence for this claim arose from the global 1E-MAIL [email protected]; FAX (617) 253-8699. deacetylation of yeast histones observed when Sir2 was GENES & DEVELOPMENT 14:1021–1026 © 2000 by Cold Spring Harbor Laboratory Press ISSN 0890-9369/00 $5.00; www.genesdev.org 1021 Downloaded from genesdev.cshlp.org on October 3, 2021 - Published by Cold Spring Harbor Laboratory Press Guarente Figure 1. Functions of Sir2 in yeast. Sir2 mediates silencing at telomeres, along with Sir3, Sir4, and Ku, (A) and at the re- peated rDNA (B) without these other fac- tors. Telomeric proteins respond to DNA double-strand breaks (DSBs) by moving to sites of damage in S-phase in a pathway requiring MEC1, RAD9, RAD53.Inasir2 mutant, homologous recombination in the rDNA increases leading to more ERCs. overexpressed (Braunstein et al. 1993). However, at- Because the deacetylase activity of Sir2 occurs prefer- tempts to demonstrate a histone deacetylase activity by entially on histone residues that are essential for silenc- Sir2 in vitro initially met with failure. ing, we infer that it is this activity, rather than the ADP– ribosyltransferase, that triggers silencing in vivo. Con- sistent with this claim, a mutation of Gly-270 of Sir2 to Sir2 is a conserved NAD-dependent histone deacetylase Ala reduces the ADP–ribosyltransferase by 93%, but re- duces the deacetylase activity by only 20% and still can Unlike SIR3 and SIR4, the SIR2 gene is broadly con- function in silencing, repression of rDNA recombina- served in organisms ranging from bacteria to humans tion, and extension of life span (Imai et al. 2000). Thus, (Brachmann et al. 1995). Studies on the bacterial homo- Sir2 is an NAD-dependent histone deacetylase that may log, cobB, led to the conclusion that this gene could sub- link metabolism and silencing in vivo (Fig. 2). The role of stitute for another bacterial gene, cobT, in the pathway the ADP–ribosyltransferase in vivo is still not clear, but of cobalamin synthesis (Tsang and Escalante-Semerena this activity is evidently separable from the deacetylase, 1998). cobT was known to encode an enzyme that trans- as a known inhibitor of mono-ADP–ribosyltransferases ferred ribose–phosphate from nicotinic acid mono- selectively inhibits the one activity of Sir2 and not the nucleotide to dimethyl benzimidazole. Thus, it seemed other (Imai et al. 2000). The ADP–ribosyltransferase may possible that Sir2 proteins might be equipped to catalyze turn out to be important to the function of Sir2 in DNA a related reaction at the nicotinamide–ribose bond in repair, as nuclear mono- and poly-ADP–ribosyltransfer- NMN and perhaps nicotinamide-adenine dinucleotide ases have been associated with DNA repair in mamma- (NAD), in the latter case resulting in transfer of ADP– ribose. Indeed, it was shown by Frye (1999) that Sir2 proteins from bacteria, yeast, or mammals were able to transfer 32P from NAD to a protein carrier, suggesting that they were ADP–ribosyl transferases. Subsequent work proved that Sir2 could, in fact, transfer ADP–ri- bose, albeit in a reaction that proceeds only weakly in vitro (Tanny et al. 1999). This latter study led to the proposal that the ADP–ribosyltransferase activity of Sir2 was essential to the in vivo function of silencing. In studying this ADP–ribosyl transferase reaction, we noticed that peptides of the amino-terminal tails of his- tone H3 or H4 could accept 32P from NAD, but only if the peptides were acetylated. Using acetylated H3, we separated the Sir2-modified product by chromatography and found by mass spectrometry that the molecular weight of the product was actually smaller by 42, indi- cating that the major modification catalyzed by Sir2 was deacetylation and not ADP ribosylation (Imai et al. Figure 2. Sir2 is an NAD-dependent histone deacetylase. The 2000). When NAD was omitted, no deacetylation by Sir2 deacetylation of lysines in the amino-terminal tails of histones occurred. NADH, NADP, or NADPH could not substi- H3 and H4 in nucleosomes (NUC) is proposed to convert active tute for NAD in this reaction. The weak ADP–ribosyl- to silenced chromatin. Sir2 is stimulated to carry out this reac- transferase reaction did not generate sufficient levels of tion by NAD, the available levels of which are likely coupled to product to allow detection by this physical method. the metabolic rate of cells. 1022 GENES & DEVELOPMENT Downloaded from genesdev.cshlp.org on October 3, 2021 - Published by Cold Spring Harbor Laboratory Press Sir2, chromatin, and aging lian cells (Kreimeyer et al. 1984; Pero et al. 1985; Meyer carbon flow through glycolysis would be high (Fig. 3A). and Hilz 1986). The glycolytic enzyme glyceraldehyde-3-P-dehydroge- nase (GAPDH) uses NAD, and the resulting NADH is recycled by the delivery of electrons to oxygen via the Is Sir2 a link between metabolic rate and aging? electron transport chain or, if oxygen is scarce, to acetyl- One of the most consistent observations in aging is the CoA to generate fermentation products.
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