Sirtuins: critical regulators at the crossroads between

Oncogene (2007) 26, 5489–5504
& 2007 Nature Publishing Group All rights reserved 0950-9232/07 $30.00
Sirtuins: critical regulators at the crossroads between cancer and aging
LR Saunders and E Verdin
Gladstone Institute of Virology and Immunology, University of California, San Francisco, CA, USA
Sirtuins (SIRTs 17), or class III histone deacetylases
(HDACs), are protein deacetylases/ADP ribosyltransferases that target a wide range of cellular proteins in the
nucleus, cytoplasm, and mitochondria for post-translational modification by acetylation (SIRT1, -2, -3 and -5)
or ADP ribosylation (SIRT4 and -6). The orthologs of
sirtuins in lower organisms play a critical role in regulating lifespan. As cancer is a disease of aging, we discuss
the growing implications of the sirtuins in protecting
against cancer development. Sirtuins regulate the cellular
responses to stress and ensure that damaged DNA is not
propagated and that mutations do not accumulate. SIRT1
also promotes replicative senescence under conditions of
chronic stress. By participating in the stress response to
genomic insults, sirtuins are thought to protect against
cancer, but they are also emerging as direct participants in
the growth of some cancers. Here, we review the growing
implications of sirtuins both in cancer prevention and as
specific and novel cancer therapeutic targets.
Oncogene (2007) 26, 5489–5504; doi:10.1038/sj.onc.1210616
Keywords: sirtuins; cancer; lifespan; lysine acetylation
Cancer is a disease of aging, as the incidence of most
cancers increases with age following an accumulation of
mutations. Mutations arise from errors during DNA
replication, environmental insults, and are induced by
intracellular reactive oxygen species that increase during
cellular stress. In many organisms, longer lifespan
correlates with lower levels of reactive oxygen species
(Finkel and Holbrook, 2000). Additionally, age-related
changes may alter the internal microenvironment to
favor tumor development (Anisimov et al., 1988). As the
global human population ages, the ability to prevent
and treat cancer becomes essential to allow people to
live longer, healthier lives.
Members of the silent information regulator 2 (Sir2)
family, which are conserved from bacteria to humans,
regulate lifespan in various organisms (Longo and
Correspondence: Dr E Verdin, Gladstone Institute of Virology and
Immunology, University of California, 1650 Owens St, San Francisco,
CA 94158, USA.
E-mail: [email protected]
Kennedy, 2006). Seven human sirtuins (SIRT1–7), also
referred to as class III histone deacetylases (HDACs),
share a catalytic domain of B275 amino acids. Because
of their established role in controlling lifespan in lower
organisms, there is growing interest in determining
whether the same effect can be seen in mammals and in
humans in particular (Figure 1).
The most effective and reproducible way to extend
lifespan is with a calorie-restricted diet (Roth et al.,
2001; Hursting et al., 2003). Calorie restriction (CR),
a 2040% lowering of caloric intake, reduces spontaneous tumor formation in several mouse models in
which a tumor suppressor is inactivated or an oncogene
is overexpressed (Jones et al., 1997; Berrigan et al.,
2002; Mai et al., 2003). Aging is associated
with increased rates of stress-induced apoptosis,
and CR lowers sensitivity to stress-induced apoptosis
(Higami et al., 1997; Ando et al., 2002). Levels of
SIRT1 are increased in response to CR, and the
enzymatic activity of SIRT1 is required for resistance
to apoptosis induced by CR (Cohen et al., 2004b;
Nemoto et al., 2004). CR also induces SIRT3 expression, but appears to decrease SIRT4 (Shi et al.,
2005; Haigis et al., 2006). Mice lacking the insulin
receptor in adipose tissue live 18% longer on a regular
diet, suggesting that a reduction in fat mass rather than
lower caloric intake mediates longer lifespan (Bluher
et al., 2003). Changes in energy balance and body mass
composition lower cancer incidence in CR mice (Huffman et al., 2007). Because the prevalence of obesity in
humans is expected to increase more than 40% by 2030,
cancer incidence is also expected to rise (Vainio et al.,
As sirtuins promote longevity, one would expect that
sirtuin activation or increased expression, should protect
against cancer. Sirtuins could mediate this protective
effect by limiting replicative lifespan, by protecting
against DNA damage and oxidative stress or by
guarding against accumulation of mutations and against
genomic instability. In contrast, the loss of sirtuin
expression, activity or regulation can bypass replicative
senescence, allow cell division to proceed without the
proper repair of DNA, and promote accumulation of
mutations and genomic instability leading to tumor
development. There is also growing evidence that tumor
cells can become ‘addicted’ to sirtuin overexpression,
which allows rapid proliferation and loss of checkpoints
to promote continued propagation in the presence of
accumulating mutations.
Sirtuins and cancer
LR Saunders and E Verdin
Figure 1 Cellular localization, enzymatic activities and targets of the mammalian sirtuins. SIRT1 responds to changes in nutrient
availability and cellular stress to promote cell survival by deacetylating histone and non-histone targets in the nucleus. SIRT2 is mainly
localized in the cytoplasm and deacetylates a-tubulin, but during mitosis, it shuttles to the nucleus and deacetylates histone H4-K16 to
promote chromatin condensation. SIRT2 may function as part of a mitotic checkpoint to ensure that cells do not pass through mitosis
if a stress signal or DNA damage is present. SIRT3, 4 and 5 localize to the mitochondria. SIRT3 deacetylates and activates acetyl-CoA
synthetase 2 (AceCS2) to enhance acetyl-coA production. SIRT4 ADP-ribosylates and represses glutamate dehydrogenase (GDH) to
suppress insulin signaling. No targets or enzymatic activity have been identified for SIRT5. SIRT6 promotes DNA repair and guards
against genomic instability in the nucleus. Its targets have not been identified. SIRT7 localizes to the nucleolus where it interacts with
RNA pol-I to promote transcription of rRNA genes. SIRT1, sirtuins.
These emerging data point to sirtuins as critical
regulators at the crossroads between cancer and aging.
Unraveling the cellular roles of the sirtuins in the control
of cell growth and as guardians against genomic damage
is likely to lead to the identification of novel cancer
prevention and treatment strategies.
Sirtuin enzymatic activities: deacetylation and
The conserved catalytic domain shared by the sirtuins
functions as a mono-ADP-ribosyltransferase and as
a b-nicotinamide adenine dinucleotide (NAD þ )-dependent lysine deacetylase (Figure 2) (Frye, 1999;
Imai et al., 2000; Landry et al., 2000; Smith et al.,
2000). The deacetylase domain in the sirtuins is different
from that of class I and II HDACs, which are Zn þ dependent enzymes (North and Verdin, 2004; Longo
and Kennedy, 2006). Sirtuins use one NAD þ molecule
and generate acetyl-ADP-ribose and nicotinamide
during the deacetylation reaction (Landry et al., 2000).
While SIRT1, 2, 3 and 5 have significant deacetylase
activity towards a histone H4 peptide, SIRT4, 6
and 7 have low to undetectable deacetylase activity
in vitro on tested substrates (Langley et al., 2002;
North et al., 2003). SIRT1 deacetylates histones and
other nuclear targets, including p53, Ku70 and FOXO
(Forkhead box, class O) in vivo (as discussed below).
Of the sirtuins, only SIRT2 deacetylates tubulin,
also a target of the class II deacetylase HDAC6
(Hubbert et al., 2002; Matsuyama et al., 2002; North
et al., 2003).
Mono-ADP-ribosylation, during which ADP-ribose
from NAD þ is transferred to an acetylated target
protein, is conserved from bacteria to humans and is
carried out by several families of proteins (Hassa et al.,
2006). Mono-ADP-ribosylated cellular proteins include
histones, high mobility group (HMG) family members,
actin, a- and b-tubulin and glutamate dehydrogenase
(GDH; Hassa et al., 2006). SIRT1 preferably transfers
mono-ADP-ribose to histone H1 (Frye, 1999). SIRT6 is
more efficient with bovine serum albumin (BSA), and
SIRT2 can also ADP-ribosylate BSA in vitro (Frye,
1999; Liszt et al., 2005). SIRT6 robustly mono-ADPribosylates itself through an intramolecular reaction
(Liszt et al., 2005). In vivo, SIRT4 ADP-ribosylates
GDH, which inhibits its activity (Haigis et al., 2006). A
conserved histidine residue in the catalytic core of the
sirtuins is important for their deacetylase and monoADP-ribosyltransferase activities (Frye, 1999). Deacetylation and ADP-ribosylation may be linked and
dependent on each other. ADP-ribose generated in the
deacetylation reaction is added to a substrate following
Sirtuins and cancer
LR Saunders and E Verdin
Figure 2 Enzymatic activities of sirtuins. The conserved catalytic domain shared by the sirtuins carries out deacetylation and ADPribosylation. In both reactions, sirtuins use b-NAD þ as a cofactor. During the deacetylation reaction (left panel), b-NAD þ is
hydrolysed to generate nicotinamide, 20 -O-acetyl-ADP-ribose and the deacetylated target. During the ADP-ribosylation reaction (right
panel), ADP-ribose is attached to an acetylated target protein and nicotinamide is released. NAD þ , nicotinamide adenine dinucleotide.
its deacetylation. Alternatively, some sirtuins might
have only mono-ADP-ribosyltransferase activity.
Regulation of sirtuins by NAD þ /NADH ratios and
inhibition by nicotinamide
Since NAD þ is a critical cofactor for sirtuin enzymatic
activity, changes in levels of NAD þ and NADH, or
their ratio, induced by diet and metabolic status, may
regulate the biological activity of sirtuins in vivo as
discussed below.
CD38, which localizes to the inner nuclear membrane,
hydrolyses NAD þ to nicotinamide (Aksoy et al.,
2006b). In CD38/ mice, NAD þ levels are increased
10- to 20-fold, and the SIRT1 substrate p53 is less
acetylated (Aksoy et al., 2006a). Nicotinamide phosphoribosyltransferase (Nampt), which is induced after
some forms of stress, converts nicotinamide to nicotinamide mononucleotide, which then reacts with ATP to
regenerate NAD þ (Yang and Sauve, 2006). Overexpression of Nampt increases NAD þ levels and induces
SIRT1 activity, with changes in gene expression
paralleling those in cells overexpressing SIRT1 (Revollo
et al., 2004). Nampt decreases as primary cells age and
undergo replicative senescence, which lowers NAD þ
levels and SIRT1 activity (van der Veer et al., 2007).
Inhibition of Nampt activity induces premature senescence in early-passage primary cells, while overexpression of Nampt delays senescence and increases survival
after oxidative stress in late-passage primary cells (van
der Veer et al., 2007). These effects of Nampt are
dependent on SIRT1, as overexpression of a catalytically inactive SIRT1 blocks lifespan extension and
accelerates senescence (van der Veer et al., 2007).
Another cellular enzyme that uses NAD þ is poly(ADPribose) polymerase (PARP), a nuclear protein involved in
DNA repair that cleaves NAD þ into nicotinamide and
ADP-ribose (Zhang, 2003). Poly(ADP-ribose) is added to
histones and other nuclear proteins including p53 (Poirier
and Savard, 1980; Mendoza-Alvarez and Alvarez-Gonzalez, 2001; Simbulan-Rosenthal et al., 2001). Activation
of PARP after DNA damage quickly depletes cells of
NAD þ with a concomitant increase in nicotinamide
levels, which may suppress the activity of the sirtuins
(Zhang, 2003). Future experiments aimed at elucidating
Sirtuins and cancer
LR Saunders and E Verdin
the consequences of and the interplay among mono-ADP
ribosylation, poly-ADP ribosylation and deacetylation
of the same target protein, like histones and p53,
will reveal how sirtuins influence cell survival after
DNA damage.
Nicotinamide promotes cell survival and longevity by
protecting cells against stress, injury and inflammatory
responses (Li et al., 2006). As a precursor to NAD þ ,
nicotinamide inhibits sirtuins by blocking the regeneration of NAD þ through interception of an ADP-ribosylenzyme-acetyl peptide intermediate (Jackson et al.,
2003; Porcu and Chiarugi, 2005). Nicotinamide extends
the replicative lifespan of primary human fibroblasts
(Lim et al., 2006), and inhibits SIRT1 with an IC50 of
o50 mM and PARP-1 with an IC50B100 mM (Yamamoto and Okamoto, 1980; Bitterman et al., 2002). Since
nicotinamide is found in mammalian cells at 50–150 mM,
nicotinamide is a physiological inhibitor of both sirtuins
and PARP (Yang and Sauve, 2006).
Regulation of SIRT1 expression in primary cells and its
overexpression in cancer
SIRT1 is overexpressed in a number of cancers.
This overexpression occurs in part at the transcriptional
level following the loss of repressors of the SIRT1
promoter. Two p53 binding sites in the SIRT1 promoter
normally repress SIRT1 expression. However, in
the absence of nutrients, Foxo3a translocates to the
nucleus, interacts with p53, inhibits its suppressive
activity and leads to increased SIRT1 expression
(Nemoto et al., 2004). p53/ mice show increased basal
expression of SIRT1 in selective tissues, including
adipose tissue, but SIRT1 levels were not further
elevated upon nutrient withdrawal (Nemoto et al.,
2004). SIRT1 levels may be higher in tumors that have
lost p53. As discussed below, SIRT1 deacetylates both
p53 and FOXO proteins, which feedback to regulate
SIRT1 expression.
Another tumor suppressor that regulates SIRT1 is
E2F1, which binds the SIRT1 promoter and regulates
basal expression and induction after DNA damage
(Wang et al., 2006). E2F1-Rb complexes bind target
promoters and recruit HDAC-containing complexes to
repress transcription of genes that control cell-cycle
progression (DeGregori and Johnson, 2006). E2F1 is
acetylated by p300/CBP-associated factor (PCAF),
which enhances its DNA-binding, transactivation activity, and stability, and transforms E2F1 into an activator
of transcription (Martinez-Balbas et al., 2000). SIRT1
also deacetylates E2F1 and may regulate apoptosis
induction in response to DNA damage through this
factor (Wang et al., 2006). The fine balance and
regulation of acetylases and deacetylases may determine
the cellular response to DNA damage: DNA repair for
low levels of DNA damage and apoptosis for extensive
DNA damage. Loss of E2F1 may have a dual effect on
SIRT expression: increased basal expression and impaired induction after DNA damage. Such dysregulation of SIRT1 expression might tip the cellular response
to DNA damage to favor apoptosis induction instead of
DNA repair (Figure 3).
SIRT1 is also regulated by the tumor suppressor gene
hypermethylated in cancer 1 (HIC1). SIRT1 mRNA and
protein levels are increased in HIC1/ mouse embryonic
fibroblasts (MEFs) (Chen et al., 2005). Similar to the
intimate feedback loop with E2F1, SIRT1 interacts with
HIC1 and represses its own expression (Chen et al.,
2005). SIRT1 also deacetylates HIC1 and promotes
HIC1 sumoylation at the same lysine acetylated by the
histone acetyltransferases (HATs) CBP/p300 (Stankovic-Valentin et al., 2007). HIC1 functions as a transcriptional repressor when deacetylated and sumoylated
(Stankovic-Valentin et al., 2007). HIC1 promoter
hypermethylation occurs during tumorigenesis and
aging, leading to the upregulation of SIRT1 (Chen
et al., 2003, 2004). The normal downregulation of
SIRT1 protein seen during aging might be lost in cells
without HIC1, making them resistant to replicative
senescence after oxidative stress, and vulnerable to
transformation if mutations are propagated. HIC þ /
mice are tumor prone and show a p53- and SIRT1dependent block in apoptosis induction in response to
DNA damage (Chen et al., 2003, 2005). Loss of HIC1
and concomitant increase in SIRT1 prolongs lifespan,
but facilitates tumor development as less apoptosis is
induced in response to DNA damage.
SIRT1 expression is also regulated at the posttranscriptional level by HuR (Abdelmohsen et al.,
2007). HuR is a ubiquitously expressed mRNA-binding
Figure 3 Activation of SIRT1 after stress promotes survival
through deacetylation of various substrates. Oxidative stress or
DNA damage induces activation of HATs and histone deacetylases. The extent of activation of these factors and the balance of
acetylation and deacetylation determines how a cell responds to the
stress signal. SIRT1 expression is induced after stress by multiple
transcription factors, such as p53, E2F1 and hypermethylated in
cancer 1, and its enzymatic activity is dependent on nicotinamide
adenine dinucleotide levels that may be regulated during stress as
well. High SIRT1 activity (left panel) promotes deacetylation of
Ku70, FOXO and p53, promotes DNA repair and cell-cycle arrest,
and blocks apoptosis induction after various stresses. If the levels
and activity of HATs predominate (right panel), acetylated forms
of Ku70, Foxo and p53 induce apoptosis. HAT, histone
acetyltransferases; SIRT, sirtuins.
Sirtuins and cancer
LR Saunders and E Verdin
protein that binds the 30 UTR of SIRT1 to stabilize the
SIRT1 transcript (Abdelmohsen et al., 2007). HuR
decreases dramatically as cells age and reach senescence
(Wang et al., 2001) and this leads to a destabilization of
SIRT1 mRNA (Abdelmohsen et al., 2007). High levels
of SIRT1 and HuR contribute to the enhanced survival
of rapidly proliferating cells after oxidative damage, as
opposed to senescent cells that are more sensitive to
oxidative damage (Abdelmohsen et al., 2007). After
oxidative damage, HuR is phosphorylated by Chk2,
which promotes dissociation of HuR from SIRT1
mRNA, leading to lower levels of SIRT1 and an
enhanced sensitivity to apoptosis induction (Abdelmohsen et al., 2007). Thus, the extent of DNA damage is
relayed to SIRT1, in part, by activation of Chk2. High
levels of DNA damage and increased activation of Chk2
decrease expression of SIRT1, tipping the balance
towards acetylation of p53 and other factors that induce
SIRT1 promotes replicative senescence during prolonged
exposure to low-level stress
Primary cells undergo a limited number of divisions
before telomere shortening sends a stress signal that
induces replicative senescence. Oncogene activation and
other stress stimuli also induce the same replicative
senescence program (Hanahan and Weinberg, 2000).
During replicative senescence in primary human fibroblasts, SIRT1 levels decrease (Michishita et al., 2005;
Abdelmohsen et al., 2007). A decrease in HuR during
senescence destabilizes the SIRT1 mRNA (Abdelmohsen et al., 2007). SIRT1 enzymatic activity also decreases
as NAD þ levels fall due to a decrease in Nampt activity
(van der Veer et al., 2007).
Several tumor suppressor genes, including p53, p19ARF
and Rb, regulate senescence by inducing a permanent
withdrawal from the cell-cycle. The transcription factor
p53 regulates expression of genes, including p21, PUMA
and Bax, to initiate cell-cycle arrest, senescence or
apoptosis (Vousden and Lu, 2002). Loss of p53 function
can compromise the ability of human cells to undergo
replicative senescence (Itahana et al., 2001). The key role
of p53 in regulating cell proliferation and stress response
is highlighted by inactivation or loss-of-function of p53
in over 50% of human tumors (Hollstein et al., 1994). In
response to stress signals, p53 is acetylated at multiple
lysines, which enhances its activity and stability (Barlev
et al., 2001). p53 is deacetylated on lysine 382 (K379 in
mouse p53) both by SIRT1 and HDAC1 (Luo et al.,
2001; Vaziri et al., 2001; Cheng et al., 2003). p53
recruitment of HATs and HDACs/SIRT1 to specific
promoters may regulate replicative senescence via
changes in gene expression (Han et al., 2006).
Increased p53 acetylation is associated with senescence (Pearson et al., 2000). As would be expected if
increased acetylation of p53 induces senescence, SIRT1
overexpression blocks oncogene-induced senescence
through deacetylation of p53 (Langley et al., 2002).
Contrary to the prediction that loss of SIRT1 would
induce senescence through increased p53 acetylation,
primary SIRT1/ MEFs are resistant to replicative
senescence in response to prolonged cell replication in
culture (Chua et al., 2005). However, induction of
replicative senescence is not impaired after acute DNA
damage or oncogene activation in SIRT1/ MEFs
(Chua et al., 2005). Thus, SIRT1 limits replicative
senescence during prolonged exposure to low levels of
stress. In addition to lower levels of total p53 and less
p53 acetylation, SIRT1/ MEFs show lower expression
of the tumor suppressor gene p19ARF (Cheng et al., 2003;
Chua et al., 2005). The increase in p19ARF and p53
expression normally observed during serial passage of
MEFs is not seen in SIRT1/ MEFs, which could
prevent induction of replicative senescence (Chua et al.,
2005). The interplay between SIRT1 and p53 and their
reciprocal regulation of expression may be critical for
replicative senescence. p19ARF does not appear to be
acetylated, and it is unclear how SIRT1 regulates p19ARF
expression, but it appears to play a critical role in how
SIRT1 regulates replicative senescence (Chua et al.,
2005). A role for SIRT1 in limiting replicative lifespan is
also suggested by knockdown of SIRT1 with siRNAs in
human diploid fibroblasts, which leads to enhanced cell
proliferation (Abdelmohsen et al., 2007). Thus, SIRT1
seems to limit replicative lifespan and induce replicative
senescence in response to chronic nonlethal forms of
stress. An in-depth analysis of specific stresses will
further define SIRT1’s role in limiting replicative
Overexpression of SIRT1 alone does not affect
replicative lifespan (Michishita et al., 2005), but Nampt
overexpression, which increases SIRT1 activity through
enhanced production of NAD þ , promotes p53 deacetylation and extends replicative lifespan of human smooth
muscle cells (van der Veer et al., 2007). These results
suggest that increased NAD þ levels regulate SIRT1 and
at least one other factor to promote longevity, or that
increased SIRT1 levels are not sufficient and SIRT1
must be activated to extend replicative lifespan. Alternatively, as NAD þ binds tetrameric p53, alters its
conformation and inhibits DNA binding at some
promoters (McLure et al., 2004), increased NAD þ may
inhibit p53, perhaps at specific promoters, to block
replicative senescence. Thus, the proper equilibrium of
SIRT1 expression levels and activation limits replicative
lifespan, along with the interaction and balance of HATs
and tumor suppressor genes, such as p53 and p19ARF.
As discussed above, cells can respond to DNA
damage either by inducing cell-cycle arrest and
DNA repair or by inducing apoptosis, depending
on the extent of the damage and the ability to repair
the damage before cell division. The balance of NAD þ
and NADH control activation of SIRT1, which may
signal the extent of DNA damage and influence the
cellular response. Proper feedback mechanisms turn
off the response once the damage is repaired. Extended
survival responses signaling through SIRT1 during
chronic stress could lead to tumor development through
inactivation of p53. Therefore, SIRT1 may induce
senescence, promote longevity and block tumor
Sirtuins and cancer
LR Saunders and E Verdin
development when cells are under chronic stress. While
senescence protects against tumor development earlier
in life, senescent cells may contribute to aging and agerelated diseases, such as cancer, by compromising tissue
renewal and repair (Campisi, 2005).
Interestingly, transgenic mice expressing a truncated
form of p53, which has high activity, are resistant to
tumor formation, yet show premature signs of aging and
have a shorter lifespan (Tyner et al., 2002). In addition to
p53, the related proteins p63 and p73 mediate cell growth
and tumorigenesis by regulating target genes similar to
those of p53, and p73 is deacetylated by SIRT1 (Dai
et al., 2007). Mice overexpressing an isoform of p63,
DNp63a, under the keratin 14 promoter for specific
expression in the basal cells of the skin, also age
prematurely (Sommer et al., 2006). In both transgenic
mice with truncated p53 and those expressing DNp63a,
SIRT1 levels are lower (Sommer et al., 2006). As the
DNp63a isoform is overexpressed in the transgenic mice
with high p53 activity and DNp63a interacts with SIRT1,
elevated levels of DNp63a might directly regulate SIRT1
stability or turnover (Sommer et al., 2006). These mouse
models suggest that lower levels of SIRT1 lead to
premature aging, but could also protect against tumorigenesis. Conditional deletion of both p53 and SIRT1
will be needed to determine whether SIRT1 exerts its
activity primarily via p53 deacetylation.
SIRT1 also promotes replicative senescence by deacetylating histone H1 and forming heterochromatin
(Figure 4). Heterochromatin, which is associated with
gene silencing, contains several distinct alterations in
acetylation and methylation, including hypoacetylated
H4-K16 (Jeppesen and Turner, 1993), which can be
carried out by SIRT1 (Vaquero et al., 2004). SIRT1 also
recruits and deacetylates linker histone H1 on K26 to
establish heterochromatin (Vaquero et al., 2004). Aging
changes the distribution of H1 subtypes (Lennox and
Cohen, 1983), and changes in chromatin mediate the
silencing of specific genes during senescence (Narita
et al., 2003). H1 protein is absent from senescent cells,
and its loss induces formation of senescence-associated
heterochromatic foci (SAHFs; Narita et al., 2003;
Funayama et al., 2006). This loss of H1 is thought to
occur post-translationally (Funayama et al., 2006),
indicating that as levels of SIRT1 decrease during
cellular aging, the resulting hyperacetylation of H1
might target it for degradation. Future experiments will
determine if the loss of histone H1 during replicative
senescence is indeed dependent on lower levels of SIRT1.
Deacetylation of SIRT1 targets in response to cellular
stress promotes survival
SIRT1 localizes to the nucleus where it deacetylates
various targets and regulates the cellular response to
stress (Figure 3). SIRT1 promotes cellular survival by
initiating cell-cycle arrest and DNA repair through
deacetylation of p53, FOXO and Ku70. While SIRT1
clearly deacetylates p53, the consequence of this
Figure 4 SIRT1 expression and activity decrease during aging. High levels of SIRT1 protein in young cells in combination with high
levels of nicotinamide adenine dinucleotide lead to deacetylation of p53 and histone proteins to promote longevity (left panel). SIRT1
levels are lower in aged cells, and higher levels of nicotinamide inhibit SIRT1 activity (right panel). The resulting hyperacetylation of
p53 can induce replicative senescence. Deacetylation of histone H1 leads to its degradation that promotes formation of senescenceassociated heterochromatic foci (SAHFs). Failure to downregulate SIRT1 during aging may promote cell survival after oxidative
damage, leading to the accumulation of mutations, and an increased risk of cancer development. SIRT, sirtuins.
Sirtuins and cancer
LR Saunders and E Verdin
modification on p53 after DNA damage is controversial.
In response to DNA damage, SIRT1/ MEFs show
hyperacetylation of p53 at multiple lysines, not just the
SIRT1 target K379, but this is not accompanied by a
further increase in expression of p53 target genes such as
p21, Bax or mdm2, nor by an increase in apoptosis
(Cheng et al., 2003; Kamel et al., 2006). A SIRT1specific inhibitor, EX-527, increases p53 acetylation
without any consequence on cell survival after DNA
damage, unlike TSA, which inhibits HDAC1 and results
in decreased survival following DNA damage (Solomon
et al., 2006). Thus, more than just SIRT1-mediated
deacetylation of p53 may be required to promote
survival after DNA damage.
MEFs from mice engineered to express p53 with seven
C terminal lysines mutated to arginines (p537KR), to
mimic unacetylated p53, respond to DNA damage
similarly to MEFs from wild-type mice, but show
enhanced stability of p537KR that allows for enhanced
induction of target genes in thymocytes after DNA
damage (Krummel et al., 2005). Immortalized p537KR
MEFs undergo senescence earlier, suggesting that
regulation of p53 acetylation may fine-tune p53 activity
over the lifespan of an organism (Krummel et al., 2005).
ES cells from mice engineered to express p53 with six C
terminal lysines mutated to arginines (p536KR) show less
transactivation of targets after DNA damage than ES
cells that are p53 þ / (Feng et al., 2005). MEFs from
these mice show no difference in p53 stability or target
transactivation after DNA damage (Feng et al., 2005).
Thymocytes from p536KR mice are slightly more resistant
to apoptosis after irradiation (IR), with impaired
upregulation of PUMA and DR5, but equivalent
induction of mdm2, Bax and Pidd, and no alteration
in p53 stability (Feng et al., 2005). Since these C
terminal lysines are also modified by ubiquitination,
neddylation, methylation and sumoylation, these studies
suggest a role for acetylation in conjunction with other
post-translational modifications in controlling p53
stability, localization and activity. A specific role for
SIRT1 deacetylation of p53 in thymocytes and other
tissues should be examined through tissue-specific
alterations in SIRT1 levels. Additionally, conditional
double-knockout mice lacking both SIRT1 and HDAC1
will address whether these deacetylases are redundant in
targeting of p53 or if both are needed in response to
certain types of stress and signaling pathways.
FOXO transcription factors, including FOXO1,
FOXO3a, FOXO4 and FOXO6, respond to DNA
damage and oxidative stress and regulate expression of
cell-cycle, DNA repair and apoptosis genes (FurukawaHibi et al., 2005). The importance of FOXO proteins in
cancer is underscored as the FOXO proteins are
important regulators of cell growth and are found as
novel fusion proteins after chromosomal translocations
in several types of cancer. The FOXOs are regulated by
post-translational modifications including phosphorylation and acetylation. After oxidative stress, FOXO
proteins are phosphorylated and relocalize to the
nucleus, where they associate with HATs to form active
transcriptional complexes, but the FOXOs themselves
are acetylated on several lysine residues by multiple
HATs, which inhibits their transactivation activity
(Fukuoka et al., 2003; Daitoku et al., 2004; Motta
et al., 2004; van der Horst et al., 2004).
SIRT1 deacetylation of FOXO suppresses transactivation of proapoptotic proteins Bim and Fas ligand by
FOXO, while promoting expression of p27kip and
GADD45a to induce cell-cycle arrest (Brunet et al.,
2004; van der Horst et al., 2004; Kobayashi et al., 2005).
While overexpression of SIRT1 inhibits apoptosis
induced by FOXO3a after cellular stress and SIRT1/
MEFs are more sensitive to oxidative damage, the
ability of FOXO3a to induce cell-cycle arrest is
enhanced by overexpression of SIRT1 and diminished
in SIRT1/ cells (Brunet et al., 2004; Kobayashi et al.,
2005). SIRT1 and FOXO1 are recruited to the
manganese superoxide dismutase promoter, and the
deacetylase activity of SIRT1 is required for transactivation of this antioxidant gene, indicating that SIRT1
also promotes survival by inducing the repair of
oxidative damage (Daitoku et al., 2004; van der Horst
et al., 2004). Thus, SIRT1 appears to promote cell-cycle
arrest and DNA repair downstream of FOXO proteins,
promoting survival rather than apoptosis. The ability of
SIRT1 to promote survival after oxidative stress agrees
with the model that SIRT1 promotes a longer lifespan,
as increased resistance to oxidative stress correlates with
longevity (Finkel and Holbrook, 2000). While the
FOXO homolog in Caenorhabditis elegans plays a key
role in lifespan regulation, demonstration of a similar
role in mammalian lifespan will require development of
conditional transgenic mouse models.
As FOXO and p53 share many transcriptional targets,
the consequences of stress signals in cells containing
wild-type FOXO and p53, versus in tumor cells containing a mutation in either or both p53 or FOXO, are
predicted to vary. The ability of SIRT1 to promote cellcycle arrest and DNA repair after DNA damage might
require both p53 and FOXO deacetylation. The fact that
p53 and FOXO interact following oxidative stress,
highlight the possibility that SIRT1 mediates survival
following cellular stress through both p53 and FOXO
(Brunet et al., 2004). In primary cells with wild-type p53
and FOXO proteins, SIRT1 will promote cell-cycle
arrest versus apoptosis after DNA damage. However, in
tumor cells lacking wild-type p53 or FOXO, SIRT1 may
have a different effect, and careful studies with various
mutants of p53, FOXO and SIRT1 are needed to
determine the interplay among these factors. CBP/p300
acetylation of FOXO proteins shifts the response
towards apoptosis (van der Heide and Smidt, 2005).
Again, overexpression of SIRT1 in primary cells may
influence the downstream signaling pathways initiated
after DNA damage and determining whether cells with
DNA damage proliferate without arresting for repair,
potentially leading to tumorigenesis. Tumor cells might
require high levels of SIRT1 to protect against apoptosis,
therefore allowing for the continued proliferation of
tumor cells with genomic instability.
Another SIRT1 target that promotes survival in cells
that have suffered DNA damage is Ku70. Ku70 is
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LR Saunders and E Verdin
mainly localized to the nucleus where it is involved in
DNA damage repair, but a small fraction is localized in
the cytoplasm where Ku70 regulates apoptosis through
sequestration of the proapoptotic protein Bax (Sawada
et al., 2003). DNA damage promotes Ku70 acetylation
on multiple lysines in its C terminus, disrupting the
association with Bax, which on release transits to the
mitochondria and initiates apoptosis (Cohen et al.,
2004a). SIRT1, along with a classI/II HDAC, maintains
Ku70 in a deacetylated state (Cohen et al., 2004a, b).
SIRT1, while mainly localized in the nucleus, can shuttle
to the cytoplasm (Tanno et al., 2006), but the site of
SIRT1-mediated deacetylation of Ku70 is not yet clear.
Only nuclear SIRT1 appears to be important for cell
survival in response to DNA damage, as mutants that
are constitutively cytoplasmic do not protect against
oxidative stress (Tanno et al., 2006). Future experiments
will determine if acetylation of Ku70 regulates its DNA
repair functions in the nucleus. For SIRT1 to promote
cell survival in cells with DNA damage, the increased
acetylation of Ku70 after DNA damage must be
promptly reversed by deacetylation of Ku70. The
factors that fine-tune Ku70 acetylation and deacetylation are not known. High levels of SIRT1 in primary
cells may tip the balance towards survival after DNA
damage. As cells age and SIRT1 levels are diminished,
DNA damage should result in increased levels of
apoptosis. In pre-malignant cells overexpressing SIRT1,
Ku70 may not be properly activated in response to
DNA damage that is too extensive to be repaired,
allowing for the accumulation of mutations without the
induction of apoptosis. Treatment of cancers with a
combination of HDAC and sirtuin inhibitors might
promote apoptosis in combination with standard
chemotherapy agents that damage DNA.
The balance between acetylation and deacetylation of
specific factors is regulated at an additional level, since
SIRT1 itself deacetylates several HATs and regulates
their enzymatic activities (Kalkhoven et al., 2002;
Thompson et al., 2004). SIRT1 directly interacts with
PCAF and p300 to promote their deacetylation and
enzymatic inactivation (Fulco et al., 2003; Motta et al.,
2004; Cohen et al., 2004a). The importance of HATs in
cancer is highlighted by their frequent mutation,
deletion and translocation in several types of cancer
(Iyer et al., 2004). Recruitment of SIRT1 to its targets
brings it in close proximity to HATs and allows SIRT1mediated deacetylation of p300, CBP and PCAF. The
balance of HAT and HDAC/SIRT1 levels and their
dysregulation in cancer alter the expression, stability
and localization of target proteins.
The critical role of SIRT1 during development may be coopted for tumor cell survival
SIRT1 clearly plays an important role in development:
knockout mice show multiple defects, and most die
before or just after birth (Cheng et al., 2003; Ford et al.,
2005). SIRT1 and p53 double-knockout mice have the
same phenotype as SIRT1/ mice indicating that p53
hyperacetylation is not responsible for the developmental defects downstream of SIRT1 (Kamel et al., 2006).
Generation of conditional mice lacking both SIRT1 and
FOXO will reveal the contribution of FOXO proteins to
SIRT1’s role in development.
SIRT1 normally limits proliferation and possibly
tumor development as small interfering RNA (siRNA)mediated SIRT1 depletion in primary human fibroblasts
enhances their proliferation (Abdelmohsen et al., 2007).
In the context of cells containing wild-type p53 and
intact cell-cycle checkpoints, SIRT1 limits tumor
formation by inducing cellular senescence. However,
loss of p53 and other tumor suppressor genes increases
SIRT1 expression and can lead to transformation and
formation of tumor cells that are addicted to SIRT1
overexpression. Various tumor cell lines cease growth
and undergo apoptosis after knockdown of SIRT1
expression via siRNA (Ford et al., 2005; Abdelmohsen
et al., 2007). Therefore, SIRT1 appears to be a key
survival factor for some tumor cells (Ford et al., 2005;
Ota et al., 2006).
SIRT1 is overexpressed in acute myeloid leukemia
(AML) blasts from patients as well as AML cell lines
(Bradbury et al., 2005). The overexpression found in
multiple types of nonmelanoma skin cancers, including
early stages, suggests that SIRT1 may be playing a
critical role in promoting proliferation in skin cancer
(Hida et al., 2006). The ability of cancer cells to undergo
senescence correlates with their response to treatment.
Interestingly, SIRT1 mRNA and protein levels are
higher in drug-resistant cancer cell lines than in the
original cell lines and in patient tumors after chemotherapy than before therapy (Chu et al., 2005). siRNAmediated knockdown of SIRT1 expression partially
reverses the drug-resistant phenotype, while SIRT1
overexpression or activation of SIRT1 with resveratrol
increases expression of the multidrug resistance protein,
MDR1 (Chu et al., 2005). Differences in the expression
level of SIRT1 in primary versus transformed cells needs
to be expanded to other cancer types, to identify cancers
which might benefit from treatment with SIRT1-specific
SIRT1-specific inhibitors may be useful chemotherapeutics to target tumors whose survival depends on
SIRT1. Overexpression of SIRT1 in tumors is likely to
affect both histone and non-histone acetylated targets.
An example of a tumor dependent on SIRT1-mediated
deacetylation of a non-histone target is B-cell lymphoma; SIRT1 deacetylates and deactivates the critical
oncogene B-cell lymphoma 6 protein (BCL6). BCL6, a
protooncogene, functions as a transcriptional repressor
and recruits HDACs and other corepressors to unique
promoters (Dhordain et al., 1998). BCL6 is specifically
expressed in mature B cells and is required for the
formation of germinal centers through repression of
genes involved in differentiation and apoptosis (Niu,
2002). Chromosomal translocations involving BCL6 are
found in non-Hodgkin’s lymphoma and BCL6 is
constitutively expressed in some B-cell lymphomas
(Niu, 2002). BCL6 is regulated by several post-translational modifications, including phosphorylation, which
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LR Saunders and E Verdin
targets BCL6 for proteasome-mediated degradation
(Niu et al., 1998), and acetylation by p300, which
inactivates BCL6 repression of targets through disruption of the interaction with HDACs (Bereshchenko
et al., 2002). Like nuclear factor-kB (NF-kB), p53 and
Ku70, BCL6 is deacetylated by both HDACs and
SIRT1 (Bereshchenko et al., 2002; Heltweg et al., 2006).
BCL6 repressor activity is controlled by competing
HATs and HDACs/SIRT1, and acetylation impairs the
oncogeneic properties and transforming capabilities of
BCL6 (Bereshchenko et al., 2002). Cambinol, a SIRT1
inhibitor, inactivates BCL6 in Burkitt’s lymphoma cells
by promoting its acetylation, and leads to apoptosis
induction (Heltweg et al., 2006). In mouse xenograft
models, cambinol alone was effective specifically against
tumors expressing BCL6 (Heltweg et al., 2006). DNA
damage does not promote BCL6 acetylation (Bereshchenko et al., 2002), but inhibition of SIRT1 with
cambinol sensitizes cells to DNA-damage-induced
apoptosis independently of p53 (Heltweg et al., 2006).
Cambinol also induced p53, FOXO3a and Ku70
acetylation (Heltweg et al., 2006), indicating that multiple targets of SIRT1 may control the response to DNA
damage. Tumor cells addicted to SIRT1 may be
sensitized to apoptosis induction by combined use of
SIRT1 and HDAC inhibitors to sensitize them to DNAdamaging chemotherapeutic agents.
In addition to its effects on specific non-histone
targets, SIRT1 may promote cancer development by
deacetylating histones. SIRT1 preferentially deacetylates H3-K9, H3-K14 and H4-K16 in vitro and in vivo
(Imai et al., 2000; Vaquero et al., 2004). Importantly,
loss of H4-K16 acetylation is a hallmark of human
tumors (Fraga et al., 2005), and while this could be due
to a loss of HAT association or function, it could also
reflect overexpression of SIRT1 in tumors. Overexpression of SIRT1, other deacetylases, and/or chromatin
remodeling factors may silence key tumor suppressor
genes as an early event in the transformation process
(Jones and Baylin, 2007). Loss of these tumor suppressor genes can relieve constraints on replicative senescence and allow tumor cells to proliferate without
normal checkpoints in place. Epigenetic changes at the
level of histone acetylation are mitotically inherited,
allowing for improper silencing to be passed on during
cell division.
Long-term silencing of genes in embryonic stem cells
is controlled by polycomb group proteins (PcG;
Schuettengruber et al., 2007). PcG proteins function as
large complexes that regulate growth, development and
differentiation by acting on chromatin to alter gene
expression, and PcGs are overexpressed in several
cancers (Gil et al., 2005). The recent ‘cancer stem cell’
hypothesis implicates genes controlling self-renewal in
pluripotent stem cells with the aberrant survival of
tumor cells that have regained the ability to self-renew
(Reya et al., 2001). In stem cells, SIRT1 directly
associates with Suz12, a member of PcG complexes
and like PcG proteins, SIRT1 is overexpressed in breast
and colon cancers (Kuzmichev et al., 2005). In potential
glioblastoma stem cells expressing the marker CD133,
SIRT1 was overexpressed B5-fold compared to CD133negative cells from tumor samples (Liu et al., 2006),
indicating a potential role for SIRT1 in driving the selfrenewal and resistance to apoptosis characteristic for
these cancer stem cells.
Activation of SIRT1 may prevent age-related cancers
Prostate cancer incidence increases with age more
rapidly than any other type of cancer, indicating that
factors that increase risk of development of prostate
cancer might also be associated with controlling lifespan
(Waterbor and Bueschen, 1995). Development of
prostate cancer depends on the activity of the androgen
receptor (AR), which functions as an oncogene, and
while androgen ablative therapy works initially, most
patients relapse (Shand and Gelmann, 2006; Singh et al.,
2006). Interestingly, the only predictor of tumor
recurrence in patients with low-grade prostate cancer
is the presence of changes in global histone acetylation
and methylation levels (Seligson et al., 2005). Hormones, including dihydrotestosterone, activate the AR
by promoting its acetylation, which enhances its
transactivation activity through increased association
with coactivators (Fu et al., 2000, 2003). SIRT1
deacetylates AR, and SIRT1 overexpression can block
hormonal activation of AR (Fu et al., 2006). Furthermore, overexpression of SIRT1 inhibits the in vitro
growth and proliferation of prostate cancer cells that
express the AR (Fu et al., 2006). This suggests that
resveratrol and other SIRT1 activators might be good
chemopreventative and chemotherapeutic agents for
prostate cancer.
The interaction of SIRT1 with several of its targets,
including the AR, may be regulated by four and a half
LIM-domain protein 2 (FHL2) (Yang et al., 2005). LIM
domain proteins function as adapters and modifiers
mediating protein interactions, and FHL2 interacts with
CBP/p300 and SIRT1 to influence their interaction with
AR (Dawid et al., 1998; Labalette et al., 2004; Yang
et al., 2005). In the normal prostate, FHL2 is localized
to the cytoplasm, but in response to RhoA activation in
prostate cancer, FHL2 relocalizes to the nucleus, and
the amount of nuclear FHL2 correlates with more
aggressive prostate cancer (Muller et al., 2002; Kahl
et al., 2006). FHL2 may cooperate with androgens to
inhibit FOXO1 activity by promoting the interaction of
FOXO1 with the AR (Yang et al., 2005). FHL2 interacts
with SIRT1 to promote interaction with FOXO and
attenuate FOXO-dependent expression of proapoptotic
genes in prostate cancer cells (Yang et al., 2005). As
FHL2 can bridge CBP/p300 and SIRT1 to target
proteins, future studies will need to dissect the precise
role of FHL2 in prostate cancer in influencing acetylation and deacetylation of targets. It is not clear if FHL2
is simply bridging HATs and SIRT1 to FOXO and AR
or playing an active role in determining access of
acetylation and deacetylation factors to targets. As
FHL2 is cytoplasmic in normal prostate cells but
nuclear in prostate cancer, small molecules that disrupt
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LR Saunders and E Verdin
the interaction of SIRT1 and its substrates may promote
apoptosis specifically in tumor cells. Furthermore, a
potential role of FHL2 in regulating SIRT1 in other
cancer types needs to be addressed. Specifically, FHL2
and SIRT1 may play a role in colon cancer as FHL2
influences the acetylation of b-catenin by CBP/p300 and
a nuclear sirtuin could play an opposing role.
Chronic inflammation-induced tumorigenesis may be
prevented by SIRT1 deacetylation of NF-jB
Chronic inflammation is a major risk factor for several
cancers and NF-kB activation plays a key role in
inflammation-associated cancers. Activation of NF-kB
in cancer is associated with resistance to apoptosis and is
a key determinant in chemotherapy response. NF-kB is
maintained in an inactive form in the cytoplasm, and
translocates to the cytoplasm in response to stress,
infection and inflammation. Once in the nucleus, NF-kB
induces transcription of cell-cycle, proliferation, migration and apoptosis genes (Piva et al., 2006). Phosphorylation of NF-kB on Ser276 recruits HAT-containing
complexes to target promoters (Zhong et al., 2002),
while acetylation of NF-kB on multiple lysines affects
both its DNA binding and transactivation activity
(Chen et al., 2002; Kiernan et al., 2003). Several class I
HDACs as well as SIRT1 can deacetylate NF-kB
(Ashburner et al., 2001; Chen et al., 2002; Zhong
et al., 2002; Yeung et al., 2004). A direct interaction
between endogenous SIRT1 and NF-kB not only
promotes the deacetylation of NF-kB on lysine 310,
but also of HATs on target promoters and local histone
proteins to actively repress target gene expression
(Yeung et al., 2004). As chronic inflammation and
activation of NF-kB may contribute to several types of
cancers, treatment with resveratrol may be a potential
chemopreventative or chemotherapeutic agent. Resveratrol inhibits NF-kB-dependent transcription, but it is
not known if this is directly and solely through
activation of SIRT1 or also dependent on resveratrol’s
effects on other regulators of NF-kB (Holmes-McNary
and Baldwin, 2000; Manna et al., 2000).
Regulation of a mitotic checkpoint by SIRT2
During mitosis, phosphorylation stabilizes SIRT2,
which shuttles to the nucleus and colocalizes with
chromatin (Dryden et al., 2003; Inoue et al., 2006). In
the nucleus, SIRT2 deacetylates H4-K16, and during the
G2/M transition, global levels of H4-K16 acetylation
decrease, which may aid in chromatin condensation
(Vaquero et al., 2006). As overexpression of SIRT2
delays mitotic exit, SIRT2 might function as a mitotic
checkpoint protein by preventing chromatin condensation in response to mitotic stress (Dryden et al., 2003;
Inoue et al., 2006). A loss of SIRT2 expression is seen in
gliomas, which might compromise the mitotic checkpoint, contributing to genomic instability and tumorigenesis (Dryden et al., 2003; Hiratsuka et al., 2003;
Inoue et al., 2006). The proposed role of SIRT2 in
regulating a mitotic checkpoint through global deacetylation of H4K-16 suggests that SIRT2 could guard
against cells undergoing division when DNA damage is
SIRT2 also colocalizes with microtubules in the
cytoplasm and deacetylates a-tubulin (North et al.,
2003). The consequences of tubulin acetylation are not
understood, but may affect tubulin stability (Piperno
et al., 1987). As several microtubule poisons are used as
chemotherapeutic agents, SIRT2 expression may contribute to the response to these drugs (Rowinsky and
Calvo, 2006). Tumors with high levels of SIRT2 may
affect their response to chemotherapy drugs that target
tubulin stability.
Mitochondrial sirtuins: SIRT3, SIRT4 and SIRT5
Three sirtuins, SIRT3, SIRT4 and SIRT5 localize to
the mitochondria. A proteomics approach recently
suggested that 20% of mitochondrial proteins are
acetylated, especially those involved in lifespan and
metabolism (Kim et al., 2006). As mitochondria are the
major site for production of reactive oxygen species that
causes oxidative stress, this organelle is key in lifespan
and aging. Mitochondria function in energy metabolism, and acetylation of key metabolic proteins may
regulate changes in energy production in response to
changes in the environment. Acetyl-CoA and NAD þ are
key indicators of cellular energy status, and acetyl-CoA
is a substrate for HATs, while NAD þ is a cofactor for
sirtuins. While the cellular role and deacetylation targets
are just starting to be identified, the mitochondrial
sirtuins will likely play a role in the altered energy
metabolism and response to oxidative stress in tumor
cells. The first targets of SIRT3 and SIRT4 have recently
been identified, and will be discussed in detail below.
SIRT5 has deacetylase activity, but no targets have yet
been identified (North et al., 2003; Michishita et al.,
SIRT3 shows high expression in brown adipose tissue,
where its expression is further increased upon CR and
exposure to cold temperatures (Shi et al., 2005). SIRT3
functions as a protein deacetylase (Onyango et al., 2002;
Schwer et al., 2002), and specifically targets K642 of the
mitochondrial protein acetyl-CoA synthetase 2. Deacetylation of acetyl-CoA synthetase 2 activates the enzyme
and promotes the production of acetyl-CoA (Hallows
et al., 2006; Schwer et al., 2006). Several genetic studies
have linked SIRT3 to aging, including in humans (Rose
et al., 2003; Glatt et al., 2007).
SIRT4 does not possess deacetylase activity, but
functions as an ADP-ribosyltransferase (Michishita
et al., 2005; Haigis et al., 2006). Its only known target
is GDH, a mitochondrial enzyme that converts glutamate to a-ketoglutarate (Haigis et al., 2006). ADPribosylation represses the activity of GDH (Haigis
et al., 2006). In contrast to SIRT1 and SIRT3 that are
upregulated during CR, SIRT4 activity may be downregulated by CR (Haigis et al., 2006). SIRT4 knockout
Sirtuins and cancer
LR Saunders and E Verdin
mice are viable, and pancreatic islets isolated from these
mice secrete higher levels of insulin, revealing a role for
SIRT4 in downregulating insulin secretion through
repression of GDH activity (Haigis et al., 2006). While
additional targets of SIRT4 remain to be identified, the
repression of GDH activity by SIRT4 may protect
against diabetes and aging. The loss of SIRT4 could
contribute to diabetes, a major risk factor for cancer
(Strickler et al., 2001). Future experiments aimed at
evaluating cancer incidence in aging SIRT4 knockout
mice, especially those on a western diet that could
increase diabetes risk, may identify a role for SIRT4 in
protecting against cancer development.
Guarding against aging and genomic instability by SIRT6
SIRT6 is a nuclear protein associated with heterochromatin that functions in base excision repair (BER). BER
serves to protect the genome from single-strand breaks
that arise spontaneously from alkylation, oxidation and
deamination events, or that are induced by chemical
mutagens (Michishita et al., 2005; Mostoslavsky et al.,
2006). SIRT6 is widely expressed with the highest levels
found in the brain and skeletal muscle and has no
detectable deacetylase activity, but displays robust
ADP-ribosyltransferase activity in vitro and can autoADP-ribosylate (North et al., 2003; Liszt et al., 2005;
Michishita et al., 2005). No ADP-ribosylated targets of
SIRT6 have been identified, but histone proteins and
DNA repair proteins are possible targets. It will also be
interesting to determine if SIRT2 and SIRT6 work in
concert in the nucleus, with SIRT6 signaling to SIRT2
when DNA damage is repaired and mitosis can proceed.
While normal human fibroblasts overexpressing
SIRT6 do not have an extended replicative lifespan,
SIRT6/ MEFs and SIRT6/ mouse embryonic stem
cells (mESCs) show a reduced proliferative rate
(Michishita et al., 2005; Mostoslavsky et al., 2006).
SIRT6/ MEFs and mESCs have an increased sensitivity to irradiation but normal UV sensitivity (Mostoslavsky et al., 2006). SIRT6/ cells show multiple
chromosomal defects including fragmentation, detached
centrosomes, gaps and translocations due to a defect in
BER (Mostoslavsky et al., 2006). Reconstitution of
SIRT6/ MEFs with wild-type and catalytically inactive
SIRT6 showed that an intact deacetylase domain of
SIRT6 is required for BER and maintenance of genomic
stability (Mostoslavsky et al., 2006). Proper regulation of
genomic stability protects against tumor formation and
aging (Lombard et al., 2005) and SIRT6/ mice show
premature aging-like symptoms and die several weeks
after birth (Mostoslavsky et al., 2006). Conditional
knockdown of SIRT6 will be needed to determine if
SIRT6 also guards against tumor formation by protecting the genome from DNA damage. SIRT6 may be a
tumor suppressor gene whose loss contributes to the
accumulation of mutations and increased genomic
stability. Future screening of human tumors for loss of
SIRT6 will determine if SIRT6 functions as a tumor
suppressor gene.
SIRT7 regulation of rRNA genes in proliferating tissues
SIRT7 localizes to nucleoli, associates with condensed
chromosomes during mitosis and is widely expressed in
proliferating mouse tissues, such as liver, testes and
spleen (Michishita et al., 2005; Ford et al., 2006). Low
levels of SIRT7 are found in nonproliferating tissues,
such as heart, brain and skeletal muscle (Michishita
et al., 2005). SIRT7 interacts with RNA Polymerase I
and promotes active transcription of rRNA genes, an
activity that requires a wild-type catalytic domain (Ford
et al., 2006). No deacetylase or ADP-ribosyltransferase
activity has been demonstrated for SIRT7, and while
SIRT7 has been reported to copurify with a high
molecular weight complex in human cells, no specific
interacting partners or targets have been identified
(North et al., 2003; Ford et al., 2005, 2006).
SIRT7 was identified in a subtractive library screen as
being overexpressed in human thyroid papillary carcinoma compared to normal thyroid cells (De Nigris
et al., 2002; Frye, 2002). As primary breast cells
(HMECs) reach senescence, expression of SIRT3 and
SIRT7 increases (Ashraf et al., 2006). Additionally,
increased SIRT7 expression was seen in breast cancers
as compared to nonmalignant breast tissue, with a
significantly higher expression of SIRT7 in nodepositive than in node-negative breast cancers (Ashraf
et al., 2006). Knockdown of SIRT7 expression in human
U20S cells induces apoptosis, indicating that SIRT7 is
required for cell survival (Ford et al., 2006). SIRT7
functions to regulate rRNA transcription and therefore
is likely essential for cell survival and proliferation in
both primary and transformed cells. It is not clear if
elevated SIRT7 levels are a cause or consequence of cells
bypassing senescence and becoming malignant, as
increased proliferation rates of tumors require increased
RNA pol-I transcription of rRNA. This potential role of
SIRT7 in driving proliferation of malignant growth is in
line with its higher expression in proliferating versus
nonproliferating tissues (Ford et al., 2006). Tumor cells
may be addicted to SIRT7 to maintain rapid proliferation, but the use of SIRT7 inhibitors as potential cancer
therapeutic agents might be impeded by an essential role
of SIRT7 in primary cells.
Therapeutic potential of sirtuin regulation
As the list of sirtuin targets rapidly expands and the
cellular function of each sirtuin becomes clearer, the
critical role that sirtuins play at the crossroads between
longevity and cancer will be further elucidated. Current
evidence suggests that several sirtuins protect against
cancer. Primary cells express high levels of SIRT1, and
SIRT1 levels normally decrease during cellular aging to
promote replicative senescence and protect against
tumorigenesis (Chua et al., 2005). As discussed above,
replicative senescence is a form of tumor suppression
and the critical role that SIRT1 plays in limiting
replicative lifespan may indicate that SIRT1 functions
as a tumor suppressor.
Sirtuins and cancer
LR Saunders and E Verdin
If SIRT1 indeed functions as a tumor suppressor, the
SIRT1 activator resveratrol may function as a beneficial
chemopreventative agent. Resveratrol is a natural
product induced in plants after a variety of stresses
and shows potential cancer chemoprotective properties
(Athar et al., 2007). Activation of SIRT1 may be a
useful chemopreventative agent in age-related cancers,
such as prostate cancer in which deacetylation of the AR
inactivates its ability to transform prostate cells (Fu
et al., 2006). Another example in which SIRT1
activators, such as resveratrol, might be useful as a
chemopreventative agent would be in promoting deacetylation and inactivation of NF-kB to prevent chronic
inflammation, a contributing factor to tumor development (Yeung et al., 2004). In addition to its ability to
function as a SIRT1 activator, resveratrol also downregulates the anti-apoptotic gene survivin and blocks
activation of the pro-survival activity of NF-kB, PI3K/
AKT and mitogen-activated protein kinase pathways
(Fulda and Debatin, 2006). Future experiments will
clarify whether SIRT1 activation by resveratrol is
upstream of all of these pathways, or if there are
multiple mechanisms by which resveratrol works to
protect against cancer.
While SIRT1 clearly plays a critical role in development, lifespan and protection against cancer development, several forms of cancer have elevated levels of
SIRT1 and may be dependent on SIRT1 for proliferation and survival. Loss of tumor-suppressor genes that
regulate SIRT1 expression may lead to the overexpression of SIRT1 (Nemoto et al., 2004; Chen et al., 2005;
Wang et al., 2006). Therefore, SIRT1-specific inhibitors
may be useful chemotherapeutic agents for cancers
‘addicted’ to SIRT1 expression. Specifically, treatment
of B-cell lymphomas with a SIRT1 inhibitor
would increase acetylation of BCL6 and inactivate it
as an oncogene (Heltweg et al., 2006). In addition to
inactivation of an oncogene, SIRT1 inhibition may also
reactivate expression of proapoptotic genes and
other tumor suppressor genes through increased histone
acetylation. Tumor cells show global changes in their
chromatin, which can in part be due to the dysregulation
of sirtuins (Jones and Baylin, 2007). Overexpression
of SIRT1, SIRT2, SIRT3 and SIRT7 are found in
tumors (De Nigris et al., 2002; Hiratsuka et al., 2003;
Bradbury et al., 2005; Ashraf et al., 2006; Hida et al.,
2006), indicating that sirtuin inhibitors may be useful
chemotherapeutic agents against cancers that overexpress these sirtuins. The increased acetylation of targets,
such as p53, Foxo, Ku70 and BCL6 induced by
inhibition of sirtuins, in conjunction with chemotherapy
agents that damage DNA, might induce tumor cell
Future work in this area will more clearly define the
precise role of each sirtuin at the crossroads between
cancer and longevity and will hopefully identify new
chemopreventative and chemotherapeutic uses of sirtuin
activators and inhibitors.
We thank Stephen Ordway and Gary Howard for editorial
assistance and John Carroll, Alisha Wilson, and Chris Goodfellow for graphics assistance. The authors apologize to all
colleagues whose work could not be included due to space
limitations. Eric Verdin is a Senior Scholar in Aging from the
Ellison Foundation for Medical Research and a member of the
scientific advisory board of Sirtris Pharmaceuticals.
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