p53 — the guardian of the genome — is mutated in more than half of all human cancers, making it a prime therapeutic target. But p53 activity is regulated by many proteins; could one of these prove to be a better candidate for cancer therapy? In the 19 October issue of Cell, Luo et al. and Vazri et al. identify another protein — Sir2 — that regulates p53's activity, and show that its inhibition potentiates p53's effects.

It has long been known that p53 activity is regulated by phosphorylation, but recent reports indicate that it is also regulated by a different type of modification — acetylation. Acetylation has previously been associated with the regulation of histones — the protein components of chromatin — but the Sir2 proteins are NAD-dependent deacetylases that are conserved in lower organisms that don't contain histones, so could they be involved in deacetylation of p53?

To investigate this, the groups isolated mammalian Sir2 homologues. Both the human SIRT1 (also known as SIR2α) and mouse Sirt1 (also known as Sir2α) interacted with p53 in vitro and in vivo, and purified Sirt1 could deacetylate p53 in vitro. The in vivo deacetylase activity of SIRT was confirmed using an antibody that was specific for acetylated p53. Acetylated p53 could not be detected following transfection of SIRT1.

So why is p53 acetylated? Acetylation seems to be induced in response to cellular stress. Luo et al. showed that p53 acetylation was induced following treatment with the DNA-damaging drug etoposide, and Vaziri et al. showed that there was an increase in p53 acetylation following γ-irradiation. Acetylation could be abolished by expression of SIRT1.

As p53 is acetylated in response to stress, it is logical to assume that this would correlate with an increase in its activity, and that deacetylation by SIRT1 would inhibit p53's activity. The groups therefore tested the ability of p53 to activate transcription using a p53-activated luciferase reporter system. Luo et al. generated a construct with p53-binding sites upstream of the transcription start site, and Vaziri et al. used the promoter of a p53 transcriptional target — the CDKN1A gene, which encodes WAF1 — to drive transcription. p53 increased luciferase activity in a dose-dependent manner, and this activity was suppressed by SIRT1. Similar results were reported in vivo, as WAF1 was induced following exposure to γ-irradiation, and a fourfold overexpression of SIRT1 reduced this induction.

So, SIRT1 inhibits p53's ability to activate transcription, but what about its ability to induce apoptosis? Luo et al. treated p53+/+ cells with either etoposide or hydrogen peroxide (oxidative stress), both of which induce apoptosis. However, expression of Sirt1 makes the cells more resistant to these types of cellular stress, so promoting cell survival.

These results indicate that p53 acetylation enhances its activity, and that this is attenuated by SIRT1. So could inhibition of SIRT1 prove a useful therapeutic approach for restoring p53 activity in cancer cells?

The answer seems to be yes. Luo et al. exploited the knowledge that Sirt1 was dependent on NAD hydrolysis for its activity to identify nicotinamide — a byproduct of this reaction — as an inhibitor of Sirt1. And both groups showed that SIRT1 was insensitive to inhibition by trichostatin A (TSA), which inhibits HDAC1, another deacetylase that acts on p53. Luo et al. showed that the two inhibitors cooperate in the induction of acetylated p53, and suggest that the combination of TSA and nicotinamide might act synergistically in cancer therapy to activate p53.

Vaziri et al. used a different approach to inhibit SIRT1 activity and potentiate that of p53. They constructed a dominant-negative SIRT1 mutant — that contained a histidine to tyrosine substitution at residue 363 and coded for a catalytically inactive protein — and showed that its expression increased the kinetics of accumulation of acetylated p53, the induction of WAF1 and apoptosis following transfection with p53.

So, with the characterization of SIRT1, we have discovered a new potential target for cancer therapy. Let's hope that this latest discovery will soon be capitalized on.