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The function of the class III histone deacetylase, Sir2, in promoting lifespan extension is well established in small model organisms. By analogy, SirT1, the mammalian orthologue of Sir2, is a candidate gene to slow down aging and forestall the onset of age-associated diseases. We have used SirT1-null mice to study the function of SirT1 in susceptibility to tumorigenesis. The number of intestinal polyps induced in mice carrying the Apc(min) mutation was unaffected by the SirT1 genotype although the average polyp size was slightly smaller in the SirT1-null animals. Similarly, the presence or absence of SirT1 had no effect on incidence and tumor load of skin papillomas induced by the classical two-stage carcinogenesis protocol. We found that resveratrol topically applied to the skin profoundly reduced tumorigenesis. This chemoprotective effect was significantly reduced but not ablated in SirT1-null mice, suggesting that part of the protection afforded by resveratrol requires the SirT1-encoded protein. Thus, our results suggest that SirT1 does not behave like a classical tumor-suppressor gene but the antitumor activity of resveratrol is mediated at least in part by SirT1.

The sirt1 gene encodes a protein deacetylase with a broad spectrum of reported substrates. Mice carrying null alleles for sirt1 are viable on outbred genetic backgrounds so we have examined them in detail to identify the biological processes that are dependent on SIRT1. Sera from adult sirt1-null mice contain antibodies that react with nuclear antigens and immune complexes become deposited in the livers and kidneys of these animals. Some of the sirt1-null animals develop a disease resembling diabetes insipidus when they approach 2 years of age although the relationship to the autoimmunity remains unclear. We interpret these observations as consistent with a role for SIRT1 in sustaining normal immune function and in this way delaying the onset of autoimmune disease.

The yeast sir2 gene and its orthologues in Drosophila and C. elegans have well-established roles in lifespan determination and response to caloric restriction. We have studied mice carrying two null alleles for SirT1, the mammalian orthologue of sir2, and found that these animals inefficiently utilize ingested food. These mice are hypermetabolic, contain inefficient liver mitochondria, and have elevated rates of lipid oxidation. When challenged with a 40% reduction in caloric intake, normal mice maintained their metabolic rate and increased their physical activity while the metabolic rate of SirT1-null mice dropped and their activity did not increase. Moreover, CR did not extend lifespan of SirT1-null mice. Thus, SirT1 is an important regulator of energy metabolism and, like its orthologues from simpler eukaryotes, the SirT1 protein appears to be required for a normal response to caloric restriction.

The SirT1 gene encodes a protein deacetylase that acts on a number of nuclear substrates. p53 was identified as a SirT1 substrate whose transcriptional activity was reported to be negatively regulated by SirT1-dependent deacetylation. We set out to determine whether developmental defects and perinatal lethality observed in SirT1-null mice were caused by p53 hyperactivity by creating mice deficient for both SirT1 and p53. Animals null for both proteins were smaller than normal at birth, had eyelid opening defects and died during the late prenatal and early postnatal periods, a phenotype indistinguishable from mice deficient for SirT1 alone. Upon re-examination of the role of SirT1 in modulating p53 activity, we found that while SirT1 interacts with p53, the SirT1 protein had little effect on p53-dependent transcription of transfected or endogenous genes and did not affect the sensitivity of thymocytes and splenocytes to radiation-induced apoptosis. These findings suggest that SirT1 does not affect many p53-mediated biological activities despite the fact that acetylated p53 has been shown to be a substrate for SirT1.

The lifespan of the nematode, Caenorhabditis elegans, can be extended by mutations affecting components of the insulin-like growth factor (IGF) signaling cascade or by overexpression of SIR2, an NAD+-dependent protein deacetylase. The mammalian homologue of SIR2, Sirt1, has been shown to modulate the activity of FoxO, a transcription factor that is downstream of the IGF signaling system. These results suggest that Sirt1 ought to affect the IGF pathway. We report here evidence that this is the case in mice. The loss of Sirt1 protein in mice results in increased expression of the IGF binding protein IGFBP1, a secreted modulator of IGF function. A number of the anatomical characteristics of Sirt1-null mice closely resemble those of transgenic mice overexpressing IGFBP1. Our data suggest that Sirt1 is part of a regulatory loop that limits the production of IGFBP1 thereby modulating IGF signaling.

The yeast sir2 gene plays a central role in mediating gene silencing and DNA repair in this organism. The mouse sir2alpha gene is closely related to its yeast homologue and encodes a nuclear protein expressed at particularly high levels in embryonic stem (ES) cells. We used homologous recombination to create ES cells null for sir2alpha and found that these cells did not have elevated levels of acetylated histones and did not ectopically express silent genes. Unlike yeast sir2 mutants, our sir2alpha null ES cells had normal sensitivity to insults such as ionizing radiation and heat shock, and they were able to silence invading retroviruses normally. These sir2alpha null cells were able to differentiate in culture normally. Our results failed to provide evidence that the mammalian SIR2alpha protein plays a role in gene silencing and suggest that the physiological substrate(s) for the SIR2alpha deacetylase may be nuclear proteins other than histones.

The yeast Sir2p protein has an essential role in maintaining telomeric and mating type genes in their transcriptionally inactive state. Mammalian cells have a very large proportion of their genome inactive and also contain seven genes that have regions of homology with the yeast sir2 gene. One of these mammalian genes, sir2alpha, is the presumptive mammalian homologue of the yeast sir2 gene. We set out to determine if sir2alpha plays a role in mammalian gene silencing by creating a strain of mice carrying a null allele of sir2alpha. Animals carrying two null alleles of sir2alpha were smaller than normal at birth, and most died during the early postnatal period. In an outbred background, the sir2alpha null animals often survived to adulthood, but both sexes were sterile. We found no evidence for failure of gene silencing in sir2alpha null animals, suggesting that either SIR2alpha has a different role in mammals than it does in Saccharomyces cerevisiae or that its role in gene silencing in confined to a small subset of mammalian genes. The phenotype of the sir2alpha null animals suggests that the SIR2alpha protein is essential for normal embryogenesis and for normal reproduction in both sexes.