The Silent Information Regulator-2 gene (Sir2) encodes an NAD-dependent histone deacetylase that links regulation of chromatin, genomic stability, and life span in S. cerevisiae. By promoting chromatin silencing, Sir2 inhibits transcription at several genetic loci and represses recombination at ribosomal DNA (rDNA) repeats. Yeast with mutations in Sir2 have increased genomic instability in the context of rDNA recombination, which in turn shortens replicative life span – a marker of reproductive aging in this organism. Conversely, extra copies of Sir2 that suppress rDNA recombination increase replicative life span. These effects of Sir2 suggest paradigms in which genes that promote genome stabilization through chromatin modulation may be important contributors to regulation of organismal life span, aging, and age-related pathology.

Consistent with a conserved role for Sir2 factors in life span regulation, increased activity of Sir2 proteins in the multi-cellular organisms C. elegans and D. melanogaster also increases life span. However, these Sir2 factors may operate through mechanisms that are independent of genome stabilization, and their physiologic molecular substrates are still unclear. In mammals, there are seven Sir2 family members, SIRT1-SIRT7. The SIRTs have been of great interest as candidate regulators of mammalian life span and aging-related processes. In this context, several mammalian SIRTs have functions that impact on aging-associated molecular pathways and disease. However, initial studies of mammalian SIRTs linked these enzymes to biochemical targets and cellular functions that are distinct from those of S. cerevisiae Sir2. For example, mammalian SIRT1 was first reported to deacetylate the p53 tumor suppressor protein; only later was SIRT1 shown to have a physiologic role in histone deacetylation, chromatin regulation, and most recently, genome stabilization. Other mammalian SIRTs (SIRT2-SIRT5) are reported to have cytoplasmic or mitochondrial substrates (though recent work suggests that sub-cellular shuttling might allow these enzymes to target histones as well). In addition, several studies had not detected histone deacetylase activity for the other nuclear SIRT proteins, SIRT6 and SIRT7. Thus, until recently, the extent to which the functional link of yeast Sir2 to chromatin and genome maintenance is evolutionarily conserved in mammals has been unclear.

The generation of mice deficient for the mammalian SIRT6 gene revealed a potential role for SIRT6 in linking regulation of life span, chromatin, and genomic stability. In this context, SIRT6 deficiency in mice leads to dramatically shortened life span and acute degenerative phenotypes that overlap with pathologies of premature aging. Moreover, SIRT6 knockout mouse cells have genomic instability and DNA damage hypersensitivity. In biochemical fractionation assays, SIRT6 protein associated preferentially with a chromatin-enriched cellular fraction. Together, these observations suggested that SIRT6 might couple chromatin regulation with DNA repair.However, a physiologic role for SIRT6 in such a process was not directly demonstrated.

We recently discovered a molecular function for SIRT6 at chromatin. We showed that SIRT6 deacetylates a specific histone residue, lysine 9 of histone H3 (H3K9), in the context of chromatin at telomeres. SIRT6 thereby stabilizes the association with telomeres of the protein WRN, a DNA processing factor that is mutated in the human progeria Werner Syndrome. In this context, depletion of SIRT6 in human cells leads to telomere dysfunction and genomic instability with end-to-end chromosomal fusions. We also identified a second physiologic context for SIRT6 function as a histone H3K9 deacetylase. Specifically, SIRT6 is recruited to the promoters of genes that have been activated by the NF-κB transcription factor, deacetylates H3K9 at these promoters to attenuate gene expression, and thereby limits NF-κB signaling. Notably, hyperactive NF-κB signaling contributes significantly to the degenerative phenotypes and early death of SIRT6-deficient mice, because in an NF-κB-haploinsufficient genetic background, SIRT6-deficient mice have milder phenotypes and live much longer than mice with SIRT6-deficiency alone. Thus, chromatin regulation by SIRT6 is important for proper telomere function and regulation of gene expression programs, and both these mechanisms of action may impact on genomic stability and aging.

In a third study, we have further expanded the known functions of SIRT6 and showed that SIRT6 is required for efficient DNA DSB repair in the context of chromatin. Biochemical analyses show that SIRT6 associates dynamically with chromatin in response to DNA damage, and stabilizes the DNA DSB repair factor, DNA-dependent protein kinase (DNA-PK), at DSBs. We suggest that the modulation of DSB repair by SIRT6 in response to chronic DNA damage over life may contribute to the effects of SIRT6 on physiologic and pathologic processes in mammalian aging.