Inhibition of SIRT1 Reactivates Silenced Cancer Genes without Loss of Promoter DNA Hypermethylation

Abstract

The class III histone deactylase (HDAC), SIRT1, has cancer relevance because it regulates lifespan in multiple organisms, down-regulates p53 function through deacetylation, and is linked to polycomb gene silencing in Drosophila. However, it has not been reported to mediate heterochromatin formation or heritable silencing for endogenous mammalian genes. Herein, we show that SIRT1 localizes to promoters of several aberrantly silenced tumor suppressor genes (TSGs) in which 5′ CpG islands are densely hypermethylated, but not to these same promoters in cell lines in which the promoters are not hypermethylated and the genes are expressed. Heretofore, only type I and II HDACs, through deactylation of lysines 9 and 14 of histone H3 (H3-K9 and H3-K14, respectively), had been tied to the above TSG silencing. However, inhibition of these enzymes alone fails to re-activate the genes unless DNA methylation is first inhibited. In contrast, inhibition of SIRT1 by pharmacologic, dominant negative, and siRNA (small interfering RNA)–mediated inhibition in breast and colon cancer cells causes increased H4-K16 and H3-K9 acetylation at endogenous promoters and gene re-expression despite full retention of promoter DNA hypermethylation. Furthermore, SIRT1 inhibition affects key phenotypic aspects of cancer cells. We thus have identified a new component of epigenetic TSG silencing that may potentially link some epigenetic changes associated with aging with those found in cancer, and provide new directions for therapeutically targeting these important genes for re-expression.

NAD+-dependent deacetylation of H4 lysine 16 by class III HDACs

“…Interestingly, these two changes occurred only in the promoter region where SirT1 was localized. This observation agrees with many studies demonstrating that changes in histone acetylation levels that were associated with transcriptional regulation were often located exclusively in regulatory regions (CalestagneMorelli and Ausio´, 2006;Pruitt et al, 2006). Additionally, the finding that SirT1 also deacetylates H1K26Ac (Vaquero et al, 2004) might indicate that H1 recruitment and its deacetylation are coordinated and concomitant with heterochromatin formation.…”

“…SirT1 is present in cancer cells at the promoters of tumor suppressor genes, and its loss results in a hyperacetylation of H4K16 and H3K9 at these promoters; this in turn induces expression of these genes without affecting DNA hypermethylation (Pruitt et al, 2006). These observations confirm previous findings demonstrating that SirT1 targets H4K16Ac (Vaquero et al, 2004).…”

“…SirT1 is overexpressed in late stages of prostate cancer (Kuzmichev et al, 2005;Figure 4b) and in breast and colon cancers (Ashraf et al, 2006;Pruitt et al, 2006). SirT1 can deacetylate the tumor-suppressor p53 and this inhibits both p53 transcriptional activity and p53-dependent apoptosis upon cellular stress.…”

Opposing effects of hMOF and SIRT1 on H4K16 acetylation and the sensitivity to the topoisomerase II inhibitor etoposide

“…Whereas class I HDACs show little substrate specificity and the direct HDAC activity of class II HDACs has been debated, the class III HDAC SIRT1 has been reported to show preference for deacetylation of H4K16 (Vaquero et al, 2004(Vaquero et al, , 2007Pruitt et al, 2006). Further, HDACs 1-8 were ubiquitously expressed in H157 cells as assessed by immunoblotting, and did not show any change in expression after VPA or TSA exposure (data not shown).…”

De la régulation du génome à la progression tumorale

“…D'autre part, ces HAT avaient déjà été identifiées dans des réarrangements chromosomiques associés à des leucémies présentant une activité HAT K16 réduite [28]. À l'inverse de cette perte globale d'acétylation de K16, il a été montré que la ré-expression, au cours du processus tumoral, de gènes épigénétiquement réprimés coïncidait avec une augmentation de l'acétylation de K16 dans leurs régions promotrices bien que les marques épigénétiques de répression ne soient pas nécessairement éliminées [30,31]. C'est pourquoi la détection de variations globales d'acétylation de K16 serait un des marqueurs de développement de certains cancers et constituerait même un moyen de prédire l'évolution clinique du cancer (par exemple, le cancer de la prostate [29]).…”