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. Author manuscript; available in PMC: 2010 Jan 9.
Published in final edited form as: Cell. 2009 Jan 9;136(1):62–74. doi: 10.1016/j.cell.2008.10.052

Figure 4. SIRT6 Deacetylates Histone H3K9 at promoters of RELA target genes.

Figure 4

(A) Western analysis of SIRT6 protein in HeLa cells stably expressing SIRT6 shRNA (sh2)

(B) SIRT6 is required for H3K9 deacetylation at promoters of RELA target genes. ChIP with α-H3K9Ac and α-H3 antibodies was performed. H3K9 acetylation at RELA target gene promoters (mean ± s.e.) is shown relative to untreated control samples and normalized to total H3 levels.

(C) SIRT6 is required to limit RELA occupancy at the promoter of RELA target gene promoters. ChIP with α-RELA antibodies was performed following a 30 minute TNF-α pulse; RELA occupancy (mean ± s.e.) at promoters relative to untreated control samples is shown.

(D)-(E) SIRT6-mediated deacetylation of nucleosomes inhibits nucleosome binding to RELA. Western analysis of H3K9Ac levels on nucleosomes following incubation with SIRT6 in NAD-dependent deacetylation or mock reactions (D). The acetylated or SIRT6-deacetylated nucleosomes were used in nucleosome binding assays with increasing amounts of GST-RELA protein (E). Extent of nucleosome binding can be estimated by comparing levels of bound (**) and unbound (*) nucleosomes. Reduced binding is observed in the SIRT6-deacetylated samples (compare 10 ug RELA samples)