SIRT6 destabilizes MAPK pathway by deacetylating Histone H3 lysine9 and ELK1 targeting in MM cells. (A) Real-time PCR analysis of MEK/ERK-signaling–related genes in wild-type (WT) and SIRT6-silenced MM cells. *P = .02, ** 0.0062 < P < .0018, ***0.007 < P < .005 (Student t test). (B) ChIP-seq binding density for H3K27ac and SIRT6 at the promoter of indicated genes in K562 cells (blue) and human H1-ES cells (green). (C-D) ChIP analyses to detect SIRT6 binding (C) and H3K9 acetylation (D) at the promoter of indicated genes, performed with SIRT6 or AcH3K9-specific antibody and immunoglobulin G (IgG) control, in SIRT6-WT and SIRT6-KD MM.1S cells. The binding of SIRT6 and AcH3K9 at promoters is shown relative to background with IgG control antibody. Data are presented as mean values ± SD of triplicates. Student t test was applied to calculate the P value: ns, not significant; *P = .001; ** 0.0006 < P < .0004; *** P = .0005. (E) Radar chart displays fold-change values of the TF activation profile in SIRT6-KD and SIRT6-WT MM.1S cells. TFIID was used as loading control. (F) ChIP quantitative polymerase chain reaction analysis of control or ELK1-KD MM1S cells. ELK1 immunoprecipitates (IPs) were measured relative to control (IgG) (mean values ± SD of triplicate experiments). GAPDH, which is bound by neither SIRT6 nor ELK1, was a negative control. *P = .04; ** 0.007 < P < .001. (G) SIRT6 binding to ELK1 and ERK2, shown by western blots of GFP-tagged SIRT6 or control IgG antibody IPs from MM.1S cells. C, cytoplasmic fraction; N, nuclear fraction; Tot, total. (H) SIRT6 occupancy at promoters of target genes in ELK1-KD versus ELK1-WT MM.1S cells determined by ChIP (mean values ± SD of triplicates). *P = .04, **.007 < P < .002 (Student t test).