Fig. 3

Effect of curcumin on DNMT3B and role of DNMT3B in PKM splicing. (a-b) Methyltransferase inhibition activity of curcumin using an in-vitro methyltransferase-assay kit, with (a) nuclear-extracts of the HNC cells, (b) purified DNMT3B enzyme and (c-e) RPS16 normalized qRT-PCR in shDNMT3B transfected cells versus shcontrol using the indicated primers for (c) DNMT3B and (d-e) PKM gene. (f) Western blot showing the protein level of DNMT3B, PKM2, and PKM1 in shDNMT3B transfected cells versus shControl in H157 cells, GAPDH act as a loading control. (g) MeDIP in shDNMT3B transfected cells versus shcontrol in H157 cells and qRT-PCR relative to input and control IgG. (h-i) ChIP in H157 cells transfected with shDNMT3B versus shcontrol using (h) BORIS and (i) RNA Pol II antibody, followed by qRT-PCR relative to input. Three independent experiments were conducted with mean values ± SD. P value using two-tailed Student’s t-test, * P < 0.05, ** P < 0.01, *** P < 0.001, ns = non-significant