Fig. 4

Activation of the enhancer by CRISPR-dCas9-VPR induces GCKR expression. HepG2 cells were co-transfected with the VPR activator plasmid and the guide RNA plasmids targeting (gR1 or gR2) or not targeting (gEm) the enhancer locus. a Total GCKR mRNA levels determined by qPCR with a GCKR Taqman gene expression assay. The data represent GCKR mRNA levels relative to non-targeting VPR + gEm, normalized to housekeeping gene RPLP0. Error bars represent standard deviation of four independent experiments (n = 4) with three, four, four, and three technical replicates, respectively. b Haplotype-specific enrichment of H3K27Ac (enrichment over input normalized to a region of GRB10 with no TF binding or active histone marks [rs6943153]) determined by ChIP-qPCR using the custom Taqman SNP Genotyping Assay for rs780094 (three independent experiments with three, two, and three technical replicates, respectively). Error bars represent the standard deviation between technical replicates (n = 8). Asterisks and the hash symbol depict statistical significance (***p ≤ 0.005; #p ≤ 0.5 and refers to comparison between haplotypes upon VPR + gEm; a one-way ANOVA with Tukey’s post-hoc test; b two-tailed t-test for comparisons between two groups)