Figure 2.

CRISPR-mediated transcriptional activation increases FMRP protein abundance at normal CGG repeat sizes. (A) Immunocytochemistry of HEK293T cells transfected with eGFP or dCas9-VP192 and gRNAs, as indicated. Single channel and merged images are shown with Cas9/GFP (green), FMRP (red), and DAPI (blue). White arrowheads indicate transfected cells. Scale bar represents 20 μm. (B) Western blots showing triplicate samples of HEK293T cells transfected with control plasmid (eGFP) or with dCas9-VP192 and indicated gRNAs and immunoblotted for FMRP, Cas9 and Tubulin. (C) Quantification of western blots from HEK293T cells transfected as indicated. Data are shown as FMRP normalized to tubulin and relative to the control plasmid (n = 6/group evaluated over at least two independent experiments. **p < 0.01, Kruskal-Wallis one-way ANOVA with Dunn’s multiple comparison test). Error bars represent SEM.