Figure 3.

Targeting dSaCas9-repressors to DUX4 has no effect on Myosin heavy chain 1, D4Z4 proximal genes, or closest-match off-target genes expressed in skeletal muscle

(A–D) Expression levels of the terminal muscle differentiation marker Myosin heavy chain 1 (MYH1) and D4Z4 proximal genes FRG1 and FRG2 were assessed by qRT-PCR in the FSHD myocyte cultures described in Figure 2. (E and F) Expression levels of (E) Lysosomal amino acid transporter 1 homolog (LAAT1), (F) Ribosome biogenesis regulatory protein homolog (RRS1), or Guanine nucleotide-binding protein G(i) subunit alpha-1 isoform 1 (GNAI1) were assessed by qRT-PCR in the relevant FSHD myocyte cultures described in Figure 2. Intron 1 of LAAT1 contains a potential off-target match to sgRNA #1. The single exon of RRS1 and the downstream flanking sequence of GNAI1 contain potential off-target matches to sgRNA #5. Data are plotted as the mean + SD value of at least four independent experiments, with relative mRNA expression for cells expressing each dCas9-epigenetic regulator alone set to 1.