Skip to main content
. 2018 Mar 13;7:e31023. doi: 10.7554/eLife.31023

Figure 6. MBD3L2 expression de-represses the D4Z4 array.

(A–C) DUX4 and DUX4 target gene expression as determined by RT-qPCR in MB2401 control (A), MB073 FSHD1 (B) or MB200 FSHD2 (C) myoblasts without (-) or with (+) doxycycline (Dox) treatment for 48 hr to induce MBD3L2 transgene expression in clonal cell lines. (D–E) DUX4-positive nuclei upon overexpression of MBD3L2 in MB200 FSHD2 myoblasts as in (C) were detected by immunofluorescence (D) and quantified by counting three fields representing >125 nuclei (E). (F–G) DUX4 and DUX4 target gene expression as determined by RT-qPCR following control (CTRL) or MBD3L family gene shRNA knockdown in MB073 FSHD1 (F) or MB200 FSHD2 (G) myotubes. Error bars denote the standard deviation from the mean of three biological replicates. Statistical significance was calculated by comparing the specific knockdown to the control knockdown for each gene using a two-tailed, two-sample Mann-Whitney U test and p was ≤0.05 for all comparisons except in (A). See also Figure 6—source data 1.

Figure 6—source data 1. Source data for Figure 6.
This file contains the source data used to make the graphs presented in Figure 6 and Figure 6—figure supplements 1–2. GraphPad Prism was utilized to visually represent the quantitative data.
elife-31023-fig6-data1.xlsx (61.6KB, xlsx)
DOI: 10.7554/eLife.31023.038

Figure 6.

Figure 6—figure supplement 1. Validation of MBD3L2 overexpression and depletion.

Figure 6—figure supplement 1.

(A–E) The ectopic (A–C) or endogenous (D–E) expression of MBD3L2 as determined by RT-qPCR in MB2401 control (A), MB073 FSHD1 (B) or MB200 FSHD2 (C) myoblasts cultured without (-) or with (+) doxycycline (Dox) for 48 hr, or in MB073 FSHD1 (D) or MB200 FSHD2 (E) myotubes expressing control (CTRL) or MBD3L gene shRNAs. Error bars denote the standard deviation from the mean of three biological replicates. See also Figure 6—source data 1.
Figure 6—figure supplement 2. Additional MBD3L knockdown experiments.

Figure 6—figure supplement 2.

(A–D) MBD3L2, DUX4, and DUX4 target gene expression as determined by RT-qPCR in two additional independent experiments with control (CTRL) or MBD3L shRNA-expressing MB073 FSHD1 (A–B) or MB200 FSHD2 (C–D) muscle cell lines differentiated into myotubes. Error bars denote the standard deviation from the mean of three biological replicates. See also Figure 6—source data 1.