Progress Update: Base editing of DUX4 somatic polyadenylation signal

Progress update by Darina Šikrová

The erroneous production of DUX4 in skeletal muscle is considered the molecular cause for FSHD. In our study, we want to explore if we could reduce the production rate of DUX4 in muscle by modifying its polyadenylation signal – a mark that signals the cell where the DUX4 production should stop. We set out to use a novel genome editing tool to do so. One of the goals was to get “clean” populations of muscle cells derived from FSHD-affected individuals with a modified DUX4 polyadenylation signal and check if it indeed leads to reduced DUX4 production. Another goal was to try optimize current version of this genome editing tool in a way that it would be suitable for preclinical in vivo studies.

After confirming the feasibility of editing our locus of interest in a model cell line, we subjected muscle cells derived from one FSHD1 and one FSHD2 individual to undergo the editing procedure and derived clonal populations with different modifications of the DUX4 polyadenylation signal. After this we compared the amount of DUX4 in successfully edited clones to clones whose polyadenylation signal was unperturbed. In general, we could detect lower levels of DUX4 in edited cells and moreover, we found that such mutagenesis of the DUX4 polyadenylation signal indeed leads to its masking, e.g. it’s not recognized/used anymore. However, we are experiencing DUX4 variability in the clones, which we would like to address by increasing the number of clones and also instead of obtaining pure edited clones try to carry out editing procedure in the whole cell population.

We also started to work on the second goal of our project and to this date we managed to create two new versions of the editor. Currently we are testing them in the model cell line if they perform in intended way as the original version of the editor.

See grant Base editing of DUX4 somatic polyadenylation signal