Transgenic mice expressing tunable levels of DUX4 develop characteristic facioscapulohumeral muscular dystrophy-like pathophysiology ranging in severity

Takako I. Jones; Guo-Liang Chew; Pamela Barraza-Flores; Spencer Schreier; Monique Ramirez; Ryan D. Wuebbles; Dean J. Burkin; Robert K. Bradley; Peter L. Jones

DUX4-FL protein was not detectable in heart or skeletal muscles from the non-recombined In order to quantitate the changes in gene expression for each severity model, qRT-PCR was used to measure overall DUX4-fl mRNA levels ( Figure 3A ). However, we have previously shown that this assay is a poor measure of DUX4-fl transgene expression using FLExDUX4 mouse models [41] , and DUX4-fl mRNA is even difficult to detect in muscle biopsies from FSHD affected subjects [16] . Since DUX4-FL functions as a transcriptional activator in both human and mouse cells [30, 68] , expression of DUX4-FL direct target genes has proven to be a more accurate indicator of DUX4-FL expression levels in both species [24, 31, 41] . Therefore, . CC-BY-NC 4.0 International license is made available under a The copyright holder for this preprint (which was not peer-reviewed) is the author/funder. It . https://doi.org/10.1101/471094 doi: bioRxiv preprint in addition to DUX4-fl mRNA, the mRNA levels of two mouse homologs of DUX4-FL direct target genes, Wfdc3 and Trim36 [41, 68] , were also assayed ( Figure 3B and C). Detectable DUX4-fl mRNA levels were extremely low in gastrocnemius muscles from all models ( Figure 3A ), consistent with previous studies [41] . Interestingly, there were no significant changes detected in DUX4-fl mRNA levels between the mild, moderate, and severe models 9 days after TMX treatments, a timepoint with prominent differences in DUX4-FL protein expression ( Figure 2 ). In contrast, both DUX4-FL target genes assayed showed significant induction in all bitransgenic animals compared with the FLExD/+ mice, indicating the presence of DUX4-FL protein. Wdfc3 and Trim36 mRNA levels are each significantly increased in muscles from the moderate and severe models compared with the mild model, and Trim36 mRNA levels are significantly increased in the severe model compared to the moderate model ( Figure 3B and C).