FLExDUX4 mice were created using a cre-dependent one-way genetic switch (FLEx) system. Homozygotes mice carrying this DUX4 conditional allele are viable and fertile. Two sets of incompatible outward facing recombination sites (loxP and lox511) flank an inverted human double homeobox 4 (DUX4) sequence, including exons 1-3 and both introns. The DUX4 gene encodes several alternative mRNA splicing variants. The hereditary muscle disorder, facioscapulohumeral muscular dystrophy (FSHD) is caused by the toxic gain-of-function of the DUX4-full-length (DUX4-fl) mRNA isoform. The DUX4-fl mRNA, which encodes a paired homeobox domain transcription factor, is typically not expressed in healthy muscle. However, in FSHD, the rare expression of DUX4-fl (in less than 1% of muscle fibers) initiates a pathogenic cascade of events including apoptosis, differentiation defects, muscle atrophy, and susceptibility to oxidative stress. Overall, FSHD is characterized by a slowly progressing muscular dystrophy that predominantly affects the skeletal muscles of the face, scapula, and upper arms but can affect muscles of the abdomen, hip girdle, and lower legs with ~20% of patients ultimately losing ambulation.
The DUX4 promoter drives expression of two short non-pathogenic isoforms (DUX4-s) and a longer cytotoxic isoform (DUX4-fl). This strain contains 4 point mutation in the 5’ splicing donor sites for the two DUX4-s mRNAs, abolishing expression of the short isoforms and only generating the pathogenic DUX4-fl mRNA isoform.
Because this construct was targeted for insertion into the Gt(ROSA)26Sor locus, DUX4-fl expression is determined by which tissue(s) express Cre recombinase. When bred to mice that express Cre recombinase, the resulting offspring will have the loxP or lox511 sites recombined, resulting in the inversion of the human DUX4-fl sequence, ending in a sense orientation.
Hemizygous and homozygous mice have low level DUX4-fl expression in the absence of Cre Recombinase. These mice exhibit alopecia, soft stool, inflammation, and muscle weakness. Homozygous are more affected, as are males compared to females.
When bred to STOCK Tg(ACTA1-cre/Esr1*)2Kesr/J mice (Stock No. 025750), resulting offspring express slightly more DUX4-fl than the FLExDUX4 hemizygous mice prior to tamoxifen induction (TMX). One TMX injection of 5 mg/kg leads to a useful mildly progressive FSHD model, while 2 injections of 10mg/kg leads to a more severe model.
Of note, the donating investigator reports that when bred to Sox2-Cre mice (Stock No. 008454), resulting offspring are embryonic lethal. When bred to ACTA1-rtTA, tetO-cre mice (Stock No. 012433), resulting offspring are embryonic lethal. When bred to ACTA1-Cre mice (Stock No. 006149), resulting offspring are still-born. When bred to UBC-CreERT2 mice (Stock No. 008085), without tamoxifen induction, offspring begin to show a severe neurological phenotype around 5 weeks of age, perhaps due to leaky DUX4 expression in the brain.
