Darko Bosnakovski1,2, Lynn M. Hartweck1, Abhijit Dandapat1, John Day3, Ramiro Nandez1, Radbod Darabi4, Rita R. Perlingeiro4, Lauren Snider5, Stephen J. Tapscott5, Janet Sowden6, Rabi Tawil6 and Michael Kyba1
1Lillehei Heart Institute and Department of Pediatrics, University of Minnesota, Minneapolis, MN 55455
2Faculty of Technology and Technical Science‐Veles, University St. Kliment Ohridski‐Bitola, Skopje, R. Macedonia
3Paul and Sheila Wellstone Center for Muscular Dystrophy and Department of Neurology, University of Minnesota, Minneapolis, MN 55455
4Lillehei Heart Institute and Department of Medicine, University of Minnesota, Minneapolis, MN 55455
5Division of Human Biology, Fred Hutchinson Cancer Research Center, Seattle, WA 98109
6Fields Center for FSHD and Neuromuscular Research and Department of Neurology, University of Rochester, Rochester, NY 14642
There is currently no animal model bearing the actual FSHD mutation (D4Z4 contraction), and the lack of a suitable model system to study the effects of this mutation has severely hampered progress in understanding FSHD. We present data from two approaches to model FSHD: the generation of a mouse bearing a doxycycline‐inducible Dux4 gene, and the differentiation of iPS cells derived from myoblasts cultures established from FSHD and control biopsies. We have inserted a construct starting at the MAL initiation of Dux4 and continuing to the end of the EcoRI site, therefore containing the pLAM polyA signal from the FSHD A haplotype, into constitutively open chromatin 5’ of the HPRT gene. Even without induction we observed that males are almost never born and carrier females present several pathologies, although not in muscle. Rare males are severely affected, and do not survive past about 1 month of age. In addition to the phenotypes observed in the females, males also present testis abnormalities, including a defect in spermatogenesis. We are currently investigating vascular pathologies, which are especially evident in embryos, and severely enhanced when doxycycline is applied to induce expression of the transgene. Although muscle is not dystrophic, these are the first animals presenting a phenotype due to the presence of human D4Z4 DNA. In a second strategy, we have derived iPS cells from myoblast cultures taken from FSHD patients and controls. We present preliminary data on the ability of these cells to differentiate into muscle in vitro, and on the chromatin state and transcription of DUX4/D4Z4 at different stages of development
