THE DUX4 GENE AT THE FSHD LOCUS ON 4Q35 ENCODES A PRO-APOPTOTIC PROTEIN

MARCOWYCZ A; KOWALJOW V; CONDE CB; ARIAS CI; COPPÉ F; BELAYEW A; ROSA AL

Nantes, Francia

Congreso; Myologie 2005; 2005

Association Francaise contre les Myopathies

Facioscapulohumeral muscular dystrophy (FSHD) is the third most common inherited myopathy after Duchenne muscular dystrophy and myotonic dystrophy. The disease is associated with partial deletions of a tandem repeat array of the D4Z4 element on chromosome 4q35. D4Z4 contains a candidate for a causative FSHD gene named double homeobox 4 (DUX4). A DUX4-related protein with the expected MW and charge has been detected in extracts of FSHD but not control primary myoblasts. We report here the impact of transient expression of DUX4 in established muscle cell lines in culture. Upon transfection of a pCIneo-DUX4 expression vector, DUX4 altered plasma membrane permeability as judged from dramatic leakage of cytoplasmic LDH into the culture medium. Caspase 3/7 activities were increased 4-fold after 48h, as compared to cells transfected with either pCINeo-DUX1 or the insertless vector, suggesting the induction of apoptosis. Cell death was also observed upon co-transfection of pcDNA3-DUX4 and GFP expression vectors, reaching 90% after 48 hours. Double immunofluorescence staining and confocal laser-scanning microscopy using specific anti-DUX4 and anti-emerin antibodies were used to study the cellular distribution of DUX4 in transfected cells. The protein localized to the cellular nucleus and altered the nuclear envelope distribution of emerin. An endogenously expressed DUX4-related protein(s) was detected with these antisera in the nuclei of cultured human adult and fetal rhabdomyosarcoma cells. A DUX4-related protein(s) was also detected in human muscle biopsy sections of control and FSHD patients. Muscle tissue localization suggested that these antigens were present in structures compatible with satellite cells. We propose that DUX4-mediated cell death is linked to the pathogenesis of FSHD. We acknowledge funding from the ABMM (Belgium), AFM (France), CONICET and FONCYT (Argentina), the Frenzel (Germany) and Fischer-Shaw (USA) families, The FSH Society and the MDA (USA). A.M. held a FRIA fellowship (Belgium).