Journal Article

Characterization of a Double Homeodomain Protein (DUX1) Encoded by a cDNA Homologous to 3.3 Kb Dispersed Repeated Elements

Hao Ding, Marie-Claire Beckers, Stéphane Plaisance, Peter Marynen, Désiré Collen and Alexandra Belayew

in Human Molecular Genetics

Volume 7, issue 11, pages 1681-1694
Published in print October 1998 | ISSN: 0964-6906
Published online October 1998 | e-ISSN: 1460-2083 | DOI:
Characterization of a Double Homeodomain Protein (DUX1) Encoded by a cDNA Homologous to 3.3 Kb Dispersed Repeated Elements

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Target genes for the helicase-like transcription factor (HLTF), a member of the SNF/SWI family, were immuno-precipitated from HeLa chromatin fragments with an anti-HLTF antibody. A 182 bp fragment (HEFT1) presented 87% sequence identity with 3.3 kb dispersed repeats from the 4q35 D4Z4 locus linked to facioscapulohumeral muscular dystrophy (FSHD). The HEFT1 loci were, however, not genetically linked to FSHD. Transfection and in vitro binding studies identified within HEFT1 a promoter whose basal activity required a GC box activated by Sp1 or Sp3. A 4.4 kb homologous transcript was found mostly in human skeletal muscle and heart. A 1.2 kb cDNA fragment was cloned that encoded a 170 amino acid protein (DUX1) with two paired-type homeodomains. In vitro translated DUX1 specifically interacted in electrophoretic mobility shift assay (EMSA) with a P5 oligonucleotide (5′-GATCTGAGTCTAATTGAGAATTACTGTAC-3′).

DUX1 co-expression activated up to 5-fold transient expression in insect cells of a minimal promoter-luciferase construct fused to P5. The presence of 20 kDa DUX1 in vivo in rhabdomyosarcoma TE671 cell extracts was shown by western blotting with a rabbit antiserum raised against a DUX1 peptide. This antiserum suppressed a TE671 protein-P5 complex in EMSA with identical migration as the in vitro translated DUX1-P5 complex. Genomic PCR experiments could not identify a gene fragment linking the HEFT1 and DUX1 sequences, which present one mismatch in their overlapping region. However, a similar gene was found in another 3.3 kb element comprising the HEFT1 promoter and a DUX1-like open reading frame. In addition, homologous gene sequences were identified in 3.3 kb elements of the D4Z4/FSHD locus, considered until now ‘junk’ DNA.

Journal Article.  10453 words.  Illustrated.

Subjects: Genetics and Genomics

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