A MUSCLE-SPECIFIC ENHANCER WITHIN INTRON-1 OF THE HUMAN DYSTROPHIN GENE IS FUNCTIONALLY DEPENDENT ON SINGLE MEF-1 E BOX AND MEF-2/AT-RICH SEQUENCE MOTIFS/
Hj. Klamut et al., A MUSCLE-SPECIFIC ENHANCER WITHIN INTRON-1 OF THE HUMAN DYSTROPHIN GENE IS FUNCTIONALLY DEPENDENT ON SINGLE MEF-1 E BOX AND MEF-2/AT-RICH SEQUENCE MOTIFS/, Nucleic acids research, 25(8), 1997, pp. 1618-1625
In previous studies we have described a 5.0 kb HindIII fragment downst
ream of muscle exon 1 that exhibits properties consistent with a muscl
e-specific transcriptional enhancer. The goal of this study has been t
o identify the sequence elements responsible for muscle-specific enhan
cer activity. Functional studies indicated that this enhancer is activ
e in pre- and pest-differentiated H962(2-1) myoblasts but functions po
orly in L6 and C2C12 myotubes. The core enhancer region was delimited
to a 195 bp Spel-Accl fragment and sequence analysis identified three
MEF-1/E box and two MEF-2/AT-rich motifs as potential muscle-specific
regulatory domains. EMSA competition and DNase footprinting indicated
that sequences within a 30 bp region containing single adjoining MEF-1
/E box and MEF-2/AT-rich motifs are target binding sites for transacti
ng factors expressed in H9C2(2-1) myotubes but not in L6 or C2C12 myot
ubes. Site-specific mutations within these motifs resulted in a signif
icant reduction in enhancer activity in H9C2(2-1) myotubes. These resu
lts suggest that the mechanisms governing DMD gene expression in muscl
e are similar to those identified in other muscle-specific genes. Howe
ver, the myogenic profile of enhancer activity and transacting factor
binding suggests a more specialized role for this enhancer that is con
sistent with its potential involvement in dystrophin gene regulation i
n cardiac muscle.