Abstract
Myogenic determination factors are basic helix-loop-helix proteins that govern specification and differentiation of muscle cells, and bind to the E-box consensus sequence CANNTG in promoter regions of muscle-specific genes. No E-box mutation has been reported to date. RAPSN encodes rapsyn, a 43 kDa postsynaptic peripheral membrane protein that clusters the nicotinic acetylcholine receptor at the motor endplate. Transcriptional regulation mechanisms of RAPSN have not been studied. We here report two novel E-box mutations in the RAPSN promoter region in eight congenital myasthenic syndrome patients. Patient 1 carries -27C→G that changes an E-box at -27 to -22 from CAGCTG to GAGCTG. An allele harboring -27C→G is not transcribed in patient's muscle. Patients 2-8 are of Oriental Jewish stock of Iraqi or Iranian origin with facial malformations, and harbor -38A→G that changes another E-box at -40 to -35 from CAACTG to CAGCTG, which does not affect the consensus CANNTG sequence. Haplotype analysis shows that -38A→G arises from a common founder. For each mutation, position +1 represents the major transcriptional start site that we determine to be 172 nucleotides upstream of the translational start site. Electrophoretic mobility shift assays reveal that -38A→G gains, and -27C→G looses, binding affinity for different components of nuclear extracts of C2C12 myotubes. Luciferase reporter assays show that both -38A→G and -27C→G attenuate reporter gene expression in C2C12 myotubes, and that -27C→G additionally attenuates reporter gene expression in MyoD- or myogenin-transfected HEK cells. The -27C→G mutation also markedly attenuates the enhancer activity of an E-box on an SV40 promoter. Impaired transcriptional activities of the RAPSN promoter region predict reduced rapsyn expression and endplate acetylcholine receptor deficiency.
Original language | English (US) |
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Pages (from-to) | 739-748 |
Number of pages | 10 |
Journal | Human molecular genetics |
Volume | 12 |
Issue number | 7 |
DOIs | |
State | Published - Apr 1 2003 |
ASJC Scopus subject areas
- Molecular Biology
- Genetics
- Genetics(clinical)