Résumé :
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Communication n° 264 Objective : Congenital myasthenic syndromes (CMS) are caused by various genetic defects of presynaptic, synaptic and postsynaptic proteins. We report a postsynaptic CMS caused by two compound heterozygous mutations of the CHRNE gene: a functional null mutation (911delT) and a novel intronic base alteration (IVS5-16G>A) in intron 5 remote from the splice acceptor site. Methods : We analyzed CHRNE transcripts for aberrant splicing in vivo and in vitro in order to delineate the potential pathogenicity of the IVS5-16G>A transition. Results : CHRNE transcripts from the patient?s muscle tissue revealed aberrant splicing of CHRNE RNA. To prove that solely IVS5-16G>A is responsible for the effects seen on RNA level, we constructed wild-type and mutated CHRNE minigenes for transfection in HEK 293 cells and subsequent RNA analysis. In the mutated construct, an exclusively used novel splice acceptor site is generated that results in two aberrant transcript variants: (i) a frameshift variant due to retention of 14 nucleotides of intron 5 in the mature mRNA; (ii) a preferentially transcribed variant using a cryptic splice donor site in exon 5 in combination with the newly created acceptor site in intron 5. In the latter transcript, the reading frame is maintained, and therefore this mRNA escapes nonsense mediated decay. Conclusions : Our results provide strong evidence that the mutation IVS5-16G>A is indeed responsible for the aberrant splicing observed in the patient?s muscle. Our data confirm that the RNA analysis may be necessary to reveal unexpected splicing aberrations due to intronic mutations that are not part of the consensus splice site.
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