Readthrough of nonsense mutation W822X in the SCN5A gene can effectively restore expression of cardiac Na+ channels W822X / 中华心血管病杂志
Chinese Journal of Cardiology
; (12): 238-241, 2011.
Article
Dans Zh
| WPRIM
| ID: wpr-272270
Responsable en Bibliothèque :
WPRO
ABSTRACT
<p><b>OBJECTIVE</b>In this study we investigated the functional restoration of nonsense mutations in the SCN5A gene.</p><p><b>METHODS</b>The readthrough-enhancing reagents were introduced to HEK293 cells to suppress one nonsense mutation W822X in the SCN5A gene. Patch-clamp was used to record the whole-cell current and dynamics. Western blot and immunofluorescence staining were used to certify the expression and the location of the sodium channel.</p><p><b>RESULTS</b>In transfected HEK293 cells, the nonsense mutation in SCN5A inhibited the expression level of full-length protein, and the sodium currents from the mutant channels were less than 3% of the wild-type level. Readthrough enhancement by decreasing translation termination efficiency with a siRNA targeting eukaryotic release factor eRF3a (a GTPase that binds eRF1), the sodium current from the mutant cDNAs was restored to as much as 30% of the wild-type. After the treatment by the readthrough-enhancing reagents, the channels from cDNA carrying W822X remained the features of wild-type phenotype, and Western blot and immunochemical staining also showed the expression of full-length channel proteins.</p><p><b>CONCLUSION</b>Readthrough-enhancing reagents could effectively suppress nonsense mutations in SCN5A and partially restore the function of sodium channel and the expression of full-length channels.</p>
Texte intégral:
1
Indice:
WPRIM
Sujet Principal:
Plasmides
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Transfection
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Canaux sodiques
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Techniques de patch-clamp
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Codon non-sens
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Petit ARN interférent
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Cellules HEK293
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Génétique
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Métabolisme
Limites du sujet:
Humans
langue:
Zh
Texte intégral:
Chinese Journal of Cardiology
Année:
2011
Type:
Article