Functional repair of a mutant chloride channel using a trans-splicing ribozyme

Christopher S. Rogers, Carlos G. Vanoye, Bruce A. Sullenger, Alfred L. George*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

74 Scopus citations


RNA repair has been proposed as a novel gene-based therapeutic strategy. Modified Tetrahymena group I intron ribozymes have been used to mediate trans-splicing of therapeutically relevant RNA transcripts, but the efficiency of the ribozyme-mediated RNA repair process has not been determined precisely and subsequent restoration of protein function has been demonstrated only by indirect means. We engineered a ribozyme that targets the mRNA of a mutant canine skeletal muscle chloride channel (cClC-1) (mutation T268M in ClC-1 causing myotonia congenita) and replaces the mutant-containing 3′ portion by trans-splicing the corresponding 4-kb wild-type sequence. Repair efficiency assessed by quantitative RT-PCR was 1.2% ± 0.1% in a population of treated cells. However, when chloride channel function was examined in single cells, a wide range of electrophysiological activity was observed, with 18% of cells exhibiting significant functional restoration and some cells exhibiting complete rescue of the biophysical phenotype. These results indicate that RNA repair can restore wild-type protein activity and reveal considerable cell-to-cell variability in ribozyme-mediated trans-splicing reaction efficiency.

Original languageEnglish (US)
Pages (from-to)1783-1789
Number of pages7
JournalJournal of Clinical Investigation
Issue number12
StatePublished - Dec 2002

ASJC Scopus subject areas

  • Medicine(all)


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