Spliceosome-mediated RNA trans-splicing as a tool for gene therapy

M. Puttaraju, Sharon F. Jamison, S. Gary Mansfield, Mariano A. Garcia-Blanco, Lloyd G. Mitchell

Research output: Contribution to journalArticlepeer-review

163 Scopus citations


We have developed RNA molecules capable of effecting spliceosome- mediated RNA trans-splicing reactions with a target messenger RNA precursor (pre-mRNA). Targeted trans-splicing was demonstrated in a HeLa nuclear extract, cultured human cells, and H1299 human lung cancer tumors in athymic mice. Trans-splicing between a cancer-associated pre-mRNA encoding the β- subunit of human chorionic gonadotropin gene 6 and pre-trans-splicing molecule (PTM) RNA was accurate both in vitro and in vivo. Comparison of targeted versus nontargeted trans-splicing revealed a moderate level of specificity, which was improved by the addition of an internal inverted repeat encompassing the PTM splice site. Competition between cis- and trans- splicing demonstrated that cis-splicing can be inhibited by trans-splicing. RNA repair in a splicing model of a nonfunctional lacZ transcript was effected in cells by a PTM, which restored significant β-galactosidase activity. These observations suggest that spliceosome-mediated RNA trans- splicing may represent a general approach for reprogramming the sequence of targeted transcripts, providing a novel approach to gene therapy.

Original languageEnglish (US)
Pages (from-to)246-252
Number of pages7
JournalNature Biotechnology
Issue number3
StatePublished - 1999
Externally publishedYes


  • Gene therapy
  • RNA repair
  • Spliceosome
  • Trans-splicing

ASJC Scopus subject areas

  • Microbiology


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