In vivo Trans-splicing of 5′ and 3′ segments of pre-mRNA directed by corresponding DNA sequences delivered by gene transfer

Robert G. Pergolizzi, Alexander E. Ropper, Rachel Dragos, Alicia C. Reid, Katsutoshi Nakayama, Yadi Tan, John R. Ehteshami, Struhan H. Coleman, Randi B. Silver, Neil R. Hackett, André Menez, Ronald Crystal

Research output: Contribution to journalArticle

25 Citations (Scopus)

Abstract

We have developed a new paradigm of in vivo gene transfer termed "segmental trans-splicing" (STS), in which individual "donor" and "acceptor" DNA sequences, delivered in vitro or in vivo, generate pre-mRNAs with 5′ and 3′ splice signals, respectively, and complementary hybridization domains through which the two pre-mRNAs interact, facilitating trans-splicing of the two mRNA fragments. To demonstrate STS, we used α-cobratoxin, a neurotoxin that binds irreversibly to postsynaptic nicotinic acetylcholine receptors. Cells or animals receiving both donor and acceptor plasmids, but neither plasmid alone, yielded RT-PCR products with the correct sequence of mature α-cobratoxin mRNA, suggesting that trans-splicing had occurred. Mice receiving intravenous administration of ≥7.5 μg donor + acceptor plasmids, but not either plasmid alone, died within 6 h. These data demonstrate that segmental trans-splicing occurs in vivo. This approach should permit the intracellular assembly of molecules hitherto too large to be accommodated within current gene transfer vectors.

Original languageEnglish
Pages (from-to)999-1008
Number of pages10
JournalMolecular Therapy
Volume8
Issue number6
DOIs
Publication statusPublished - 1 Jan 2003
Externally publishedYes

    Fingerprint

ASJC Scopus subject areas

  • Molecular Medicine
  • Molecular Biology
  • Genetics
  • Pharmacology
  • Drug Discovery

Cite this

Pergolizzi, R. G., Ropper, A. E., Dragos, R., Reid, A. C., Nakayama, K., Tan, Y., Ehteshami, J. R., Coleman, S. H., Silver, R. B., Hackett, N. R., Menez, A., & Crystal, R. (2003). In vivo Trans-splicing of 5′ and 3′ segments of pre-mRNA directed by corresponding DNA sequences delivered by gene transfer. Molecular Therapy, 8(6), 999-1008. https://doi.org/10.1016/j.ymthe.2003.08.022