DNA transposition by protein transduction of the piggyBac transposase from lentiviral Gag precursors. Issue 4 (21st November 2013)

• Record Type: Journal Article
• Authors: Cai, Yujia; Bak, Rasmus O.; Krogh, Louise Bechmann; Staunstrup, Nicklas H.; Moldt, Brian; Corydon, Thomas J.; Schrøder, Lisbeth Dahl; Mikkelsen, Jacob Giehm
• Is Part Of: Nucleic acids research. Volume 42:Issue 4(2014)
• Page Start: e28
• Page End: e28
• Publication Date: 2013-11-21
• Subjects: Nucleic acids -- Periodicals; Molecular biology -- Periodicals; 572.805
• DOI: 10.1093/nar/gkt1163
• Languages: English
• ISSNs: 0305-1048

Abstract

Abstract: DNA transposon-based vectors have emerged as gene vehicles with a wide biomedical and therapeutic potential. So far, genomic insertion of such vectors has relied on the co-delivery of genetic material encoding the gene-inserting transposase protein, raising concerns related to persistent expression, insertional mutagenesis and cytotoxicity. This report describes potent DNA transposition achieved by direct delivery of transposase protein. By adapting integrase-deficient lentiviral particles (LPs) as carriers of the hyperactive piggyBac transposase protein (hyPBase), we demonstrate rates of DNA transposition that are comparable with the efficiency of a conventional plasmid-based strategy. Embedded in the Gag polypeptide, hyPBase is robustly incorporated into LPs and liberated from the viral proteins by the viral protease during particle maturation. We demonstrate lentiviral co-delivery of the transposase protein and vector RNA carrying the transposon sequence, allowing robust DNA transposition in a variety of cell types. Importantly, this novel delivery method facilitates a balanced cellular uptake of hyPBase, as shown by confocal microscopy, and allows high-efficiency production of clones harboring a single transposon insertion. Our findings establish engineered LPs as a new tool for transposase delivery. We believe that protein transduction methods will increase applicability and safety of DNA transposon-based vector technologies.