title: Comparative genomic integration profiling of Sleeping Beauty transposons mobilized with high efficacy from integrase-defective lentiviral vectors in primary human cells.
abstract: It has been previously shown that integrase-defective HIV-1-based gene vectors can serve, with moderate efficiency, as substrate for DNA transposition by a transiently expressed Sleeping Beauty (SB) transposase. Here, we describe the enhanced gene transfer properties of a HIV-1/SB hybrid vector that allows efficient DNA transposition, facilitated by the hyperactive SB100X transposase, from vector DNA intermediates in primary human cells. Potent transposase-dependent integration of genetic cargo carried by the hybrid HIV-1/SB vector (up to 160-fold above background) is reported in human cell lines as well as in primary human fibroblasts and keratinocytes. The efficiency of transgene integration in context of the newly developed hybrid vector is comparable with that of conventional lentiviral vectors (LVs). Integration profiles of integrating HIV-1-derived vectors and SB transposons mobilized from LVs are investigated by deep sequencing of a large number of integration sites. A significant bias of lentiviral integrations in genes is reported, confirming that biological properties of the viral integration machinery facilitate preferred insertion into actively transcribed genomic regions. In sharp contrast, lentiviral insertions catalyzed by the SB100X transposase are far more random with respect to genes. Based on these properties, HIV-1/SB vectors may become valuable tools for genetic engineering and therapeutic gene transfer.
authors.affiliation: Department of Human Genetics, University of Aarhus, Aarhus, Denmark.
authors.forename: Brian
authors.initials: B
authors.lastname: Moldt
authors.forename: Csaba
authors.initials: C
authors.lastname: Miskey
authors.forename: Nicklas Heine
authors.initials: NH
authors.lastname: Staunstrup
authors.forename: Andreas
authors.initials: A
authors.lastname: Gogol-Döring
authors.forename: Rasmus O
authors.initials: RO
authors.lastname: Bak
authors.forename: Nynne
authors.initials: N
authors.lastname: Sharma
authors.forename: Lajos
authors.initials: L
authors.lastname: Mátés
authors.forename: Zsuzsanna
authors.initials: Z
authors.lastname: Izsvák
authors.forename: Wei
authors.initials: W
authors.lastname: Chen
authors.forename: Zoltán
authors.initials: Z
authors.lastname: Ivics
authors.forename: Jacob Giehm
authors.initials: JG
authors.lastname: Mikkelsen
mesh_terms: D001483:Base Sequence; D002460:Cell Line; D004247:DNA / Q000235:genetics / Q000378:metabolism; D004251:DNA Transposable Elements; D005347:Fibroblasts; D018014:Gene Transfer Techniques; D005822:Genetic Vectors / Q000235:genetics*; D016678:Genome; D015497:HIV-1 / Q000235:genetics*; D059014:High-Throughput Nucleotide Sequencing; D006801:Humans; D019426:Integrases / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D015603:Keratinocytes; D017422:Sequence Analysis, DNA; D019076:Transgenes; D019895:Transposases / Q000235:genetics* / Q000378:metabolism*; D016662:Virus Integration / Q000235:genetics*
chemical_list: D004251:DNA Transposable Elements; D004247:DNA; D019426:Integrases; D019895:Transposases; C494471:sleeping beauty transposase, human