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Comparative Genome-Wide Profiling of Transposon and Retrovirus Integration Sites in Primary Human CD4+ T Cells

ABSTRACT: We mapped Sleeping Beauty (SB) and piggyBac (PB) insertions in primary human CD4+ T cells and compared their insertion profiles with those of the MLV retrovirus and the HIV lentivirus with respect to proximity to genes, TSSs, CpG islands, DNase I hypersensitive sites, repetitive elements, chromatin marks and transcriptional status of genes.

ORGANISM(S): Homo sapiens

PROVIDER: GSE58744 | GEO | 2015/07/01

SECONDARY ACCESSION(S): PRJNA253316

REPOSITORIES: GEO

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Comparative Genome-Wide Profiling of Transposon and Retrovirus Integration Sites in Primary Human CD4+ T Cells

Project description:We mapped Sleeping Beauty (SB) and piggyBac (PB) insertions in primary human CD4+ T cells and compared their insertion profiles with those of the MLV retrovirus and the HIV lentivirus with respect to proximity to genes, TSSs, CpG islands, DNase I hypersensitive sites, repetitive elements, chromatin marks and transcriptional status of genes. Examination of two transposon systems in one cell type.

2015-07-01 | E-GEOD-58744 | biostudies-arrayexpress

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Project description:DNA transposon-based gene delivery vectors represent a promising new branch of randomly integrating vector development for gene therapy. For the side-by-side evaluation of the piggyBac and Sleeping Beauty systems - the only DNA transposons currently employed in clinical trials - during therapeutic intervention, we treated the mouse model of Tyrosinemia type I. with liver-targeted gene delivery using both transposon vectors. For genome-wide mapping of transposon insertion sites we developed a new Next Generation Sequencing procedure called Streptavidin-Based Enrichment Sequencing, which allowed us to identify approximately 1 million integration sites for both systems. We revealed that a high proportion of piggyBac integrations are clustered in hot regions and found that they are frequently recurring at the same genomic positions among treated animals, indicating that the genome-wide distribution of Sleeping Beauty-generated integrations is closer to random. We also revealed that the piggyBac transposase protein exhibits prolonged activity, which predicts the risk of oncogenesis by generating chromosomal double-strand breaks. Safety concerns associated with prolonged transpositional activity draw attention to the importance of squeezing the active state of the transposase enzymes into a narrower time window.

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Sleeping Beauty transposon mutagenesis in mouse intestinal organoids identifies genes involved in tumor progression and metastasis [Atxn1]

Project description:To identify genes important for colorectal cancer (CRC) development and metastasis, we established a new metastatic mouse organoid model using Sleeping Beauty (SB) transposon mutagenesis. Intestinal organoids derived from mice carrying actively mobilizing SB transposons, an activating KrasG12D, and an inactivating ApcΔ716 allele, were transplanted to immunodeficient mice. Analysis of SB insertion sites in tumors identified numerous candidate cancer genes (CCGs) identified previously in intestinal SB screens performed in vivo, in addition to new CCGs, such as Atxn1. To provide functional validation, we knocked out Atxn1 in mouse tumor organoids carrying an activating KrasG12D, and an inactivating ApcΔ716 allele, and transplanted to mice. Tumor development was promoted when these gene were knocked out, demonstrating that these are potent tumor suppressors.

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Sleeping Beauty Expression data from mouse

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Sleeping Beauty Expression data from mouse

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Sleeping Beauty transposon mutagenesis identified genes involved in colorectal cancer development in the inflammatory microenvironment.

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