Abstract
Retroviral vectors integrate in genes and regulatory elements and may cause transcriptional deregulation of gene expression in target cells. Integration into transcribed genes also has the potential to deregulate gene expression at the posttranscriptional level by interfering with splicing and polyadenylation of primary transcripts. To examine the impact of retroviral vector integration on transcript splicing, we transduced primary human cells or cultured cells with HIV-derived vectors carrying a reporter gene or a human β-globin gene under the control of a reduced-size locus-control region (LCR). Cells were randomly cloned and integration sites were determined in individual clones. We identified aberrantly spliced, chimeric transcripts in more than half of the targeted genes in all cell types. Chimeric transcripts were generated through the use of constitutive and cryptic splice sites in the HIV 5′ long terminal repeat and gag gene as well as in the β-globin gene and LCR. Compared with constitutively spliced transcripts, most aberrant transcripts accumulated at a low level, at least in part as a consequence of nonsense-mediated mRNA degradation. A limited set of cryptic splice sites caused the majority of aberrant splicing events, providing a strategy for recoding lentiviral vector backbones and transgenes to reduce their potential posttranscriptional genotoxicity.
| Original language | English |
|---|---|
| Pages (from-to) | 1653-1666 |
| Number of pages | 14 |
| Journal | Journal of Clinical Investigation |
| Volume | 122 |
| Issue number | 5 |
| DOIs | |
| Publication status | Published - May 1 2012 |
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ASJC Scopus subject areas
- Medicine(all)
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Lentiviral vector integration in the human genome induces alternative splicing and generates aberrant transcripts. / Moiani, Arianna; Paleari, Ylenia; Sartori, Daniela; Mezzadra, Riccardo; Miccio, Annarita; Cattoglio, Claudia; Cocchiarella, Fabienne; Lidonnici, Maria Rosa; Ferrari, Giuliana; Mavilio, Fulvio.
In: Journal of Clinical Investigation, Vol. 122, No. 5, 01.05.2012, p. 1653-1666.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Lentiviral vector integration in the human genome induces alternative splicing and generates aberrant transcripts
AU - Moiani, Arianna
AU - Paleari, Ylenia
AU - Sartori, Daniela
AU - Mezzadra, Riccardo
AU - Miccio, Annarita
AU - Cattoglio, Claudia
AU - Cocchiarella, Fabienne
AU - Lidonnici, Maria Rosa
AU - Ferrari, Giuliana
AU - Mavilio, Fulvio
PY - 2012/5/1
Y1 - 2012/5/1
N2 - Retroviral vectors integrate in genes and regulatory elements and may cause transcriptional deregulation of gene expression in target cells. Integration into transcribed genes also has the potential to deregulate gene expression at the posttranscriptional level by interfering with splicing and polyadenylation of primary transcripts. To examine the impact of retroviral vector integration on transcript splicing, we transduced primary human cells or cultured cells with HIV-derived vectors carrying a reporter gene or a human β-globin gene under the control of a reduced-size locus-control region (LCR). Cells were randomly cloned and integration sites were determined in individual clones. We identified aberrantly spliced, chimeric transcripts in more than half of the targeted genes in all cell types. Chimeric transcripts were generated through the use of constitutive and cryptic splice sites in the HIV 5′ long terminal repeat and gag gene as well as in the β-globin gene and LCR. Compared with constitutively spliced transcripts, most aberrant transcripts accumulated at a low level, at least in part as a consequence of nonsense-mediated mRNA degradation. A limited set of cryptic splice sites caused the majority of aberrant splicing events, providing a strategy for recoding lentiviral vector backbones and transgenes to reduce their potential posttranscriptional genotoxicity.
AB - Retroviral vectors integrate in genes and regulatory elements and may cause transcriptional deregulation of gene expression in target cells. Integration into transcribed genes also has the potential to deregulate gene expression at the posttranscriptional level by interfering with splicing and polyadenylation of primary transcripts. To examine the impact of retroviral vector integration on transcript splicing, we transduced primary human cells or cultured cells with HIV-derived vectors carrying a reporter gene or a human β-globin gene under the control of a reduced-size locus-control region (LCR). Cells were randomly cloned and integration sites were determined in individual clones. We identified aberrantly spliced, chimeric transcripts in more than half of the targeted genes in all cell types. Chimeric transcripts were generated through the use of constitutive and cryptic splice sites in the HIV 5′ long terminal repeat and gag gene as well as in the β-globin gene and LCR. Compared with constitutively spliced transcripts, most aberrant transcripts accumulated at a low level, at least in part as a consequence of nonsense-mediated mRNA degradation. A limited set of cryptic splice sites caused the majority of aberrant splicing events, providing a strategy for recoding lentiviral vector backbones and transgenes to reduce their potential posttranscriptional genotoxicity.
UR - http://www.scopus.com/inward/record.url?scp=84860594261&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=84860594261&partnerID=8YFLogxK
U2 - 10.1172/JCI61852
DO - 10.1172/JCI61852
M3 - Article
C2 - 22523069
AN - SCOPUS:84860594261
VL - 122
SP - 1653
EP - 1666
JO - Journal of Clinical Investigation
JF - Journal of Clinical Investigation
SN - 0021-9738
IS - 5
ER -