Transcription Start Regions in the Human Genome Are Favored Targets for MLV Integration

Wu, Xiaolin; Li, Yuan; Crise, Bruce; Burgess, Shawn M.

doi:10.1126/science.1083413

Cited by 1,216 publications

(1,236 citation statements)

References 17 publications

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“…In contrast to the genomic integration profile of retroviral systems and other transposon systems ( piggyBac and Tol2 ), which show integration preferences for actively transcribed genes,51, 52, 53 SB -based integration is random 37, 54. Compared to computationally generated control datasets, SB100X -mediated genomic integration of the PEDF gene delivered by the pFAR4 vector into primary human RPE cells showed a close-to-random profile.…”

Section: Discussionmentioning

confidence: 83%

Engineering of PEDF-Expressing Primary Pigment Epithelial Cells by the SB Transposon System Delivered by pFAR4 Plasmids

Thumann

Harmening

Prat-Souteyrand

et al. 2017

Molecular Therapy - Nucleic Acids

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Neovascular age-related macular degeneration (nvAMD) is characterized by choroidal blood vessels growing into the subretinal space, leading to retinal pigment epithelial (RPE) cell degeneration and vision loss. Vessel growth results from an imbalance of pro-angiogenic (e.g., vascular endothelial growth factor [VEGF]) and anti-angiogenic factors (e.g., pigment epithelium-derived factor [PEDF]). Current treatment using intravitreal injections of anti-VEGF antibodies improves vision in about 30% of patients but may be accompanied by side effects and non-compliance. To avoid the difficulties posed by frequent intravitreal injections, we have proposed the transplantation of pigment epithelial cells modified to overexpress human PEDF. Stable transgene integration and expression is ensured by the hyperactive Sleeping Beauty transposon system delivered by pFAR4 miniplasmids, which have a backbone free of antibiotic resistance markers. We demonstrated efficient expression of the PEDF gene and an optimized PEDF cDNA sequence in as few as 5 × 103 primary cells. At 3 weeks post-transfection, PEDF secretion was significantly elevated and long-term follow-up indicated a more stable secretion by cells transfected with the optimized PEDF transgene. Analysis of transgene insertion sites in human RPE cells showed an almost random genomic distribution. The results represent an important contribution toward a clinical trial aiming at a non-viral gene therapy of nvAMD.

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Section: Discussionmentioning

confidence: 83%

Engineering of PEDF-Expressing Primary Pigment Epithelial Cells by the SB Transposon System Delivered by pFAR4 Plasmids

Thumann

Harmening

Prat-Souteyrand

et al. 2017

Molecular Therapy - Nucleic Acids

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“…In a large series of HPV-associated tumors, viral integration sites were found preferentially distributed in transcribed cellular sequences (Klimov et al, 2002;Wentzensen et al, 2002;Ziegert et al, 2003). Extensive analyses of retrovirus integration sites have shown that active genes (Schroder et al, 2002) or transcription start regions (Wu et al, 2003) are preferential integration targets. Fragile sites have also been reported to facilitate HPV DNA integration (Thorland et al, 2003), but other factors may influence the nature of the genetic rearrangements that implicate insertion of foreign DNA and illegitimate recombination.…”

Section: Discussionmentioning

confidence: 99%

MYC activation associated with the integration of HPV DNA at the MYC locus in genital tumors

et al. 2006

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To determine whether integration of human papillomavirus (HPV) DNA sequences could lead to the deregulation of genes implied in oncogenesis, we analysed the HPV integration sites in a series of nine cell lines derived from invasive genital carcinomas. Using in situ hybridization, HPV16 or 18 sequences were found at chromosome band 8q24, the localization of MYC, in IC1, IC2, IC3, IC6 and CAC-1 cells and at other sites in IC4, IC5, IC7 and IC8 cells. We then localized viral sequences at the molecular level and searched for alterations of MYC structure and expression in these cells. MYC genomic status and viral integration sites were also analysed in primary tumors from which IC1, IC2, IC3 and IC6 cells were derived. In IC1, IC2 and CAC-1 cells, HPV DNA was located within 58 kb of MYC, downstream, upstream, or within MYC. In IC3 and IC6 cells, HPV DNA was located 400-500 kb upstream of MYC. Amplification studies showed that, in IC1, IC2 and IC3, viral and MYC sequences were coamplified in an amplicon between less than 50 and 800 kb in size. MYC amplification was also observed in primary tumors, indicating that this genetic alteration, together with viral insertion at the MYC locus, had already taken place in vivo. MYC was not amplified in the other cell lines. MYC mRNA and protein overexpression was observed in the five cell lines in which the HPV DNA was inserted close to the MYC locus, but in none of the lines where the insertion had occurred at other sites. MYC activation, triggered by the insertion of HPV DNA sequences, can be an important genetic event in cervical oncogenesis.

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“…The numerous proteins that have been suspected to influence integration can be grouped into three main categories: those that (i) interact with IN directly; (ii) bind to the viral DNA to influence the process; or (iii) participate in the final gap repair step. Of equivalent interest is deciphering the roles cellular proteins are likely to play in genome-wide patterns of retroviral integration [34][35][36][37][38][39][40][41][42][43][44][45][46]. Experimental capabilities in this area changed with the availability of the human genome sequence, commensurately advancing bioinformatics, and more recently, a second wave of high-throughput sequencing methodologies [47,48]).…”

Section: Host Cell Factors and Integrationmentioning

confidence: 99%

“…Broadly sketched, lentiviruses favor integrating in active transcription units without favoring the promoter regions in particular [34,35,38,39,43,45,46], MLV favors the promoter regions of genes (transcription start sites and CpG islands) [35,38], and ALV appears, in general, to show less specificity for transcription units [36,37,42]. MMTV shows none at all and is so far the most random integrator [57].…”

Section: Host Cell Factors and Integrationmentioning

confidence: 99%

Integrase, LEDGF/p75 and HIV replication

Poeschla

2008

Cell. Mol. Life Sci.

115

104

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HIV integrates a DNA copy of its genome into a host cell chromosome in each replication cycle. The essential DNA cleaving and joining chemistry of integration is known, but there is less understanding of the process as it occurs in a cell, where two complex and dynamic macromolecular entities are joined: the viral pre-integration complex and chromatin. Among implicated cellular factors, much recent attention has coalesced around LEDGF/p75, a nuclear protein that may act as a chromatin docking factor or receptor for lentiviral pre-integration complexes. LEDGF/p75 tethers HIV integrase to chromatin, protects it from degradation, and strongly influences the genome-wide pattern of HIV integration. Depleting the protein from cells and/or over-expressing its integrase-binding domain blocks viral replication. Current goals are to establish the underlying mechanisms and to determine whether this knowledge can be exploited for antiviral therapy or for targeting lentiviral vector integration in human gene therapy.

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Transcription Start Regions in the Human Genome Are Favored Targets for MLV Integration

Cited by 1,216 publications

References 17 publications

Engineering of PEDF-Expressing Primary Pigment Epithelial Cells by the SB Transposon System Delivered by pFAR4 Plasmids

Engineering of PEDF-Expressing Primary Pigment Epithelial Cells by the SB Transposon System Delivered by pFAR4 Plasmids

MYC activation associated with the integration of HPV DNA at the MYC locus in genital tumors

Integrase, LEDGF/p75 and HIV replication

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