Site-specific integration by adeno-associated virus is directed by a cellular DNA sequence.

Giraud, Catherine; Winocour, Ernest; Berns, Kenneth I.

doi:10.1073/pnas.91.21.10039

Cited by 139 publications

(126 citation statements)

References 31 publications

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“…from HA16 cells. PCR using a TNNT1 (derived from exon 14) and an AAV-specific primer (derived from the cap gene) resulted in one band of the expected size, the identity of which was confirmed by restriction analysis (data not shown).…”

Section: Tnnt1 Rearrangements In Latently Infectedmentioning

confidence: 92%

“…It was shown by using an episomal model system that site specificity is determined by cellular sequences (14) and that a 33-nt sequence is necessary and sufficient for this targeted nonhomologous recombination event to occur (15). This 33-nt chromosomal sequence is similar to the minimal viral origin of DNA replication, consisting of an RBS and a TRS (16), leading to the concept that Rep-mediated DNA replication is involved in the integration mechanism.…”

mentioning

confidence: 99%

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Adeno-associated virus site-specifically integrates into a muscle-specific DNA region

Dutheil

Shi

Dupressoir

et al. 2000

Proc. Natl. Acad. Sci. U.S.A.

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The nonpathogenic human virus adeno-associated virus type 2 (AAV) has evolved the potentially unique strategy to establish latency by site-specifically integrating its genome into human chromosome 19 (19q13.3-qter) at a locus designated AAVS1. This nonhomologous, site-specific recombination of viral DNA with the human genome provides a basis for developing targeted gene therapy vectors. To assess whether the region surrounding AAVS1 might have contributed to the selection of the specific integration site, we have investigated this locus. Here, we show that AAVS1 is closely linked to the slow skeletal troponin T gene, TNNT1, which has been mapped previously to 19q13.4. In support of this idea, we demonstrate that site-specific AAV DNA integration can result in the formation of TNNT1-AAV junctions. The question now arises whether muscle represents a natural target tissue for latent AAV infection. This possibility is of additional interest in view of recent observations that muscle tissue is particularly well suited for AAV-mediated gene transfer. The question also occurs whether latent infection by AAV can lead to phenotypic changes of the multinucleated muscle fiber cells.T he human parvovirus adeno-associated virus type 2 (AAV) has adopted the strategy of alternate life styles to ensure a successful relationship with its host (1). This nonpathogenic virus productively replicates only in the presence of helper virus co-or superinfection (2). In the absence of such helper conditions, AAV has evolved a strategy potentially unique among eukaryotic viruses. On infection of the non-coinfected host cell, AAV establishes latency by integrating its genome sitespecifically into a region on the q arm of human chromosome 19 (19q13.3-qter) (3, 4), designated AAVS1.The 4.7-kb linear single-stranded genome contains two ORFs and is flanked by inverted terminal repeats (ITRs) (5). The right ORF encodes three capsid proteins, and the left ORF encodes the four nonstructural proteins (Rep proteins) that are involved in regulating all aspects of the viral life style (2).The 145-nt ITRs are the only viral sequences required in cis for DNA replication, packaging of the viral genome into the capsid, and site-specific DNA integration. Within the ITRs, a Rep binding site (RBS) allows for specific recruitment of the large Rep proteins (i.e., Rep68 and Rep78) to the origin of replication (6-8). A Rep-specific endonuclease site (terminal resolution site, TRS) is separated from the RBS by a 13-nt spacer (9-11). Together, RBS and TRS can act as a minimal origin for Rep-mediated DNA replication (12).To dissect the requirements for site specificity in targeted AAV DNA integration, an 8.2-kb fragment of AAVS1 has been isolated from embryonic lung fibroblast (WI38) DNA, and a 4-kb sequence was determined (13). It was shown by using an episomal model system that site specificity is determined by cellular sequences (14) and that a 33-nt sequence is necessary and sufficient for this targeted nonhomologous recombination event to occur (15). This 33-n...

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Section: Tnnt1 Rearrangements In Latently Infectedmentioning

confidence: 92%

mentioning

confidence: 99%

Adeno-associated virus site-specifically integrates into a muscle-specific DNA region

Dutheil

Shi

Dupressoir

et al. 2000

Proc. Natl. Acad. Sci. U.S.A.

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show abstract

“…To map cis-acting elements of AAVS1 essential for sitespecific AAV integration, Giraud et al 10 utilized an Epstein-Barr virus (EBV)-derived shuttle plasmid that can be stably maintained as an episome in eukaryotic cells and subsequently recovered as a bacterial plasmid. Various portions of the AAVS1 locus were cloned into the EBV shuttle plasmid and individual constructs were used to establish shuttle plasmid-bearing cell lines.…”

Section: Aavs1 and Integrationmentioning

confidence: 99%

Adeno-associated virus integration: virus versus vector

Smith

2008

Gene Ther

147

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“…In addition, genes introduced by rAAV can provide continuous production of the recombinant transgene after a single application (9 -11). Infection with wild-type AAV alone leads to the establishment of a long-term latency, which is due primarily to site-specific integration in the AAVS1 site on human chromosome 19 (12), although some forms persist as episomes or are integrated at other sites (13).…”

mentioning

confidence: 99%

Gene Delivery in Renal Tubular Epithelial Cells Using Recombinant Adeno-Associated Viral Vectors

Chen¹,

Agarwal²,

Glushakova³

et al. 2003

Journal of the American Society of Nephrology

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ABSTRACT. Gene therapy has the potential to provide a therapeutic strategy for numerous renal diseases such as diabetic nephropathy, chronic rejection, Alport syndrome, polycystic kidney disease, and inherited tubular disorders. In previous studies using cationic liposomes or adenoviral or retroviral vectors to deliver genes into the kidney, transgene expression has been transient and often associated with adverse host immune responses, particularly with the use of adenoviral vectors. The unique properties of recombinant adeno-associated viral (rAAV) vectors permit long-term stable transgene expression with a relatively low host immune response. The purpose of the present study was to evaluate gene expression in the rat kidney after intrarenal arterial infusion of a rAAV (serotype 2) vector encoding green fluorescence protein (GFP) induced by a cytomegalovirus-chicken beta-actin hybrid promoter. The left kidney of experimental animals was treated with either saline or transduced with rAAV2-GFP (0.125 ml/100 g body wt, 1 × 1010/ml infectious units) through the renal artery. A time-dependent expression of GFP was observed in all kidneys injected with rAAV2-GFP, with maximal expression observed at 6 wk posttransduction. The expression of GFP was restricted to cells in the S3 segment of the proximal tubule and intercalated cells in the collecting duct, the latter identified by co-localization with H+-ATPase. No transduction was observed in the glomeruli or the intrarenal vasculature. These studies demonstrate successful transgene expression in tubular epithelial cells, specifically in the S3 segment of the proximal tubule and intercalated cells, after intrarenal administration of a rAAV vector and provide the impetus for further studies to exploit its use as a tool for gene therapy in the kidney.E-mail: agarwal@nersp.nerdc.ufl.edu

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Site-specific integration by adeno-associated virus is directed by a cellular DNA sequence.

Cited by 139 publications

References 31 publications

Adeno-associated virus site-specifically integrates into a muscle-specific DNA region

Adeno-associated virus site-specifically integrates into a muscle-specific DNA region

Adeno-associated virus integration: virus versus vector

Gene Delivery in Renal Tubular Epithelial Cells Using Recombinant Adeno-Associated Viral Vectors

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