This Article
Right arrow Full Text
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrowReprints and Permissions
Right arrow Copyright Information
Right arrow Books from ASM Press
Right arrow MicrobeWorld
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Rinaudo, D.
Right arrow Articles by Toniatti, C.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Rinaudo, D.
Right arrow Articles by Toniatti, C.

 Previous Article  |  Next Article 

Journal of Virology, January 2000, p. 281-294, Vol. 74, No. 1
0022-538X/0/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.

Conditional Site-Specific Integration into Human Chromosome 19 by Using a Ligand-Dependent Chimeric Adeno-Associated Virus/Rep Protein

Daniela Rinaudo, Stefania Lamartina, Giuseppe Roscilli, Gennaro Ciliberto, and Carlo Toniatti*

Department of Genetics, Istituto di Ricerche di Biologia Molecolare, I.R.B.M.---Piero Angeletti, 00040 Pomezia (Rome), Italy

Received 7 May 1999/Accepted 21 September 1999

It is of great interest for gene therapy to develop vectors that drive the insertion of a therapeutic gene into a chosen specific site on the cellular genome. Adeno-associated virus (AAV) is unique among mammalian viruses in that it integrates into a distinct region of human chromosome 19 (integration site AAVS1). The inverted terminal repeats (ITRs) flanking the AAV genome and the AAV-encoded nonstructural proteins Rep78 and/or Rep68 are the only viral elements necessary and sufficient for site-specific integration. However, it is also known that unrestrained Rep activity may cause nonspecific genomic rearrangements at AAVS1 and/or have detrimental effects on cell physiology. In this paper we describe the generation of a ligand-dependent form of Rep, obtained by fusing a C-terminally deleted Rep68 with a truncated form of the hormone binding domain of the human progesterone receptor, which does not bind progesterone but binds only its synthetic antagonist RU486. The activity of this chimeric protein, named Rep1-491/P, is highly dependent on RU486 in various assays: in particular, it triggers site-specific integration at AAVS1 of an ITR-flanked cassette in a ligand-dependent manner, as efficiently as wild-type Rep68 but without generating unwanted genomic rearrangement at AAVS1.

* Corresponding author. Mailing address: Istituto di Ricerche di Biologia Molecolare, I.R.B.M.---P. Angeletti, Via Pontina Km 30.600, 00040 Pomezia (Rome), Italy. Phone: 39-06-91093668. Fax: 39-06-91093654. E-mail: toniatti{at}irbm.it.

Journal of Virology, January 2000, p. 281-294, Vol. 74, No. 1
0022-538X/0/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.


This article has been cited by other articles:

  • van Nierop, G. P., de Vries, A. A. F., Holkers, M., Vrijsen, K. R., Goncalves, M. A. F. V. (2009). Stimulation of homology-directed gene targeting at an endogenous human locus by a nicking endonuclease. Nucleic Acids Res 37: 5725-5736 [Abstract] [Full Text]  
  • Sharma, N., Moldt, B., Dalsgaard, T., Jensen, T. G., Mikkelsen, J. G. (2008). Regulated gene insertion by steroid-induced {Phi}C31 integrase. Nucleic Acids Res 36: e67-e67 [Abstract] [Full Text]  
  • McAlister, V. J., Owens, R. A. (2007). Preferential Integration of Adeno-Associated Virus Type 2 into a Polypyrimidine/Polypurine-Rich Region within AAVS1. J. Virol. 81: 9718-9726 [Abstract] [Full Text]  
  • Young, S. M. Jr., Samulski, R. J. (2001). Adeno-associated virus (AAV) site-specific recombination does not require a Rep-dependent origin of replication within the AAV terminal repeat. Proc. Natl. Acad. Sci. USA 10.1073/pnas.241508998v1 [Abstract] [Full Text]  
  • Minamino, T., Gaussin, V., DeMayo, F. J., Schneider, M. D. (2001). Inducible Gene Targeting in Postnatal Myocardium by Cardiac-Specific Expression of a Hormone-Activated Cre Fusion Protein. Circ. Res. 88: 587-592 [Abstract] [Full Text]  
  • Lamartina, S., Ciliberto, G., Toniatti, C. (2000). Selective Cleavage of AAVS1 Substrates by the Adeno-Associated Virus Type 2 Rep68 Protein Is Dependent on Topological and Sequence Constraints. J. Virol. 74: 8831-8842 [Abstract] [Full Text]  
  • Lamartina, S., Sporeno, E., Fattori, E., Toniatti, C. (2000). Characteristics of the Adeno-Associated Virus Preintegration Site in Human Chromosome 19: Open Chromatin Conformation and Transcription-Competent Environment. J. Virol. 74: 7671-7677 [Abstract] [Full Text]  
  • Young, S. M. Jr., Samulski, R. J. (2001). Adeno-associated virus (AAV) site-specific recombination does not require a Rep-dependent origin of replication within the AAV terminal repeat. Proc. Natl. Acad. Sci. USA 98: 13525-13530 [Abstract] [Full Text]  


Copyright © 2009 by the American Society for Microbiology.   For an alternate route to Journals.ASM.org, visit: http://intl-journals.asm.org | More Info»