Journal article
Concatamerization of Adeno-Associated Virus Circular Genomes Occurs through Intermolecular Recombination
Journal of virology, Vol.73(11), pp.9468-9477
11/1999
DOI: 10.1128/JVI.73.11.9468-9477.1999
PMCID: PMC112981
PMID: 10516055
Abstract
Long-term recombinant AAV (rAAV) transgene expression in muscle has been associated with the molecular conversion of single-stranded rAAV genomes to high-molecular-weight head-to-tail circular concatamers. However, the mechanisms by which these large multimeric concatamers form remain to be defined. To this end, we tested whether concatamerization of rAAV circular intermediates occurs through intra- or intermolecular mechanisms of amplification. Coinfection of the tibialis muscle of mice with rAAV alkaline phosphatase (Alkphos)- and green fluorescent protein (GFP)-encoding vectors was used to evaluate the frequency of circular concatamer formation by intermolecular recombination of independent viral genomes. The GFP shuttle vector also encoded ampicillin resistance and contained a bacterial origin of replication to allow for bacterial rescue of circular intermediates from Hirt DNA of infected muscle samples. The results demonstrated a time-dependent increase in the abundance of rescued plasmids encoding both GFP and Alkphos, which reached 33% of the total circular intermediates by 120 days postinfection. Furthermore, these large circular concatamers were capable of expressing both GFP- and Alkphos-encoding transgenes following transient transfection in cell lines. These findings demonstrate that concatamerization of AAV genomes in vivo occurs through intermolecular recombination of independent monomer circular viral genomes and suggest new viable strategies for delivering multiple DNA segments at a single locus. Such developments will expand the utility of rAAV for splicing large gene inserts or large promoter-gene combinations carried by two or more independent rAAV vectors.
Details
- Title: Subtitle
- Concatamerization of Adeno-Associated Virus Circular Genomes Occurs through Intermolecular Recombination
- Creators
- Jusan Yang - Department of Anatomy and Cell BiologyWeihong Zhou - Department of Anatomy and Cell BiologyYulong Zhang - Department of Anatomy and Cell BiologyTerese Zidon - Department of Anatomy and Cell BiologyTerry Ritchie - Department of Anatomy and Cell BiologyJohn F Engelhardt - Department of Anatomy and Cell Biology
- Resource Type
- Journal article
- Publication Details
- Journal of virology, Vol.73(11), pp.9468-9477
- Publisher
- American Society for Microbiology
- DOI
- 10.1128/JVI.73.11.9468-9477.1999
- PMID
- 10516055
- PMCID
- PMC112981
- ISSN
- 0022-538X
- eISSN
- 1098-5514
- Language
- English
- Date published
- 11/1999
- Academic Unit
- Roy J. Carver Department of Biomedical Engineering; Anatomy and Cell Biology; Radiation Oncology; Internal Medicine
- Record Identifier
- 9984025682002771
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