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Journal of Virology, December 2001, p. 12279-12287, Vol. 75, No. 24
0022-538X/01/$04.00+0   DOI: 10.1128/JVI.75.24.12279-12287.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.

Involvement of Cellular Double-Stranded DNA Break Binding Proteins in Processing of the Recombinant Adeno-Associated Virus Genome

Lorena Zentilin,1 Alessandro Marcello,1 and Mauro Giacca1,2,*

Molecular Medicine Laboratory, International Centre for Genetic Engineering and Biotechnology, 34012 Trieste,1 and Laboratorio di Biologia Molecolare, Scuola Normale Superiore, 56126 Pisa,2 Italy

Received 2 July 2001/Accepted 29 August 2001

Unlike postmitotic tissues in vivo, transduction of cultured cells is poor with recombinant adeno-associated virus (rAAV). The ability of rAAV to transduce cells is greatly enhanced by a variety of agents that induce DNA damage and is elevated in cells defective in the ataxia telangiectasia gene product (ATM), showing increased genomic instability. Here we show that DNA double-stranded break (DSB) repair pathways are involved in the regulation of rAAV transduction efficiency. By quantitative chromatin immunoprecipitation, we found that Ku86 and Rad52 proteins associate with viral DNA inside transduced cells. Both proteins are known to competitively recognize hairpin structures and DNA termini and to promote repair of DSBs, the former by facilitating nonhomologous end joining and the latter by initiating homologous recombination. We found that rAAV transduction is increased in Ku86-defective cells while it is inhibited in Rad52 knockout cells. These results suggest that binding of Rad52 to the rAAV genome might be involved in processing of the vector genome through a homologous recombination pathway.

* Corresponding author. Mailing address: Molecular Medicine Laboratory, International Centre for Genetic Engineering and Biotechnology, Padriciano 99, 34012 Trieste, Italy. Phone: 39-040-3757.324. Fax: 39-040-226555. E-mail: giacca{at}icgeb.trieste.it.

Journal of Virology, December 2001, p. 12279-12287, Vol. 75, No. 24
0022-538X/01/$04.00+0   DOI: 10.1128/JVI.75.24.12279-12287.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.


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