Delayed expression of adeno-associated virus vector DNA.
Two previous reports indicated that recombinant adeno-associated virus (rAAV) vectors were dependent on helper adenovirus (Ad) for efficient conversion of single-stranded (ss) rAAV DNA to the double-stranded (ds) form. This finding is somewhat paradoxical, however, since during a latent infection wild-type (wt)-AAV is rapidly converted to a ds form in the absence of Ad. Our hypothesis was that the effect observed in the previous studies was due to kinetic factors, i.e. to a relative delay in conversion to ds-DNA rather than to an absolute requirement for Ad. To test this, Hela cells were infected with a rAAV-CMV-green fluorescent protein (GFP) vector either in the presence or absence of Ad. Within the first 2 days, Ad infection resulted in a 4-fold increase in AAV vector expression and an augmentation of conversion to a ds-AAV DNA. By 6 days, however, the total number of GFP-expressing cells in the Ad-free culture had exceeded the original number in the Ad co-infected cells, and the conversion to ds-DNA episomes was substantial and ongoing.
Afione
S A
SA
Department of Physiology and Pediatrics, Johns Hopkins University, Baltimore, Md., USA.
Wang
J
J
Walsh
S
S
Guggino
W B
WB
Flotte
T R
TR
D000256:Adenoviridae / Q000502:physiology; D002461:Cell Line, Transformed; D004274:DNA, Recombinant / Q000378:metabolism*; D000229:Dependovirus / Q000235:genetics / Q000502:physiology*; D005822:Genetic Vectors / Q000235:genetics / Q000502:physiology*; D006367:HeLa Cells; D006801:Humans; D013997:Time Factors; D014162:Transfection; D014170:Transformation, Genetic; D014407:Tumor Cells, Cultured; D016662:Virus Integration; D017735:Virus Latency
D004274:DNA, Recombinant