Development and characterization of novel empty adenovirus capsids and their impact on cellular gene expression.
Adenovirus (Ad) has been extensively studied as a eukaryotic viral vector. As these vectors have evolved from first-generation vectors to vectors that contain either very few or no viral genes ("gutless" Ad), significant reductions in the host innate immune response upon infection have been observed. Regardless of these vector improvements an unknown amount of toxicity has been associated with the virion structural proteins. Here we demonstrate the ability to generate high particle numbers (10(11) to 10(12)) of Ad empty virions based on a modification of Cre/lox gutless Ad vectors. Using a battery of analyses (electron microscopy, atomic force microscopy, confocal images, and competition assays) we characterized this reagent and determined that it (i) makes intact virion particles, (ii) competes for receptor binding with wild-type Ad, and (iii) enters the cell proficiently, demonstrating an ability to carry out essential steps of viral entry. To further study the biological impact of these Ad empty virions on infected cells, we carried out DNA microarray analysis. Compared to that for recombinant Ad, the number of mRNAs modulated upon infection was significantly reduced but the expression signatures were similar. This reagent provides a valuable tool for studies of Ad in that researchers can examine the effect of infection in the presence of the virion capsid alone.
Stilwell
Jackie L
JL
UNC Gene Therapy Center, University of North Carolina, Chapel Hill, North Carolina 27599, USA.
McCarty
Douglas M
DM
Negishi
Atsuko
A
Superfine
Richard
R
Samulski
R Jude
RJ
D000256:Adenoviridae / Q000502:physiology*; D002213:Capsid / Q000502:physiology*; D004279:DNA, Viral / Q000648:ultrastructure; D004591:Electrophoresis, Polyacrylamide Gel; D015967:Gene Expression Regulation, Viral / Q000502:physiology*; D018625:Microscopy, Atomic Force; D008854:Microscopy, Electron; D014764:Viral Proteins / Q000235:genetics / Q000648:ultrastructure
D004279:DNA, Viral; D014764:Viral Proteins