title: Efficient photoreceptor-targeted gene expression in vivo by recombinant adeno-associated virus.
abstract: We describe a general approach for achieving efficient and cell type-specific expression of exogenous genes in photoreceptor cells of the mammalian retina. Recombinant adeno-associated virus (rAAV) vectors were used to transfer the bacterial lacZ gene or a synthetic green fluorescent protein gene (gfp) to mouse or rat retinas after injection into the subretinal space. Using a proximal murine rod opsin promoter (+86 to -385) to drive expression, reporter gene product was found exclusively in photoreceptors, not in any other retinal cell type or in the adjacent retinal pigment epithelium. GFP-expressing photoreceptors typically encompassed 10-20% of the total retinal area after a single 2-microl injection. Photoreceptors were transduced with nearly 100% efficiency in the region directly surrounding the injection site. We estimate approximately 2.5 million photoreceptors were transduced as a result of the single subretinal inoculation. This level of gene transfer and expression suggests the feasibility of genetic therapy for retinal disease. The gfp-containing rAAV stock was substantially free of both adenovirus and wild-type AAV, as judged by plaque assay and infectious center assay, respectively. Thus, highly purified, helper virus-free rAAV vectors can achieve high-frequency tissue-specific transduction of terminally differentiated, postmitotic photoreceptor cells.
authors.affiliation: School of Optometry and Neuroscience Group, University of California, Berkeley, CA 94720, USA.
authors.forename: J G
authors.initials: JG
authors.lastname: Flannery
authors.forename: S
authors.initials: S
authors.lastname: Zolotukhin
authors.forename: M I
authors.initials: MI
authors.lastname: Vaquero
authors.forename: M M
authors.initials: MM
authors.lastname: LaVail
authors.forename: N
authors.initials: N
authors.lastname: Muzyczka
authors.forename: W W
authors.initials: WW
authors.lastname: Hauswirth
mesh_terms: D000818:Animals; D004274:DNA, Recombinant; D000229:Dependovirus; D015870:Gene Expression; D018014:Gene Transfer Techniques; D005822:Genetic Vectors; D007763:Lac Operon; D051379:Mice; D008810:Mice, Inbred C57BL; D010786:Photoreceptor Cells; D051381:Rats; D017207:Rats, Sprague-Dawley; D012160:Retina
chemical_list: D004274:DNA, Recombinant