PAX6 MiniPromoters drive restricted expression from rAAV in the adult mouse retina

• Published in: Molecular therapy. Methods & clinical development 2016-01, Vol.3 (C), p.16051-16051, Article 16051
• Main Authors: Hickmott, Jack W, Chen, Chih-Yu, Arenillas, David J, Korecki, Andrea J, Lam, Siu Ling, Molday, Laurie L, Bonaguro, Russell J, Zhou, Michelle, Chou, Alice Y, Mathelier, Anthony, Boye, Sanford L, Hauswirth, William W, Molday, Robert S, Wasserman, Wyeth W, Simpson, Elizabeth M
• Format: Article
• Language: English
• Subjects: Aniridia; Binding sites; Bioinformatics; Blindness; Consortia; Deoxyribonucleic acid; Diabetic retinopathy; DNA; Engineering research; Gene therapy; Genomes; Glaucoma; Horizontal cells; Medical research; Mutation; Neighborhoods; Pax6 protein; Plasmids; Promoters; Regulation; Regulatory sequences; Retina; Software; Stem cells; Transcription factors
• ISSN: 2329-0501; 2329-0501
• DOI: 10.1038/mtm.2016.51

Current gene therapies predominantly use small, strong, and readily available ubiquitous promoters. However, as the field matures, the availability of small, cell-specific promoters would be greatly beneficial. Here we design seven small promoters from the human paired box 6 (PAX6) gene and test them in the adult mouse retina using recombinant adeno-associated virus. We chose the retina due to previous successes in gene therapy for blindness, and the PAX6 gene since it is: well studied; known to be driven by discrete regulatory regions; expressed in therapeutically interesting retinal cell types; and mutated in the vision-loss disorder aniridia, which is in need of improved therapy. At the PAX6 locus, 31 regulatory regions were bioinformatically predicted, and nine regulatory regions were constructed into seven MiniPromoters. Driving Emerald GFP, these MiniPromoters were packaged into recombinant adeno-associated virus, and injected intravitreally into postnatal day 14 mice. Four MiniPromoters drove consistent retinal expression in the adult mouse, driving expression in combinations of cell-types that endogenously express Pax6: ganglion, amacrine, horizontal, and Müller glia. Two PAX6-MiniPromoters drive expression in three of the four cell types that express PAX6 in the adult mouse retina. Combined, they capture all four cell types, making them potential tools for research, and PAX6-gene therapy for aniridia.