title: Characterization of Ribozymes Targeting a Congenital Night Blindness Mutation in Rhodopsin Mutation.
abstract: The G90D mutation in the rhodopsin gene leads to autosomal dominant congenital stationary night blindness (CSNB) in patients. This occurs because the G90D mutant protein cannot efficiently bind chromophore and is constitutively active. To combat this mutation, we designed and characterized two different hammerhead ribozymes to cleave G90D transcript. In vitro testing showed that the G90D1 ribozyme efficiently and specifically cleaved the mutant transcript while G90D2 cleaved both WT and mutant transcript. AAV-mediated delivery of G90D1 under the control of the mouse opsin promoter (MOP500) to G90D transgenic eyes showed that the ribozyme partially retarded the functional degeneration (as measured by electroretinography [ERG]) associated with this mutation. These results suggest that with additional optimization, ribozymes may be a useful part of the gene therapy knockdown strategy for dominant retinal disease.
authors.affiliation: Department of Cell Biology, University of Oklahoma Health Sciences Center, 940 Stanton L. Young Blvd. BMSB 781, 73104, Oklahoma City, OK, USA. Shannon-conley@ouhsc.edu.
authors.forename: Shannon M
authors.initials: SM
authors.lastname: Conley
authors.affiliation: Department of Molecular Genetics and Microbiology, University of Florida, 32611, Gainesville, FL, USA.
authors.forename: Patrick
authors.initials: P
authors.lastname: Whalen
authors.affiliation: Department of Molecular Genetics and Microbiology, University of Florida, 32611, Gainesville, FL, USA. lewin@ufl.edu.
authors.forename: Alfred S
authors.initials: AS
authors.lastname: Lewin
authors.affiliation: Department of Cell Biology, University of Oklahoma Health Sciences Center, 940 Stanton L. Young Blvd. BMSB 781, 73104, Oklahoma City, OK, USA. muna-naash@ouhsc.edu.
authors.forename: Muna I
authors.initials: MI
authors.lastname: Naash
mesh_terms: D000818:Animals; D055162:Biocatalysis; D000229:Dependovirus / Q000235:genetics; D004596:Electroretinography; D015785:Eye Diseases, Hereditary / Q000235:genetics* / Q000503:physiopathology / Q000628:therapy; D040181:Genetic Diseases, X-Linked / Q000235:genetics* / Q000503:physiopathology / Q000628:therapy; D015316:Genetic Therapy / Q000379:methods; D005822:Genetic Vectors / Q000235:genetics; D006801:Humans; D008822:Mice, Transgenic; D050505:Mutant Proteins / Q000235:genetics; D009154:Mutation; D009216:Myopia / Q000235:genetics* / Q000503:physiopathology / Q000628:therapy; D009755:Night Blindness / Q000235:genetics* / Q000503:physiopathology / Q000628:therapy; D012313:RNA / Q000235:genetics / Q000378:metabolism; D016337:RNA, Catalytic / Q000235:genetics / Q000378:metabolism*; D012243:Rhodopsin / Q000235:genetics*; D013379:Substrate Specificity; D014158:Transcription, Genetic / Q000235:genetics
chemical_list: D050505:Mutant Proteins; D016337:RNA, Catalytic; C427204:hammerhead ribozyme; D012313:RNA; D012243:Rhodopsin
keywords: Congenital stationary night blindness; G90D; Gene therapy; Retinal degeneration; Rhodopsin; Ribozyme