title: Generation, characterization, and molecular cloning of the Noerg-1 mutation of rhodopsin in the mouse.
abstract: We performed genome-wide mutagenesis of C57BL/6J mice using the mutagen N-ethyl-N-nitrosourea (ENU) and screened the third generation (G3) offspring for visual system alterations using electroretinography and fundus photography. Several mice in one pedigree showed characteristics of retinal degeneration when tested at 12-14 weeks of age: no recordable electroretinogram (ERG), attenuation of retinal vessels, and speckled pigmentation of the fundus. Histological studies showed that the retinas undergo a photoreceptor degeneration with apoptotic loss of outer nuclear layer nuclei but visual acuity measured using the optomotor response under photopic conditions persists in spite of considerable photoreceptor loss. The Noerg-1 mutation showed an autosomal dominant pattern of inheritance in progeny. Studies in early postnatal mice showed degeneration to occur after formation of partially functional rods. The Noerg-1 mutation was mapped genetically to chromosome 6 by crossing C57BL/6J mutants with DBA/2J or BALB/cJ mice to produce an N2 generation and then determining the ERG phenotypes and the genotypes of the N2 offspring at multiple loci using SSLP and SNP markers. Fine mapping was accomplished with a set of closely spaced markers. A non-recombinant region from 112.8 Mb to 115.1 Mb was identified, encompassing the rhodopsin (Rho) coding region. A single nucleotide transition from G to A was found in the Rho gene that is predicted to result in a substitution of Tyr for Cys at position 110, in an intradiscal loop. This mutation has been found in patients with autosomal dominant retinitis pigmentosa (RP) and results in misfolding of rhodopsin expressed in vitro. Thus, ENU mutagenesis is capable of replicating mutations that occur in human patients and is useful for generating de novo models of human inherited eye disease. Furthermore, the availability of the mouse genomic sequence and extensive DNA polymorphisms made the rapid identification of this gene possible, demonstrating that the use of ENU-induced mutations for functional gene identification is now practical for individual laboratories.
authors.affiliation: Department of Neurobiology and Physiology and Center for Functional Genomics, Northwestern University, Evanston, Il 60208, USA. larry-pinto@northwestern.edu
authors.forename: Lawrence H
authors.initials: LH
authors.lastname: Pinto
authors.forename: Martha H
authors.initials: MH
authors.lastname: Vitaterna
authors.forename: Kazuhiro
authors.initials: K
authors.lastname: Shimomura
authors.forename: Sandra M
authors.initials: SM
authors.lastname: Siepka
authors.forename: Erin L
authors.initials: EL
authors.lastname: McDearmon
authors.forename: Deborah
authors.initials: D
authors.lastname: Fenner
authors.forename: Stephen L
authors.initials: SL
authors.lastname: Lumayag
authors.forename: Chiaki
authors.initials: C
authors.lastname: Omura
authors.forename: Anne W
authors.initials: AW
authors.lastname: Andrews
authors.forename: Matthew
authors.initials: M
authors.lastname: Baker
authors.forename: Brandon M
authors.initials: BM
authors.lastname: Invergo
authors.forename: Marissa A
authors.initials: MA
authors.lastname: Olvera
authors.forename: Edward
authors.initials: E
authors.lastname: Heffron
authors.forename: Robert F
authors.initials: RF
authors.lastname: Mullins
authors.forename: Val C
authors.initials: VC
authors.lastname: Sheffield
authors.forename: Edwin M
authors.initials: EM
authors.lastname: Stone
authors.forename: Joseph S
authors.initials: JS
authors.lastname: Takahashi
mesh_terms: D019943:Amino Acid Substitution; D000818:Animals; D015153:Blotting, Western; D002874:Chromosome Mapping; D003001:Cloning, Molecular; D004247:DNA / Q000096:biosynthesis / Q000235:genetics; D004596:Electroretinography; D005038:Ethylnitrosourea / Q000494:pharmacology; D005838:Genotype; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D009153:Mutagens / Q000494:pharmacology; D009154:Mutation / Q000235:genetics*; D009801:Oculomotor Muscles / Q000502:physiology; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D012160:Retina / Q000002:abnormalities; D012162:Retinal Degeneration / Q000235:genetics / Q000473:pathology; D012243:Rhodopsin / Q000235:genetics*
chemical_list: D009153:Mutagens; D012333:RNA, Messenger; D004247:DNA; D012243:Rhodopsin; D005038:Ethylnitrosourea