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 nonrecombinant 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.
