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Generation, characterization, and molecular cloning of the Noerg-1 mutation of rhodopsin in the mouse

Published online by Cambridge University Press:  06 December 2005

LAWRENCE H. PINTO ,
MARTHA H. VITATERNA ,
KAZUHIRO SHIMOMURA ,
SANDRA M. SIEPKA ,
ERIN L. MCDEARMON ,
DEBORAH FENNER ,
STEPHEN L. LUMAYAG ,
CHIAKI OMURA ,
ANNE W. ANDREWS  and
MATTHEW BAKER
...Show all authors
LAWRENCE H. PINTO
Affiliation:
Department of Neurobiology and Physiology and Center for Functional Genomics, Northwestern University, Evanston
MARTHA H. VITATERNA
Affiliation:
Department of Neurobiology and Physiology and Center for Functional Genomics, Northwestern University, Evanston
KAZUHIRO SHIMOMURA
Affiliation:
Department of Neurobiology and Physiology and Center for Functional Genomics, Northwestern University, Evanston
SANDRA M. SIEPKA
Affiliation:
Department of Neurobiology and Physiology and Center for Functional Genomics, Northwestern University, Evanston
ERIN L. MCDEARMON
Affiliation:
Department of Neurobiology and Physiology and Center for Functional Genomics, Northwestern University, Evanston
DEBORAH FENNER
Affiliation:
Department of Neurobiology and Physiology and Center for Functional Genomics, Northwestern University, Evanston
STEPHEN L. LUMAYAG
Affiliation:
Department of Neurobiology and Physiology and Center for Functional Genomics, Northwestern University, Evanston
CHIAKI OMURA
Affiliation:
Department of Neurobiology and Physiology and Center for Functional Genomics, Northwestern University, Evanston
ANNE W. ANDREWS
Affiliation:
Department of Neurobiology and Physiology and Center for Functional Genomics, Northwestern University, Evanston
MATTHEW BAKER
Affiliation:
Department of Neurobiology and Physiology and Center for Functional Genomics, Northwestern University, Evanston
BRANDON M. INVERGO
Affiliation:
Department of Neurobiology and Physiology and Center for Functional Genomics, Northwestern University, Evanston
MARISSA A. OLVERA
Affiliation:
Department of Ophthalmology & Visual Sciences, University of Iowa, Iowa City
EDWARD HEFFRON
Affiliation:
Department of Ophthalmology & Visual Sciences, University of Iowa, Iowa City
ROBERT F. MULLINS
Affiliation:
Department of Ophthalmology & Visual Sciences, University of Iowa, Iowa City
VAL C. SHEFFIELD
Affiliation:
Departments of Pediatrics and Genetics, University of Iowa, Iowa City Howard Hughes Medical Institute, Carver College of Medicine, Iowa City
EDWIN M. STONE
Affiliation:
Department of Ophthalmology & Visual Sciences, University of Iowa, Iowa City
JOSEPH S. TAKAHASHI
Affiliation:
Department of Neurobiology and Physiology and Center for Functional Genomics, Northwestern University, Evanston Howard Hughes Medical Institute, Carver College of Medicine, Iowa City
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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 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.

Keywords

Chemical mutagenesisENUForward geneticsRetinaMouse modelsRetinitis pigmentosaMolecular cloningPositional cloningGene discoveryRhodopsinProtein foldingDisulfide bond formationRodsRod photoreceptorRho-C110Y mutationOptomotor responseVisual acuity
Type
Research Article
Information
Visual Neuroscience , Volume 22 , Issue 5 , September 2005 , pp. 619 - 629
Copyright
© 2005 Cambridge University Press

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