Ethylnitrosourea-induced mutation in mice leads to the expression of a novel protein in the eye and to dominant cataracts.

in Genetics (2001), 157(3), 1313-20

A novel ENU-induced mutation in the mouse leading to a nuclear and zonular opacity of the eye lens (Aey1) was mapped to chromosome 1 between the markers D1Mit303 and D1Mit332. On the basis of the ... [more ▼]

A novel ENU-induced mutation in the mouse leading to a nuclear and zonular opacity of the eye lens (Aey1) was mapped to chromosome 1 between the markers D1Mit303 and D1Mit332. On the basis of the chromosomal position, the gamma-crystallin encoding gene cluster (Cryg) and the betaA2-crystallin encoding gene Cryba2 were tested as candidate genes. An A --> T mutation destroys the start codon of the Cryge gene in the mutants; this mutation was confirmed by the absence of a restriction site for NcoI in the corresponding genomic fragment of homozygous mutants. The next in-frame start codon is 129 bp downstream; this predicted truncated gammaE-crystallin consists of 131 amino acids, resulting in a molecular mass of 14 kD. However, another open reading frame was observed just 19 bp downstream of the regular Cryge start codon, resulting in a protein of 119 amino acids and a calculated molecular weight of 13 kD. Western blot analysis using polyclonal antibodies against gamma-crystallins or the novel Aey1-specific protein demonstrated the specific expression of the Aey1 protein in the cataractous lenses only; the truncated form of the gammaE-crystallin could not be detected. Therefore, it is concluded that the novel protein destroys the sensitive cellular structure of the eye lens. [less ▲]

Characterization of a new, dominant V124E mutation in the mouse alphaA-crystallin-encoding gene.

in Investigative Ophthalmology and Visual Science (2001), 42(12), 2909-15

PURPOSE: During an ethylnitrosourea (ENU) mutagenesis screening, mice were tested for the occurrence of dominant cataracts. The purpose of the study was morphologic description, mapping of the mutant gene ... [more ▼]

PURPOSE: During an ethylnitrosourea (ENU) mutagenesis screening, mice were tested for the occurrence of dominant cataracts. The purpose of the study was morphologic description, mapping of the mutant gene, and characterization of the underlying molecular lesion in a particular mutant, Aey7. METHODS: Isolated lenses were photographed and histologic sections of the eye were analyzed according to standard procedures. Linkage analysis was performed with a set of microsatellite markers covering all autosomal chromosomes. cDNA was amplified after reverse transcription of lens mRNA. For PCR, cDNA or genomic DNA was used as a template. RESULTS: Nuclear opacity and posterior suture anomaly were visible at eye opening and progressed to a nuclear and zonular cataract at 2 months of age. The opacity as well as the microphthalmia was more pronounced in the homozygotes than in the heterozygotes. The mutation was mapped to chromosome 17 between the markers D17Mit133 and D17Mit180. This position made the alphaA-crystallin-encoding gene (Cryaa) an excellent candidate gene. Sequence analysis revealed a mutation of a T to an A at position 371 in the Cryaa cDNA. The mutation was confirmed by an additional MnlI restriction site in the genomic DNA of homozygous mutants leading to replacement of Val with Glu at codon 124 affecting the C-terminal region of the alphaA-crystallin. CONCLUSIONS: The Aey7 mutant represents the first dominant mouse cataract mutation affecting the Cryaa gene. The mutation leads to progressive opacification of the lens. Compared with the beta- and gamma-crystallin-encoding genes, mutations in the alpha-crystallin-encoding genes are rare. [less ▲]

The Notch ligand Jagged1 is required for inner ear sensory development.

in Proceedings of the National Academy of Sciences of the United States of America (2001), 98(7), 3873-8

Within the mammalian inner ear there are six separate sensory regions that subserve the functions of hearing and balance, although how these sensory regions become specified remains unknown. Each sensory ... [more ▼]

Within the mammalian inner ear there are six separate sensory regions that subserve the functions of hearing and balance, although how these sensory regions become specified remains unknown. Each sensory region is populated by two cell types, the mechanosensory hair cell and the supporting cell, which are arranged in a mosaic in which each hair cell is surrounded by supporting cells. The proposed mechanism for creating the sensory mosaic is lateral inhibition mediated by the Notch signaling pathway. However, one of the Notch ligands, Jagged1 (Jag1), does not show an expression pattern wholly consistent with a role in lateral inhibition, as it marks the sensory patches from very early in their development--presumably long before cells make their final fate decisions. It has been proposed that Jag1 has a role in specifying sensory versus nonsensory epithelium within the ear [Adam, J., Myat, A., Roux, I. L., Eddison, M., Henrique, D., Ish-Horowicz, D. & Lewis, J. (1998) Development (Cambridge, U.K.) 125, 4645--4654]. Here we provide experimental evidence that Notch signaling may be involved in specifying sensory regions by showing that a dominant mouse mutant headturner (Htu) contains a missense mutation in the Jag1 gene and displays missing posterior and sometimes anterior ampullae, structures that house the sensory cristae. Htu/+ mutants also demonstrate a significant reduction in the numbers of outer hair cells in the organ of Corti. Because lateral inhibition mediated by Notch predicts that disruptions in this pathway would lead to an increase in hair cells, we believe these data indicate an earlier role for Notch within the inner ear. [less ▲]

Genome-wide, large-scale production of mutant mice by ENU mutagenesis.

in Nature Genetics (2000), 25(4), 444-7

In the post-genome era, the mouse will have a major role as a model system for functional genome analysis. This requires a large number of mutants similar to the collections available from other model ... [more ▼]

In the post-genome era, the mouse will have a major role as a model system for functional genome analysis. This requires a large number of mutants similar to the collections available from other model organisms such as Drosophila melanogaster and Caenorhabditis elegans. Here we report on a systematic, genome-wide, mutagenesis screen in mice. As part of the German Human Genome Project, we have undertaken a large-scale ENU-mutagenesis screen for dominant mutations and a limited screen for recessive mutations. In screening over 14,000 mice for a large number of clinically relevant parameters, we recovered 182 mouse mutants for a variety of phenotypes. In addition, 247 variant mouse mutants are currently in genetic confirmation testing and will result in additional new mutant lines. This mutagenesis screen, along with the screen described in the accompanying paper, leads to a significant increase in the number of mouse models available to the scientific community. Our mutant lines are freely accessible to non-commercial users (for information, see http://www.gsf.de/ieg/groups/enu-mouse.html). [less ▲]

Mutation in the betaA3/A1-crystallin encoding gene Cryba1 causes a dominant cataract in the mouse.

in Genomics (1999), 62(1), 67-73

During the mouse ENU mutagenesis screen, mice were tested for the occurrence of dominant cataracts. One particular mutant was discovered as a progressive opacity (Po). Heterozygotes show opacification of ... [more ▼]

During the mouse ENU mutagenesis screen, mice were tested for the occurrence of dominant cataracts. One particular mutant was discovered as a progressive opacity (Po). Heterozygotes show opacification of a superficial layer of the fetal nucleus, which progresses and finally forms a nuclear opacity. Since the homozygotes have already developed the total cataract at eye opening, the mode of inheritance is semidominant. Linkage analysis was performed using a set of genome-wide microsatellite markers. The mutation was mapped to chromosome 11 distal of the marker D11Mit242 (9.3 +/- 4.4 cM) and proximal to D11Mit36 (2.3 +/- 2.3 cM). This position makes the betaA3/A1-crystallin encoding gene Cryba1 an excellent candidate gene. Mouse Cryba1 was amplified from lens mRNA. Sequence analysis revealed a mutation of a T to an A at the second base of exon 6, leading to an exchange of Trp by Arg. Computer analysis predicts that the fourth Greek key motif of the affected betaA3/A1-crystallin will not be formed. Moreover, the mutation leads also to an additional splicing signal, to the skipping of the first 3 bp of exon 6, and finally to the deletion of the Trp residue. Both types of mRNA are present in the homozygous mutant lenses. The mutation will be referred to as Cryba1(po1). This particular mouse mutation provides an excellent animal model for a human congenital zonular cataract with suture opacities, which is caused by a mutation in the homologous gene. [less ▲]