V76D mutation in a conserved gD-crystallin region leads to dominant cataracts in mice.

During a large-scale ENU mutagenesis screen, a mouse mutant with a dominant cataract was detected and referred to as Aey4. Aim of this study was the morphological description of the mutant, the mapping of the mutation, and the characterization of the underlying molecular lesion. The slit-lamp examination revealed a strong nuclear cataract surrounded by a homogeneous milky opacity in the inner cortex.

Crygf(Rop): the first mutation in the Crygf gene causing a unique radial lens opacity.

Purpose: The Rop (radial opacity) mutation, which was recovered in a mutagenicity screen after paternal treatment with procarbazine, was analyzed to determine phenotype, chromosomal localization, candidate genes, and molecular lesion.

Methods: Native lenses were photographed under a dissecting microscope. Histologic sections of the eye were made according to standard procedures.

Ethylnitrosourea-induced base pair substitution affects splicing of the mouse gammaE-crystallin encoding gene leading to the expression of a hybrid protein and to a cataract.

A novel ENU-induced mutation in the mouse leading to a nuclear and cortical opacity of the eye lens (ENU418) was mapped to proximal chromosome 1 by a genome-wide mapping approach. It suggests that the cluster of gamma-crystallin encoding genes (Cryg) and the betaA2-crystallin encoding gene Cryba2 are excellent candidate genes. An A --> G exchange in the middle of intron 1 of the Cryge gene was found as the only alteration cosegregating with the cataractous phenotype.

Characterization of a mutation in the lens-specific MP70 encoding gene of the mouse leading to a dominant cataract.

During an ethylnitrosourea mutagenesis screen, Aey5, a new mouse mutation exhibiting an autosomal dominant congenital cataract was isolated. The cataractous phenotype is visible at the eye opening and progresses to a nuclear and zonular cataract at 2 months of age with no difference in onset or severity between heterozygous and homozygous mutants. Histological analysis revealed that fiber cell differentiation continues at the lens bow region, but the cell nuclei do not degrade normally and remain in the deeper cortex.

Molecular characterization of Pax6(2Neu) through Pax6(10Neu): an extension of the Pax6 allelic series and the identification of two possible hypomorph alleles in the mouse Mus musculus.

Phenotype-based mutagenesis experiments will increase the mouse mutant resource, generating mutations at previously unmarked loci as well as extending the allelic series at known loci. Mapping, molecular characterization, and phenotypic analysis of nine independent Pax6 mutations of the mouse recovered in mutagenesis experiments is presented. Seven mutations result in premature termination of translation and all express phenotypes characteristic of null alleles, suggesting that Pax6 function requires all domains to be intact.

A 6-bp deletion in the Crygc gene leading to a nuclear and radial cataract in the mouse.

Purpose: A mouse mutant expressing a bilateral nuclear and radial cataract was found after paternal treatment with chlorambucil. The purpose of this study was to establish the linkage of the mutation to a particular chromosome to allow molecular characterization. Moreover, the mutants were examined morphologically.

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

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.

Regulation of the human SIX3 gene promoter.

A 2-kb promoter fragment of SIX3, a human transcription factor essential for vertebrate eye development, has been characterized in a gene reporter assay system. The peak of activity implies the 2-kb sequence of SIX3, whereas 5'-deletion constructs of the promoter decreases successively to 60% of the activity starting from the entire promoter. In contrast, cutting off 300 bp of the 3' promoter extinguishes its activity completely.

Muscle actin genes: a first step towards computational classification of tissue specific promoters.

Tissue-specific gene expression is governed by enhancer and promoter sequences determining the specificity most probably by their internal organization of transcription factor binding sites. In case of muscle-specific gene expression excellent compilations of sequence regions responsible for the tissue-specificity are available. We took advantage of such a compilation in order to elucidate organizational features that are directly correlated with promoter specificity.

Epaxial-adaxial-hypaxial regionalisation of the vertebrate somite: evidence for a somitic organiser and a mirror-image duplication.

During vertebrate neural tube formation, the initially lateral borders between the neural and epidermal ectoderm fuse to form the definitive dorsal region of the embryo, while the initially dorsally located notochord-floor plate complex is being internalised. Along the definitive dorso-ventral body axis, one can distinguish an epaxial (dorsal to the notochord) and a hypaxial (ventral to the notochord) body region. The mesodermal somites on both sides of the notochord and neural tube give rise to the trunk skeleton and skeletal muscle.

Sequence analysis of zebrafish chondromodulin-1 and expression profile in the notochord and chondrogenic regions during cartilage morphogenesis.

Chondromodulin-I (ChM-I) is suggested in higher vertebrate systems to function as a key regulatory protein for cartilage development. To further understand the process of chondrogenesis and the function of ChM-I, we have cloned the zebrafish cDNA for chondromodulin-1 (chm1) and have mapped the chm1 gene locus. The expression profile of chm1 was determined during zebrafish embryonic development and compared to that of type II collagen (col2a1).

Neural plate patterning: upstream and downstream of the isthmic organizer.

Two organizing centres operate at long-range distances within the anterior neural plate to pattern the forebrain, midbrain and hindbrain. Important progress has been made in understanding the formation and function of one of these organizing centres, the isthmic organizer, which controls the development of the midbrain and anterior hindbrain. Here we review our current knowledge on the identity, localization and maintenance of the isthmic organizer, as well as on the molecular cascades that underlie the activity of this organizing centre.

Comparative analysis of the genomic organization of Pax9 and its conserved physical association with Nkx2-9 in the human, mouse, and pufferfish genomes.

As a first step towards the identification of cis-regulatory elements of Pax9 by means of comparative genomics, we have analyzed genome regions encompassing the Pax9 gene in three vertebrate species, humans, mice (Mus musculus), and the Japanese pufferfish (Fugu rubripes). We show the genomic organization of Pax9 and its physical association with Nkx2-9 conserved in the three species. We discuss about possible implications of the conserved synteny between Pax9 and Nkx2-9 in a context of vertebrate evolution.

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

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.

First pass annotation of promoters on human chromosome 22.

The publication of the first almost complete sequence of a human chromosome (chromosome 22) is a major milestone in human genomics. Together with the sequence, an excellent annotation of genes was published which certainly will serve as an information resource for numerous future projects. We noted that the annotation did not cover regulatory regions; in particular, no promoter annotation has been provided.

Antagonistic action of Six3 and Prox1 at the gamma-crystallin promoter.

Gamma-crystallin genes are specifically expressed in the eye lens. Their promoters constitute excellent models to analyse tissue-specific gene expression. We investigated murine CRYGE/f promoters of different length in lens epithelial cell lines.

Characterization of a 1-bp deletion in the gammaE-crystallin gene leading to a nuclear and zonular cataract in the mouse.

Purpose: A previous study had found a mouse mutant to have bilateral nuclear cataract with zonular opacity after paternal irradiation with gamma-rays. The mutation was then demonstrated to be allelic with the Cat2 group of dominant cataract mutations and was referred to as Cat2(nz) in a later study. Because several members of this group have been confirmed as mutations in the gene cluster coding for gamma-crystallins (CRYG:), these genes were now tested as candidates for Cat2(nz).

From developmental biology to developmental toxicology.

Progress derived from the human genome project will have tremendous impact on toxicology. Questions concerning genetic susceptibility or resistance to toxic compound exposure and the dissection of the molecular mechanisms involved will be at the forefront of future toxicological research. In recent years, it was recognized that many of the molecular control mechanisms of embryogenesis have been conserved during evolution.

Enzyme-activity mutants in Mus musculus. I. Phenotypic description and genetic characterization of ethylnitrosourea-induced mutations.

The specific activity of erythrocyte enzymes was measured to detect gene mutations in F(1)-offspring of male mice treatment with different doses (80, 160, or 250 mg/kg body weight) of ethylnitrosourea (ENU). Altogether 13,230 offspring were screened for 10 enzyme activities. Mutants with reduced activity as well as mutants with enhanced activity were found.

Saturation mutagenesis for dominant eye morphological defects in the mouse Mus musculus.

We have summarized our extensive series of mutagenesis experiments to isolate dominant mutations in the mouse that express eye morphological defects. Thirty-two experimental groups in which parental mice were exposed to chemical mutagens or irradiation and a historical control group of the laboratory are presented. The largest series of experiments included parental exposure to ethylnitrosourea or irradiation.

Role of Stat3 in lipopolysaccharide-induced IL-10 gene expression.

IL-10 is a unique cytokine because it is anti-inflammatory and immunosuppressive. IL-10 is regulated at the level of transcription, but the critical motifs and the relevant transcription factors controlling this gene have remained elusive to date. We now report that a sequence at -120 bp in the human IL-10 promoter binds Stat3 but no other Stat proteins.

Human adrenoleukodystrophy protein and related peroxisomal ABC transporters interact with the peroxisomal assembly protein PEX19p.

Four ABC half transporters (ALDP, ALDRP, PMP70, and PMP69) have been identified in the mammalian peroxisomal membrane but no function has been unambiguously assigned to any of them. To date X-linked adrenoleukodystrophy (X-ALD) is the only human disease known to result from a defect of one of these ABC transporters, ALDP. Using the yeast two-hybrid system and in vitro GST pull-down assays, we identified the peroxin PEX19p as a novel interactor of ALDP, ALDRP, and PMP70.