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Biomedical subjects

W Pretsch

Publications and source records attributed to W Pretsch.

At least 19 recordsLinked to original sources
PubMed2001

A 76-bp deletion in the Mip gene causes autosomal dominant cataract in Hfi mice.

Hfi is a dominant cataract mutation where heterozygotes show hydropic lens fibers and homozygotes show total lens opacity. The Hfi locus was mapped to the distal part of mouse chromosome 10 close to the major intrinsic protein (Mip), which is expressed only in cell membranes of lens fibers. Molecular analysis of Mip revealed a 76-bp deletion that resulted in exon 2 skipping in Mip mRNA. In Hfi/Hfi this deletion resulted in a complete absence of the wildtype Mip. In contrast, Hfi/+ animals had the same amount of wildtype Mip as +/+. Results from pulse-chase expression studies excluded hetero-oligomerization of wildtype and mutant Mip as a possible mechanism for cataract formation in the Hfi/+. We propose that the cataract phenotype in the Hfi heterozygote mutant is due to a detrimental gain of function by the mutant Mip resulting in either cytotoxicity or disruption in processing of other proteins important for the lens. Cataract formation in the Hfi/Hfi mouse is probably a combined result of both the complete loss of wildtype Mip and a gain of function of the mutant Mip.

Animals↗
PubMed2000

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. Of the 36 independently observed mutations, 20 were homozygous lethal. Genetic and biochemical characterizations were routinely performed. These mutants provide insight into the mechanism of ENU mutagenesis and can serve as models for structure-function studies of the corresponding enzymes.

Animals↗
PubMed2000

A comparison of enzyme activity mutation frequencies in germ cells of mice (Mus musculus) and golden hamsters (Mesocricetus auratus) after exposure to 2 + 2 Gy gamma-irradiation.

The radiation-induced germ cell mutation rate has been investigated in two species of mammals. Mice and golden hamsters of both sexes were exposed to 2 + 2 Gy gamma-irradiation with a 24 h fractionation interval and mated to untreated partners. In mice, specific locus mutations were examined as positive controls and the obtained mutation rates (per locus and gamete x10(-5)) were 51.4, 10.1, 13.6 and 17.4 for irradiated post-spermatogonia, spermatogonia and 1-7 and >7 days post-treatment oocytes, respectively. Offspring of mice and golden hamsters were screened for activity alterations of 10 erythrocyte enzymes coded by at least 14 loci. The observed mutation rates per locus per gamete x10(-5) for treated post-spermatogonial stages, spermatogonia and oocytes 1-7 and >7 days post-treatment were 6.5, 1.5, 8.8 and 7.0, respectively, for mice and 16.7, 0, 7.6 and 0, respectively, for golden hamsters. There is a significant difference for mutation rates in mouse oocytes 1-7 days post-treatment compared with the control. No differences in the frequencies of mutations in the various germ cell stages could be observed between mice and golden hamsters. A critical assumption for the extrapolation of experimental mutagenesis studies to humans is that no species effects exist in sensitivity to mutation induction by irradiation. Our results do not contradict this assumption.

Animals↗
PubMed1999

Glutathione reductase activity deficiency in homozygous Gr1a1Neu mice does not cause haemolytic anaemia.

A glutathione reductase (GR) mutant with approximately 50% residual enzyme activity in blood compared with wild-type was detected amongst offspring of isopropyl methanesulphonate-treated male mice. Homozygous mutants with only 2% residual enzyme activity were recovered in progeny of inter se matings of heterozygotes. Results of linkage studies indicate a mutation at the Gr1 structural locus on chromosome 8. The loss of GR activity was evident both in blood and in other tissue extracts. Erythrocyte and organo-somatic indices did not show differences between wild-types and homozygous mutants, indicating no association between the GR deficiency and haemolytic anaemia in this potential animal model.

Anemia, Hemolytic↗
PubMed1998

Three murine cataract mutants (Cat2) are defective in different gamma-crystallin genes.

A number of murine cataract mutations have been localized to chromosome 1 close to the gamma-crystallin gene cluster (Cryg) (Everett et al., 1994, Genomics 20: 429-434; Löster et al., 1994, Genomics 23: 240-242). Based on the size of the mapping or allelism tests they have not been shown to be genetically distinct and have been assigned to locus symbol Cat2. Here we assign three mutations to the respective gamma-crystallin gene. Using a systematic candidate gene approach to analyze the entire Cryg cluster, an A-->G transition was found in exon 2 of Cryga for the ENU-436 mutation and is designated Cryga1Neu. The mutant allele Crygbnop (formerly Cat2(nop)) is caused by a replacement of 11 bp by 4 bp in the third exon of Crygb, while a C-->G transversion in exon 3 of Cryge has been found for the Cryget (formerly Cat2(t)) mutation. For the mutation Cryga1Neu, an Asp-->Gly exchange is deduced, whereas the mutations Crygbnop and Cryget lead to the formation of in-frame stop codons and give rise to truncated proteins of 144 and 143 amino acids, respectively. The effects of the mutations upon gamma-crystallin structure are likely to be quite different. The Cryga1Neu mutation is expected to affect the link between Greek-key motifs 2 and 3, whereas both Crygbnop and Cryget mutations are supposed to truncate the fourth Greek-key motif. All three mutations are predicted to alter protein folding of the gamma-crystallins and result in lens cataract, but the phenotype for each is quite distinctive.

Amino Acid Sequence↗
PubMed1998

Molecular, genetic and biochemical characterization of lactate dehydrogenase-A enzyme activity mutations in Mus musculus.

Four independent heterozygous lactate dehydrogenase (LDH) mutations with approximately 60% of wild-type enzyme activity in whole blood have been recovered. The mutant line Ldh1a2Neu proved to be homozygous lethal, whereas for the three lines Ldh1a7Neu, Ldh1a11Neu, and Ldh1a12Neu homozygous mutants with about 20% residual activity occurred in the progeny of heterozygous inter se matings. However, the number of homozygous mutants was less than expected, suggesting an increased lethality of these animals. Various physicochemical and kinetic properties of LDH are altered. Exons of the Ldh1 gene were PCR amplified and sequenced to determine the molecular lesion in the mutant alleles. Ldh1a2Neu carried an A/T-->G/C transition in codon 112 (in exon 3), resulting in an Asn-->Asp substitution; Asn112 is part of the helix alpha D, which is involved in the coenzyme-binding domain. Ldh1a7Neu contained an A/T-->C/G transversion within the codon for residue 194 in exon 4, causing an Asp-->Ala substitution, which may affect the arrangement of the substrate-binding site. Three base substituions were discovered for the mutation Ldh1a11Neu in exon 7: the transition C/G-->T/A, a silent mutation, and two transversions C/G-->A/T and C/G-->G/C, both missense mutations, which led to the amino acid replacements A1a319-->Glu and Thr321-->Ser, respectively, located in the alpha H helix structure of the COOH tail of LDHA. We suggest that the mutation in the result of a gene conversion event between Ldh1a wild-type gene and the pseudogene Ldhl-ps. The alteration Ile-->Thr of codon 241 in exon 6 caused by the base pair change T/A-->C/G was identified in the mutation Ldh1a12Neu; Ile241 is included in the helix alpha 2G, a structure that is indirectly involved in coenzyme binding. Each of the sequence alterations has a potential impact on the structure of the LDHA protein, which is consistent with the decreased LDH activity and biochemical and physiological alterations.

Amino Acid Sequence↗
PubMed1997

Genetic mapping of a mouse ocular malformation locus, Tcm, to chromosome 4.

The Tcm mutation in the mouse is an autosomal dominant ocular malformation manifesting as microphthalmia, iris dysplasia, cataract, and coloboma. As a first step to cloning the Tcm gene, we report the localization of the Tcm mutation with respect to known microsatellite markers. Backcross progeny carrying the Tcm mutation were produced by mating Tcm/+ heterozygous mice to normal C57BL/6 partners. Genomic DNA from each mouse was subjected to PCR analysis to identify simple sequence length polymorphisms. Our results locate Tcm to Chr 4 and suggest candidate genes responsible for the Tcm phenotype. Finally, ocular histopathology was done in 3-week-old animals to define the extent of the malformation.

Abnormalities, Multiple↗
PubMed1997

The mouse Cat4 locus maps to chromosome 8 and mutants express lens-corneal adhesion.

Cat4 is the second largest allelism group in the collection of mouse dominant eye mutations recovered in Neuherberg and carriers express anterior polar cataract, central corneal opacity, and lens-corneal adhesions. We have mapped the Cat4 locus of the mouse to central Chromosome (Chr) 8 at position cM 31. Histological characterization of Cat4(a) heterozygotes and homozygotes indicates failure of separation of the lens vesicle from the surface ectoderm. Human anterior segment ocular dysgenesis (ASOD) is autosomal dominant, carriers express an eye phenotype similar to that of Cat4(a) carriers, and it has been mapped to a region of 4q homologous to mouse central Chr 8. Thus, on the basis of phenotype and map position, Cat4 may be a mouse model of human ASOD. The genes Junb, Jund1, Mel, and Zfp42 are discussed as possible candidates for Cat4.

Animals↗
PubMed1997

Detection of a point mutation (A to G) in exon 5 of the murine Mgf gene defines a novel allele at the Steel locus with a weak phenotype.

A new mutation at the locus encoding the mast cell growth factor (Mgf) is described and designated as MgfSl-3Neu. Homozygous mutants have a light grey fur, sometimes with white patches. Homozygotes are fertile, but with reduced litter size, when mated inter se. Analysis of haematological parameters indicated no difference between mutant and wild-type mice. Sequence analysis of the cDNA obtained from the brain of homozygous mutants revealed an A-->G exchange at position 400 leading to a predicted amino acid exchange from Asn-->Leu at position 122. As a consequence of the predicted amino acid exchange an extension of the alpha-helical context and a decreased hydropathicity of the region at positions 101-125 can be deduced. This single amino acid exchange is outside of the known important domains of MGF and explains the weak phenotype of MgfSl-3Neu.

Alleles↗
PubMed1997

Mapping of the autosomal dominant cataract mutation (Coc) on mouse chromosome 16.

PURPOSE: To characterize the mouse cataract mutation Coc. METHODS: Coc is an X-radiation-induced autosomal dominant cataract mutation maintained on a murine C3H inbred strain. The affected heterozygotes were outcrossed to C57BL/6, and (C3H Coc/+ x C57BL/6) mice that were Coc/+ were then backcrossed to C57BL/6 to generate a panel of 103 progeny for mapping. For linkage analysis, microsatellites from each autosome were selected. The maximum distance between markers was 30 centimorgans (cM). RESULTS: The initial genome-wide screen of 14 backcrossed progeny indicated that the Coc locus resides on chromosome 16. Further mapping with additional markers from chromosome 16 for all 103 backcrossed progeny positioned Coc between markers D16Mit134 and D16Mit63. This region is syntenic to human chromosome 3. CONCLUSIONS: Mapping of the Coc locus to mouse chromosome 16 provides the positional information necessary to identify the candidate gene responsible for the Coc phenotype. The molecular characterization of the gene disrupted in the Coc mutation will provide insight into the mechanisms involved in cataract formation.

Animals↗
PubMed1996

The mouse Pax2(1Neu) mutation is identical to a human PAX2 mutation in a family with renal-coloboma syndrome and results in developmental defects of the brain, ear, eye, and kidney.

We describe a new mouse frameshift mutation (Pax2(1Neu)) with a 1-bp insertion in the Pax2 gene. This mutation is identical to a previously described mutation in a human family with renal-coloboma syndrome [Sanyanusin, P., McNoe, L. A., Sullivan, M. J., Weaver, R. G. & Eccles, M. R. (1995) Hum. Mol. Genet. 4, 2183-2184]. Heterozygous mutant mice exhibit defects in the kidney, the optic nerve, and retinal layer of the eye, and in homozygous mutant embryos, development of the optic nerve, metanephric kidney, and ventral regions of the inner ear is severely affected. In addition, we observe a deletion of the cerebellum and the posterior mesencephalon in homozygous mutant embryos demonstrating that, in contrast to mutations in Pax5, which is also expressed early in the mid-hindbrain region, loss of Pax2 gene function alone results in the early loss of the mid-hindbrain region. The mid-hindbrain phenotype is similar to Wnt1 and En1 mutant phenotypes, suggesting the conservation of gene regulatory networks between vertebrates and Drosophila.

Abnormalities, Multiple↗
PubMed1996

Purkinje cell lineage and the topographic organization of the cerebellar cortex: a view from X inactivation mosaics.

We utilized a strain of mice, derived from a radiation mutagenesis experiment and carrying an activity-attenuated allele of the X-linked enzyme glucose-6-phosphate dehydrogenase (G6PD), to analyze the development of the cell lineage leading to cerebellar Purkinje neurons. Due to random X inactivation during early embryonic development, X- linked genes can be used to distinguish between clonally related populations of cells in X inactivation mosaics. Following histochemical staining for G6PD activity, the numeric proportions of Purkinje cells expressing either the wild-type or the mutant enzyme and the spatial distribution of these cellular phenotypes and their relation to anatomically and genetically defined cerebellar compartments were analyzed. Our data suggest that cerebellar Purkinje neurons originate from a limited pool of some 129 precursors. The size of this pool is different from the one derived from chimeric mice, allowing us to deduce the relative timing of Purkinje cell lineage restriction. Our data also show that Purkinje neurons of distinct lineage are extensively intermingled within the cerebellar cortex. Together, these findings suggest both a role for cell-cell communication in the development of genetically defined cerebellar compartments and a temporal window during which such cellular interactions may take place.

Animals↗
PubMed1995

Molecular analysis of four ENU induced triosephosphate isomerase null mutants in Mus musculus.

Four ENU-induced mutations were previously identified at the triosephosphate isomerase (TPI) locus in mouse germinal mutation experiments. Each of the mutants is associated with a 50% loss of enzymatic activity in the F1 (heterozygous) animals. Exons of the TPI gene from control mice and heterozygous mutant mice were PCR amplified and sequenced as necessary to determine the molecular lesion in the mutant alleles. Mutants Tpi*M-1NEU and Tpi*M-2NEU carried the same T:A to A:T transversion in exon 6, resulting in a Leu to Gln substitution at residue 192. Amino acid residue 192 is located in alpha-helix H6 of the protein. Tpi*M-4NEU contained a T:A to A:T transversion within the codon for residue 162 in exon 5, also causing a Leu to Gln substitution. This mutation is located at the beginning of beta-strand B6, within a highly conserved sequence region surrounding the active site residue Glu 165. Sequence analysis of Tpi*M-3NEU revealed an A:T to C:G transversion, changing the stop codon to a codon for Cys, with the resulting addition of 19 predominantly hydrophobic amino acids to the protein. All four mutations occurred at an A:T base pair. In each case, the mutation site was flanked on both sides by G:C base pairs. Each of the sequence alterations has a potential impact on the structure of the TPI protein that is consistent with the existence of a null allele. In addition to providing insight into the molecular basis of ENU induced germ cell mutations and the differences in mutation spectra among organisms, these mutants represent models for structure-function studies of this highly conserved enzyme.

Animals↗
PubMed1994

Close linkage of the dominant cataract mutations (Cat-2) with Idh-1 and cryge on mouse chromosome 1.

The murine dominant gene Cat-2 was located on chromosome 1 between the loci of fuzzy and leaden. Subsequent linkage analysis revealed one recombinant between Cat-2t and isocitrate dehydrogenase-1, and one between Cat-2t and gamma E-crystallin among 338 offspring in three-point backcrosses. The resulting genetic distance between the loci is 0.3 +/- 0.3 cM. The very close linkage between the Cat-2 and the gamma-crystallin gene cluster together with the finding of reduced gamma-crystallin transcripts in mutant lenses suggest strongly that the gamma-crystallin genes may be candidate genes for the Cat-2 mutations.

Animals↗
PubMed1994

Molecular analysis of four lactate dehydrogenase-A mutants in the mouse.

Four electrophoretic and/or enzyme-activity variants of murine LDH-A subunit (Ldhla-m1Neu, Ldhla-m5Neu, Ldhla-m6Neu, Ldhla-m9Neu), induced by procarbazine hydrochloride or ethylnitrosourea (ENU), were analyzed at the DNA level. The exons of the Ldhl gene from homozygous mutants were amplified by PCR and sequenced. Three mutations resulted from nucleotide substitutions in exon 5: the transitions A-->G at codons 216 (Ldhla-m5Neu) and 225 (Ldhla-m6Neu), and the transversion G-->C (Ldhla-m1Neu) at codon 222. The mutations resulted in the replacements of Glu by Gly (Ldhla-m5Neu), Gln by Arg (Ldhla-m6Neu) and Asp by His (Ldhla-m1Neu). The fourth base substitution, the transition T-->C (Ldhla-m9Neu), has been found at the GT donor splice site following the first exon; this mutation affected the efficiency of transcription. All ENU-induced mutations were A/T-->G/C transitions. The mutation events could be correlated with the biochemical and physiological alterations observed in affected mice.

Amino Acid Sequence↗