Cytoplasmic mislocalization of POU3F4 due to novel mutations leads to deafness in humans and mice.

POU3F4 is a POU domain transcription factor that is required for hearing. In the ear, POU3F4 is essential for mesenchymal remodeling of the bony labyrinth and is the causative gene for DFNX2 human nonsyndromic deafness. Ear abnormalities underlie this form of deafness, characterized previously in multiple spontaneous, radiation-induced and transgenic mouse mutants. Here, we report three novel mutations in the POU3F4 gene that result in profound hearing loss in both humans and mice. A p.Gln79* mutation was identified in a child from an Israeli family, revealed by massively parallel sequencing (MPS). This strategy demonstrates the strength of MPS for diagnosis with only one affected individual. A second mutation, p.Ile285Argfs*43, was identified by Sanger sequencing. A p.Cys300* mutation was found in an ENU-induced mutant mouse, schwindel (sdl), by positional cloning. The mutation leads to a predicted truncated protein, similar to the human mutations, providing a relevant mouse model. The p.Ile285Argfs*43 and p.Cys300* mutations lead to a shift of Pou3f4 nuclear localization to the cytoplasm, demonstrated in cellular localization studies and in the inner ears of the mutant mice. The discovery of these mutations facilitates a deeper comprehension of the molecular basis of inner ear defects due to mutations in the POU3F4 transcription factor.

Generation of ENU-induced mouse mutants with hypocholesterolemia: novel tools for dissecting plasma lipoprotein homeostasis.

Pathologic plasma lipoprotein cholesterol levels play a key role in the development and pathogenesis of human atherosclerotic cardiovascular diseases. Plasma cholesterol homeostasis is regulated by genetic predispositions and environmental factors. Animal models showing aberrant plasma cholesterol levels are used for the identification and analysis of novel causative genes. Here, we searched for inherited hypocholesterolemia phenotypes in randomly mutant mice which may contribute to the detection of disease protective alleles. In the Munich ENU mouse mutagenesis project, clinical chemistry blood analysis was carried out on more than 15,500 G1 offspring and 230 G3 pedigrees of chemically mutagenized inbred C3H mice to detect dominant and recessive mutations leading to a decreased plasma total cholesterol level. We identified 66 animals consistently showing hypocholesterolemia. Transmission of the altered phenotype to the subsequent generations led to the successful establishment of 14 independent hypocholesterolemic lines. Line-specific differences were detected by clinical chemistry analysis of plasma HDL cholesterol, LDL cholesterol and triglycerides. Thus, we successfully established a novel panel of ENU-derived mutant mouse lines for their use in the identification of alleles selectively influencing the plasma cholesterol homeostasis. Such findings may be subsequently used for humans and other species.

Hypercholesterolemia in ENU-induced mouse mutants.

Hypercholesterolemia is caused by multiple environmental factors and genetic predispositions, and plays an important role in the development and pathogenesis of various human diseases. In this study, we aimed to establish randomly mutant mouse lines showing hypercholesterolemia for their further use in the detection of novel causative alleles. In the Munich ENU Mouse Mutagenesis Project, clinical chemistry blood analysis was performed on more than 15,000 G1 mice and 230 G3 pedigrees of chemically mutagenized mice to detect dominant and recessive mutations leading to an increased plasma total cholesterol level. Using inbred C3HeB/FeJ mice we identified more than 100 animals consistently showing hypercholesterolemia. Transmission of the altered phenotype to the subsequent generations led to the production of nine hypercholesterolemic lines. A single line showed further obvious deviations in the analysis of additional clinical chemistry blood parameters. Thus, the lines produced will contribute to the search for alleles that selectively cause primary hypercholesterolemia.