Genome function in Mouse with Combination Mutagenesis

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DESCRIPTION (provided by applicant): This proposal seeks 3 years of support to complete functional analyses of a 50 Mb region of mouse Chromosome 5 comprising nearly 2% of the mouse genome. Under previous funding, mutations were induced by randomly mutagenizing the genome with ethylnitrosourea (ENU), and selecting those on proximal Chr 5 using a breeding scheme that exploited a chromosome inversion called rump-white (Rw). A set of deletion complexes spanning much of the Rw region was created to aid in the genetic analysis of these mutations. Though screens were conducted for several phenotypes, embryonic lethal mutations (total of 34, representing 32 complementation groups) were the largest class recovered. Additionally, two male mutations causing infertility, one causing deafness, and 2 affecting behavior were recovered. The lethal mutations act throughout development, from pre-implantation through birth. The mutant phenotypes include: failure of the inner cell mass to proliferate; abnormal placental formation; gastrulation defects; homeotic-like skeletal transformations; craniofacial abnormalities; and cardiovascular defects. Genetic mapping experiments have localized most of these mutations to relatively small intervals within the Rw region. Thus far, 5 lethals and 1 infertility allele have been cloned. This proposal has two specific aims. The first is to positionally clone the remaining infertility mutation and 20 of the lethal alleles. The second is to complete a gross phenotypic characterization of this lethal mutation set, using analytic methods appropriate for various embryological stages. In combination, these experiments will link the molecular defects to mutant phenotypes, and thereby illuminate genetic mechanisms of normal mammalian development. Many of the uncloned mutations currently map to regions containing none or few known lethal genes, ensuring that several new essential genes or functional elements will be identified. This establishment of phenotype/genotype relationships will pave the way for more detailed investigations by ourselves and the mammalian development community. Overall, these experiments will eventually provide an unbiased glimpse into the functional elements encoded by a representative region of the genome. The project uses the mouse as model to identify important genes affecting embryonic development and fertility in humans. Ultimately, we anticipate this work will assist treatment of certain male infertilities, increase our understanding of the genes required for normal embryonic development, and potentially lead to improved genetic diagnosis of certain birth defects.