Genetic and Environmental Factors in Deletion Disorders

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DESCRIPTION (Investigator's abstract): A genomic disorder occurs de novo in region-specific repeats. Prototypic of genomic disorders are Prader-Willi and Angelman syndromes (PWS and AS), which arise in -1/7,500 births from a 4 Mb deletion of chromosome 15q11-q13. Our recent studies have shown the presence of 2-5 copies of paralogous sequences (duplicons) at or near each of a distal and two proximal PWS/AS breakpoint hotspots. The duplicons are derived in large part from genomic duplications of a novel gene (HERC2), while the tight clustering of breakpoints suggests a specific recombination mechanism leads to these deletions. We hypothesize that these duplicated sequences are involved in inter- and intra-chromosomal misalignment and homologous recombination, and that transcription of the duplicons in germ cells facilitates the recombination process. Finally, paternal occupational exposure to hydrocarbons has been associated with PWS, and we have shown that UV, X-rays, hydrocarbons and other chemicals enhance recombination of a similar genomic duplication in mice. Therefore, both genetic and environmental factors appear to predispose to chromosomal deletion events in PWS and AS. We propose four Specific Aims to examine the molecular mechanisms, and role of genetic and environmental susceptibility, in chromosome 15q1 1-q13 rearrangements: Aim 1. To determine the structure (number, orientation, sequence, gene content, and population variation) of HERC2-duplicons at 15Q11 and 15q13 by molecular cytogenetic and sequence echnologies. Aim 2. To identify sequence-specific hotspot(s) for chromosome recombination, we will clone and sequence deletion breakpoints using two novel assays, one detecting aberrant HERC2-related transcripts, and sequencing / informatics. Aim 3. We will examine genetic susceptibility (effect of inter-repeat distance, transcription and number of repeat copies) to intra- and inter-chromosomal deletion using a HERC2-GFP duplication model, including hotspot sequences (Aim 2), with assessment of the male germ cell rearrangement frequency using flow cytometry to detect GFP-tagged recombinants. Aim 4. We will determine whether environmental factors enhance the frequency of germ line recombination within Hprt- and HERC2-duplication mouse models, and similarly will examine genetic susceptibility using Ku80 about and p53- mice deficient in recombination pathways. These studies will identify the structure, variation and molecular pathological mechanisms underlying rearrangement of complex duplicated sequences in the mammalian germ line. Identification of environmental risk factors for germ line chromosome deletions may allow prevention of pre-conceptional exposure in the human population.