Structural chromosomal rearrangements are an important cause of human genetic disorders associated with mental retardation and other human developmental abnormalities. The etiology of recurring constitutional chromosomal rearrangements such as deletions, duplications, and marker chromosome formation is poorly understood. The investigators seek to address this problem by a complete genomic and molecular characterization of the chromosome region 15q11-q13, which is associated with an unusually high frequency and diversity of recurring rearrangements including one of the most frequent interstitial deletions observed in humans (1/10,000-1/20,000 births). Due to genomic imprinting, paternal deletions produce Prader-Willi syndrome (PWS) and maternal deletions produce the distinct genetic disorders, Angelman syndrome (AS). This region is also involved in 50 percent of all supernumerary marker chromosomes, and less frequently in intrachromosomal duplications, triplications, and jumping translocations. Evidence now suggests the presence of four hotspots for chromosome breakage in this region: two proximal to the PWS/AS region and two distal. Complete molecular characterization of this region may lead to a more basic understanding of the mechanism of chromosomal rearrangements. Specific goals of the project include the development of a high-resolution physical map of the approximately 8 Mb (reduced in size by 2 Mb) chromosome segment 15cen-15q14 using YAC and BAC contigs and STS content mapping. The goal is 1 STS/50 kb. The map will be anchored at the chromosome 15 centromere by YAC and BAC walking to close the current 1-2cM gap (previously a 12 cM gap). Progress will be monitored by interphase FISH analysis and by genetic mapping in ovarian teratomas with newly developed STRs and/or SNPs. The four recurring breakpoints (hotspots) will be cloned and sequenced to test the hypothesis that multiple copies of a large duplicated gene cluster (200-600 kb) mediate intrachromosomal and interchromosomal rearrangements. Features of these sequences, such as copy number, distribution, and orientation will be analyzed in normal controls and parents of children with de novo abnormalities to assess polymorphism or to identify any specific predisposing factors or genomic rearrangements. These sequences will also be studied among other great ape species for evidence of """"""""evolutionary hotspots"""""""" for chromosomal rearrangement. The proposed orientation of the 4 duplicated clusters (direct-inverted-direct-direct) provides a framework for proposing and testing specific models to account for the diverse rearrangements observed in chromosome 15. An extensive patient collection will be developed representing each class of rearrangement (deletion, duplication, triplication, marker chromosome, jumping translocation) and each of the 4 breakpoint hotspots. For each patient and class of abnormality, the investigators will determine the content, parental origin, and specific breakpoint as well as determine whether the rearrangements are due to meiotic unequal crossing-over or intrachromosomal events. Specific breakpoints will be determined by FISH and pulse field gel electrophoresis, followed by sequencing in selected cases. Development of rapid and precise breakpoint specific assays and methylation assays may facilitate patient analysis and clinical diagnosis.
