Genomic analysis of the chromosome 15q11-q13 Prader-Willi syndrome region and characterization of transcripts for GOLGA8E and WHCD1L1 from the proximal breakpoint region

<p>Abstract</p> <p>Background</p> <p>Prader-Willi syndrome (PWS) is a neurobehavioral disorder characterized by neonatal hypotonia, childhood obesity, dysmorphic features, hypogonadism, mental retardation, and behavioral problems. Although PWS is most often caused by a paternal interstitial deletion of a 6-Mb region of chromosome 15q11-q13, the identity of the exact protein coding or noncoding RNAs whose deficiency produces the PWS phenotype is uncertain. There are also reports describing a PWS-like phenotype in a subset of patients with full mutations in the <it>FMR1 </it>(fragile X mental retardation 1) gene. Taking advantage of the human genome sequence, we have performed extensive sequence analysis and molecular studies for the PWS candidate region.</p> <p>Results</p> <p>We have characterized transcripts for the first time for two UCSC Genome Browser predicted protein-coding genes, <it>GOLGA8E </it>(golgin subfamily a, 8E) and <it>WHDC1L1 </it>(WAS protein homology region containing 1-like 1) and have further characterized two previously reported genes, <it>CYF1P1 </it>and <it>NIPA2</it>; all four genes are in the region close to the proximal/centromeric deletion breakpoint (BP1). <it>GOLGA8E</it> belongs to the golgin subfamily of coiled-coil proteins associated with the Golgi apparatus. Six out of 16 golgin subfamily proteins in the human genome have been mapped in the chromosome 15q11-q13 and 15q24-q26 regions. We have also identified more than 38 copies of <it>GOLGA8E</it>-like sequence in the 15q11-q14 and 15q23-q26 regions which supports the presence of a <it>GOLGA8E</it>-associated low copy repeat (LCR). Analysis of the 15q11-q13 region by PFGE also revealed a polymorphic region between BP1 and BP2. <it>WHDC1L1 </it>is a novel gene with similarity to mouse <it>Whdc1 </it>(WAS protein homology region 2 domain containing 1) and human JMY protein (junction-mediating and regulatory protein). Expression analysis of cultured human cells and brain tissues from PWS patients indicates that <it>CYFIP1 </it>and <it>NIPA2</it> are biallelically expressed. However, we were not able to determine the allele-specific expression pattern for <it>GOLGA8E </it>and <it>WHDC1L1 </it>because these two genes have highly related sequences that might also be expressed.</p> <p>Conclusion</p> <p>We have presented an updated version of a sequence-based physical map for a complex chromosomal region, and we raise the possibility of polymorphism in the genomic orientation of the BP1 to BP2 region. The identification of two new proteins <it>GOLGA8E</it> and <it>WHDC1L1</it> encoded by genes in the 15q11-q13 region may extend our understanding of the molecular basis of PWS. In terms of copy number variation and gene organization, this is one of the most polymorphic regions of the human genome, and perhaps the single most polymorphic region of this type.</p

Physical Map of 1p36, Placement of Breakpoints in Monosomy 1p36, and Clinical Characterization of the Syndrome

Monosomy 1p36 is the most common terminal deletion syndrome. This contiguous gene deletion syndrome is presumably caused by haploinsufficiency of a number of genes. We have constructed a contig of overlapping large-insert clones for the most distal 10.5 Mb of 1p36, evaluated the deletion sizes in 61 subjects with monosomy 1p36 from 60 families, and created a natural deletion panel. We found pure terminal deletions, interstitial deletions, derivative chromosomes, and more complex rearrangements. Breakpoints were “binned” into 0.5-Mb regions. Analyses revealed some clustering of breakpoints but no single common breakpoint. Determination of the parental origin showed that 60% of de novo 1p36 terminal deletions arose from the maternally inherited chromosome. Of the 61 subjects, 30 were examined systematically through a protocol at the Texas Children's Hospital General Clinical Research Center. Specifically, we report hearing evaluations, palatal and ophthalmological examinations, echocardiograms, neurological assessments, and thyroid function tests. To our knowledge, this systematic molecular and clinical characterization of monosomy 1p36 is the largest and most comprehensive study of this deletion syndrome to date. Many cytogenetically visible, apparent terminal deletions are more complex than anticipated by cytogenetics, as revealed at the molecular level by our study. Our clinical findings allow for the more accurate recognition of the syndrome and for proper medical evaluation

Development of a comparative genomic hybridization microarray and demonstration of its utility with 25 well-characterized 1p36 deletions

Chromosomal abnormalities, such as deletions and duplications, are characterized by specific and often complex phenotypes resulting from an imbalance in normal gene dosage. However, routine chromosome banding is not sensitive enough to detect subtle chromosome aberrations (<5-10 Mb). Array-based comparative genomic hybridization (array CGH) is a powerful new technology capable of identifying chromosomal imbalance at a high resolution by co-hybridizing differentially labeled test and control DNAs to a microarray of genomic clones. We used a previously assembled contig of large-insert clones that span 10.5 Mb of the most distal region of 1p36 to design a microarray. The array includes 97 clones from 1p36, 41 clones from the subtelomeric regions of all human chromosomes, and three clones from each of the X and Y chromosomes. We used this microarray to study 25 subjects with well-characterized deletions of 1p36. All array CGH results agree with the deletion sizes and locations of the breakpoints in these subjects as determined previously by FISH and microsatellite analyses. Terminal deletions, interstitial deletions, derivative chromosomes and complex rearrangements were also identified. We anticipate that array CGH will change the diagnostic approach to many congenital and acquired genetic diseases such as mental retardation, birth defects and cancer