Wilson's disease (WD), also known as hepatolenticular degeneration is a rare, autosomal recessive disorder. A defect in the hepatic secretion of copper leads to acute or chronic liver disease, neurologic symptoms which can manifest as tremors or by rigidity and contractures and psychiatric symptoms. The biochemical defect is unknown, and this project will apply positional cloning techniques in order to identify the gene. The disease has been localized to 13q14.3-q21.1 between D13S31 and D13S59 at distances of 0.4 and 1.2 cM respectively. However, odds for this location, versus the more proximal location, between D13S31 and D13S25 are currently only 7 times more likely. Physical mapping in this region infers that 1 cM is approximately equal to 1,000kb, suggesting that WD lies 400kb from D13S31. In an attempt to clone the WD gene a 1 Mb YAC contig around D13S31 has been constructed. Highly polymorphic markers are being developed at D13531, D13S59 and D13S25 that should provide excellent odds for the location of the disease interval. Colored FISH will be performed to determine the orientation of the contig, and once the location of the disease is refined, the YAC contig will be extended in that direction. Additional highly polymorphic markers at the ends of the contig will be generated and typed in WD families exhibiting recombination between WND and the closely linked loci. This should define the """"""""smallest co- segregating region"""""""" (SCR) in which the disease gene must lie. Candidate genes in genomic clones derived from the SCR will be screened for by 1) the presence of conserved sequences 2) CpG islands 3) positive signals on Northern blots 4)direct hybridization of cDNA libraries to YACs 5) exon trapping. Genes will be isolated from cDNA libraries, sequenced, and PCR based systems will be developed that amplify cDNA and genomic DNA. PCR products from WD patients and normal individuals will be subjected to single stranded conformation analysis, denaturing gradient gel analysis and hybridization to mutS protein to identify mutations. Any variant that is detected will be sequenced. The existence of a gene with mutation/s specific to WD patients will be strong evidence that the gene has been isolated. Direct comparisons of candidate gene sequences between affected and normal individuals will be performed with PCR amplification and direct sequencing if preliminary screening approaches for mutations are not fruitful.
| Kalinsky, H; Funes, A; Zeldin, A et al. (1998) Novel ATP7B mutations causing Wilson disease in several Israeli ethnic groups. Hum Mutat 11:145-51 |
| Bowcock, A M; Tomfohrde, J; Weissenbach, J et al. (1994) Refining the position of Wilson disease by linkage disequilibrium with polymorphic microsatellites. Am J Hum Genet 54:79-87 |
