Scott C. Wildenberg

Evidence That a Locus for Familial High Myopia Maps to Chromosome 18p

Myopia, or nearsightedness, is the most common human eye disorder. A genomewide screen was conducted to map the gene(s) associated with high, early-onset, autosomal dominant myopia. Eight families that each included two or more individuals with >=-6.00 diopters (D) myopia, in two or more successive generations, were identified. Myopic individuals had no clinical evidence of connective-tissue abnormalities, and the average age at diagnosis of myopia was 6.8 years. The average spherical component refractive error for the affected individuals was -9.48 D. The families contained 82 individuals; of these, DNA was available for 71 (37 affected). Markers flanking or intragenic to the genes for Stickler syndrome types 1 and 2 (chromosomes 12q13.1-q13.3 and 6p21.3, respectively), Marfan syndrome (chromosome 15q21.1), and juvenile glaucoma (chromosome 1q21-q31) were also analyzed. No evidence of linkage was found for markers for the Stickler syndrome types 1 and 2, the Marfan syndrome, or the juvenile glaucoma loci. After a genomewide search, evidence of significant linkage was found on chromosome 18p. The maximum LOD score was 9.59, with marker D18S481, at a recombination fraction of .0010. Haplotype analysis further refined this myopia locus to a 7.6-cM interval between markers D18S59 and D18S1138 on 18p11.31.

Evidence for Locus Heterogeneity in Puerto Ricans with Hermansky-Pudlak Syndrome

Hermansky-Pudlak syndrome (HPS) consists of ocu-locutaneous albinism, a platelet storage-pool deficiency, and ceroid lipofuscinosis. In a recent report on the cloning of an HPS gene, all 22 Puerto Rican HPS patients were homozygous for a 16-bp duplication in exon 15. This presumably reflected a founder effect for the HPS mutation in Puerto Rico. Nevertheless, we ascertained two individuals from central Puerto Rico who lacked the 16-bp duplication, exhibited significant amounts of normal-size HPS mRNA by northern blot analysis, and had haplotypes in the HPS region that were different from the haplotype of every 16-bp-duplication patient. Moreover, these two individuals displayed no mutations in their cDNA sequences, throughout the entire HPS gene. Both patients exhibited pigment dilution, impaired visual acuity, nystagmus, a bleeding diathesis, and absent platelet dense bodies, confirming the diagnosis of HPS. These findings indicate that analysis of Puerto Rican patients for the 16-bp duplication in HPS cannot exclude the diagnosis of HPS. In addition, HPS most likely displays locus heterogeneity, consistent with the existence of several mouse strains manifesting both pigment dilution and a platelet storage-pool deficiency.

Detection of a Tsp509I polymorphism in the 3' UTR of the human tyrosinase related protein-1 (TYRP) gene.

We have identified a Tsp509I polymorphism in the 3′ UTR of the human tyrosinase related protein-1 gene (TYRP). TYRP is one of several genes involved in melanin pigment production.

Genetic linkage analysis using pooled DNA and infrared detection of tailed STRP primer patterns

The mapping of a disease locus to a specific chromosomal region is an important step in the eventual isolation and analysis of a disease causing gene. Conventional mapping methods analyze large multiplex families and/or smaller nuclear families to find linkage between the disease and a chromosome marker that maps to a known chromosomal region. This analysis is time consuming and tedious, typically requiring the determination of 30,000 genotypes or more. For appropriate populations, we have instead utilized pooled DNA samples for gene mapping which greatly reduces the amount of time necessary for an initial chromosomal screen. This technique assumes a common founder for the disease locus of interest and searches for a region of a chromosome shared between affected individuals. Our analysis involves the PCR amplification of short tandem repeat polymorphisms (STRP) to detect these shared regions. In order to reduce the cost of genotyping, we have designed unlabeled tailed PCR primers which, when combined with a labeled universal primer, provides for an alternative to synthesizing custom labeled primers. The STRP pattern is visualized with an infrared fluorescence based automated DNA sequencer and the patterns quantitated by densitometric analysis of the allele pattern. Differences in the distribution of alleles between pools of affected and unaffected individuals, including a reduction in the number of alleles in the affected pool, indicate the sharing of a region of a chromosome. We have found this method effective for markers 10 - 15 cM away from the disease locus for a recessive genetic disease.

Three novel polymorphisms in the gene responsible for the Hermansky-Pudlak syndrome

Acknowledgments: This work was supported by NIH grant #AR44649.