Faculty Bibliography

Increased activity of erythrocyte sodium-lithium countertransport is associated with essential hypertension. Sodium-lithium countertransport is highly heritable, but no single gene product mediating the exchange or explaining the association of increased sodium-lithium countertransport activity and hypertension has been identified. We performed a linkage study by using erythrocyte sodium-lithium countertransport as a quantitative phenotype and genome-wide markers at an average resolution of approximately 10 cM to identify quantitative trait loci explaining sodium-lithium countertransport activity. A peak LOD score of 2.83 was detected on chromosome 15q at D15S642, a marker previously shown to be linked to blood pressure. Several genes mapped to this region are possible candidates for factors affecting erythrocyte sodium-lithium countertransport and/or blood pressure. Further studies confirming the presence of a quantitative trait locus in this region and evaluating these candidate genes may help explain the association of elevated sodium-lithium countertransport and hypertension.

Clinical expression of Hirschsprung disease (HSCR) requires the interaction of multiple susceptibility genes. Molecular genetic analyses have revealed that interactions between mutations in the genes encoding the RET receptor tyrosine kinase and the endothelin receptor type B (EDNRB) are central to the genesis of HSCR. We have established two locus noncomplementation assays in mice, using allelic series at Ednrb in the context of Ret kinase-null heterozygotes, to understand the clinical presentation, incomplete penetrance, variation in length of aganglionic segment, and sex bias observed in human HSCR patients. Titration of Ednrb in the presence of half the genetic dose of Ret determines the presentation of an enteric phenotype in these strains, revealing or abrogating a sex bias in disease expression depending on the genotype at Ednrb. RET and EDNRB signaling pathways are also critical for the normal development of other tissues, including the kidneys and neural crest-derived melanocytes. Our data demonstrate that interaction between these genes is restricted to the enteric nervous system and does not affect renal, coat color, and retinal choroid development.

Association studies of candidate genes with complex traits have generally used one or a few single nucleotide polymorphisms (SNPs), although variation in the extent of linkage disequilibrium (LD) within genes markedly influences the sensitivity and precision of association studies. The extent of LD and the underlying haplotype structure for most candidate genes are still unavailable. We sampled 193 blacks (African-Americans) and 160 whites (European-Americans) and estimated the intragenic LD and the haplotype structure in four genes of the renin-angiotensin system. We genotyped 25 SNPs, with all but one of the pairs spaced between 1 and 20 kb, thus providing resolution at small scale. The pattern of LD within a gene was very heterogeneous. Using a robust method to define haplotype blocks, blocks of limited haplotype diversity were identified at each locus; between these blocks, LD was lost owing to the history of recombination events. As anticipated, there was less LD among blacks, the number of haplotypes was substantially larger, and shorter haplotype segments were found, compared with whites. These findings have implications for candidate-gene association studies and indicate that variation between populations of European and African origin in haplotype diversity is characteristic of most genes.

Evidence for genomic regions influencing systolic and diastolic blood pressure (BP) were assessed in a whole genome linkage analysis in 211 African American and 160 white families as part of the GenNet network of the National Heart, Lung and Blood Institute-sponsored Family Blood Pressure Program. Multipoint regression and variance components linkage methods were used to analyze 372 polymorphic markers. Statistically compelling evidence for linkage (P values .0057 and .00023, respectively) was found on chromosome 1. Our results support the idea that BP regulation is most likely governed by multiple genetic loci, each with a relatively weak effect on BP in the population at large.

What has been learnt about individual human biology and common diseases 50 years on from the discovery of the structure of DNA? Unfortunately the double helix has not, so far, revealed as much as one would have hoped. The primary reason is an inability to determine how nurture fits into the DNA paradigm. We argue here that the environment exerts its influence at the DNA level and so will need to be understood before the underlying causal factors of common human diseases can be fully recognized.

Single-nucleotide polymorphisms (SNPs) constitute the great majority of variations in the human genome, and as heritable variable landmarks they are useful markers for disease mapping and resolving population structure. Redundant coverage in overlaps of large-insert genomic clones, sequenced as part of the Human Genome Project, comprises a quarter of the genome, and it is representative in terms of base compositional and functional sequence features. We mined these regions to produce 500,000 high-confidence SNP candidates as a uniform resource for describing nucleotide diversity and its regional variation within the genome. Distributions of marker density observed at different overlap length scales under a model of recombination and population size change show that the history of the population represented by the public genome sequence is one of collapse followed by a recent phase of mild size recovery. The inferred times of collapse and recovery are Upper Paleolithic, in agreement with archaeological evidence of the initial modern human colonization of Europe.

Contemporary genotyping and sequencing methods do not provide information on linkage phase in diploid organisms. The application of statistical methods to infer and reconstruct linkage phase in samples of diploid sequences is a potentially time- and labor-saving method. The Stephens-Smith-Donnelly (SSD) algorithm is one such method, which incorporates concepts from population genetics theory in a Markov chain-Monte Carlo technique. We applied a modified SSD method, as well as the expectation-maximization and partition-ligation algorithms, to sequence data from eight loci spanning >1 Mb on the human X chromosome. We demonstrate that the accuracy of the modified SSD method is better than that of the other algorithms and is superior in terms of the number of sites that may be processed. Also, we find phase reconstructions by the modified SSD method to be highly accurate over regions with high linkage disequilibrium (LD). If only polymorphisms with a minor allele frequency >0.2 are analyzed and scored according to the fraction of neighbor relations correctly called, reconstructions are 95.2% accurate over entire 100-kb stretches and are 98.6% accurate within blocks of high LD.

Genetic studies of Hirschsprung disease, a common congenital malformation, have identified eight genes with mutations that can be associated with this condition. Mutations at individual loci are, however, neither necessary nor sufficient to cause clinical disease. We conducted a genome-wide association study in 43 Mennonite family trios using 2,083 microsatellites and single-nucleotide polymorphisms and a new multipoint linkage disequilibrium method that searches for association arising from common ancestry. We identified susceptibility loci at 10q11, 13q22 and 16q23; the gene at 13q22 is EDNRB, encoding a G protein-coupled receptor (GPCR) and the gene at 10q11 is RET, encoding a receptor tyrosine kinase (RTK). Statistically significant joint transmission of RET and EDNRB alleles in affected individuals and non-complementation of aganglionosis in mouse intercrosses between Ret null and the Ednrb hypomorphic piebald allele are suggestive of epistasis between EDNRB and RET. Thus, genetic interaction between mutations in RET and EDNRB is an underlying mechanism for this complex disorder.