Search for functional alleles in the human genome with focus on cardiovascular disease candidate genes

The genetic investigation of human disorders largely through linkage mapping has led to the discovery of candidate genes and mutations as risk factors for those disorders where there is a high degree of penetrance. While twin studies have provided evidence that there are major genetic contributions to multifactorial diseases like coronary artery disease, it has proven difficult to find and replicate significant genetic associations for such diseases. Recent advances in technology, throughput and understanding of widespread human genetic variation at the genomic level (e.g., the HapMap project) have allowed the application of more genetic markers in larger sample studies, but we are still lacking a complete picture of genetic contributions to major multifactorial diseases.Searching for genetic variants with evidence of a direct molecular impact on the expression and function of genes vital to disease development and progression is one valid approach to this problem. There is a growing appreciation that one major class of variation acts at the level of mRNA expression. Traditional tools for studying this class of variation (e.g., reporter gene assays) in the laboratory have severe limitations, mainly in that they lack the in vivo context where the alleles are hypothesized to have a functional impact. This dissertation relies heavily on the application of a relatively novel technique, the measurement of allelic expression imbalances (AEI) between chromosomes in primary human tissues. Using these measurements as phenotypic traits, we demonstrate that cis-acting alleles exerting molecular affects on mRNA expression can often be readily mapped. In the largest survey to date of AEI in primary human tissues we find that AEI in disease candidate genes is quite common, and that the functional contributors to these expression phenotypes are often not regulatory polymorphisms, but polymorphisms found directly within the mRNAs and affecting mRNA processing and functions. Computational analysis of mRNA structures and genetic variation within human genomes indicates that modulation of mRNA structural plasticity to polymorphism is likely one contributor to human phenotypic variability.Focusing on a number of cardiovascular disease candidate genes I make a number of novel findings: (1) a strong ACE AEI phenotype common in the African-American population is mapped to specific upstream regulatory alleles and is significantly associated with relevant clinical phenotypes, (2) SOD2 is subject to extremely common and extensive AEI in the human population suggesting potential positive selection, and (3) our results call into question the strength of many previous association studies based on polymorphisms in ACE, CCL2 and NOS3 where there is weak evidence supporting putative functional alleles.