Functional studies and population genetic studies of a 25kb haplotype upstream of the AKT1 gene associated with metabolic syndrome

We report here the identification of a highly conserved haplotype, denoted H1 (ancestral) and H2 (derived), of three SNPs in high linkage disequilibrium present upstream of the AKT1 gene and within the coding region of a novel zinc finger gene. Haplotype H2 is associated with lower insulin resistance, lower fasting glucose levels and a decreased risk for the development of metabolic syndrome in American populations. In this dissertation we report the characterization of this 12kb haplotype in terms of a novel transcript unit (Z-FAMS) within the 12kb H1/H2 region, effects of the constituent polymorphisms on gene expression, and population genetics of the H2 haplotype. We characterized the putative zinc finger gene, Z-FAMS , and showed that it is transcribed and translated in human skeletal muscle where it localizes to the nucleus. SNP rs10141867 of H2 is present within the Z-FAMS coding sequence. Through a series of in vitro reporter assays, we showed that the H2 allele of this SNP has strong enhancer effects in mouse muscle cells conferring a 15.7-fold induction. Using a modified allele-specific expression assay in human skeletal muscle we showed that the Z-FAMS transcript containing the H2 sequence was overexpressed by 1.75 fold over the H1 sequence. The high frequency of the intact H2 haplotype in diverse American populations and its strong association with insulin resistance phenotypes suggested that this region may have been subject to positive selection. The 18kb region harboring the 12kb H1/H2 haplotype is flanked by regions of high recombination (10-15%). To query a model of positive selection in a small insulated genomic region we conducted population genetic studies of the H2 haplotype in 60 isolated global populations, based on a model of haplotype diversity and frequency. Over 95% of all global populations expressed the H2 haplotype at varying frequencies, ranging from 8%-54%. Using the program Network we built haplotype trees for the H2 region and control regions on global populations. The diversity of haplotypes in the H1/H2 region was very low, ranging from 0%-5.2%, compared to control regions. The research described in this thesis is significant for characterizing the novel transcript unit Z-FAMS, for defining functional consequences of SNPs included in a haplotype associated with metabolic syndrome, and for beginning to describe a model where positive selection may occur in regions flanked by high recombination.