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    Natasha Meyer O'Brown.
    Understanding the molecular and genomic events responsible for adaptation is a major goal of evolutionary genetics. With recent advances in sequencing technology, it has now become possible to compare genomes across taxa and uncover the genetic lesions that differentiate populations and species. One of the most distinctive traits that differs between marine and freshwater sticklebacks is the number of bony lateral plates along their flanks. While marine fish tend to have 32 armor plates, most freshwater fish have reduced this number to 7 or less. This reduction in armor plates has previously been mapped to expression changes in the ectodysplasin (EDA) gene (Colosimo et al, 2005). We now have a complete threespine stickleback (Gasterosteus aculeatus) reference genome, along with several additional genomes from various marine and freshwater populations (Jones et al, 2012). The work in this thesis uncovers an armor plate enhancer that lies downstream of EDA and regulates EDA expression in the developing armor plates. Within this enhancer lies a single base pair change that is shared between all low-plated freshwater fish. Altering this single base in the enhancer changes the domain of enhancer activity by restricting activity to the anterior armor plates that remain in freshwater fish. Furthermore, this thesis goes on to show that the restricted domain of enhancer activity in freshwater fish is due to diminished Wnt responsiveness. Whole genome sequencing projects have also identified genomic differences that separate humans and chimpanzees (Chimpanzee Sequencing and Analysis Consortium, 2005). The rapid and dramatic expansion of the human brain is one of the most distinctive traits that separates us from chimpanzees, yet little is known about the genetic basis for human-specific cortical expansion. Using a comparative genomics approach, our lab previously identified 510 genomic locations where highly conserved regulatory sequences had been specifically lost in the human lineage. One of these locations is a gene rich-region on chimpanzee chromosome 7q22 previously associated with tumor suppressor activity. By cloning the ancestral sequence into a basal promoter-LacZ construct, we have shown that the highly conserved sequence normally serves as an enhancer that drives expression specifically in progenitor cells of the developing forebrain during the critical periods of cortical neurogenesis. In order to functionally test the effects of losing this sequence in the human lineage, we have generated "humanized" knockout mice that are missing the same regulatory sequence. Using RNA-sequencing and ATAC-sequencing to compare knockout to wildtype littermates, I have found that deletion of this enhancer element significantly changes the expression of hundreds of genes, including genes located both nearby and far away from the human deletion, and also alters the local chromatin landscape. The genes with increased expression in mutant embryonic brains are highly enriched for cell division and proliferation functions. I have found that deletion homozygous mice are viable and fertile, and show increased cortical neuron and astrocyte numbers in adult cortex and hippocampus. Gene expression and genetic complementation studies indicate that many of the phenotypes in the knockout mice are due to altered regulation of a gene flanking the deletion that encodes N-acyl phosphatidyl ethanolamine phospholipase D (Napepld). Interestingly, homozygous enhancer knockout mice with increased neuron and glia numbers also show behavioral improvement in memory and learning as measured by the Morris water maze. Collectively, these experiments provide new insights into the genomic and developmental events underlying human brain development and evolution.
    Digital Access   2015