Date of Award
Doctor of Philosophy (PhD)
Precisely coordinated expression of distinct sets of genes is essential for cellular development and function, especially in complex multicellular organisms. This regulation is achieved by the action of transcription factors (TF), proteins that bind specific genomic locations and alter the activity state and packaging of the DNA to promote or repress gene expression. However, while tremendous effort has defined networks of transcription factors that work together to drive specific phenotypes, little is known about their differential activity at the hundreds or thousands of sites where they bind. There are also many questions regarding the basic principles of the packaging of DNA within the nucleus, its influence on gene expression, and how transcription factors regulate this process.
To address these questions, I have investigated gene regulatory mechanisms that control the development of rod and cone photoreceptors. Photoreceptors are the neurons of the retina responsible for the initial conversion of a visual stimulus into an electrical signal. Photoreceptors are a complex, accessible, and highly disease relevant neuronal cell population, the lessons from which are relevant for many other cell types across the body. A primary TF in photoreceptor development is CRX. This TF when mutated in humans can cause severe vision loss, and its deletion in mice leads to a severe condition with no functional photoreceptors.
First, I determined the functional consequences of human disease-causing variants when modelled in mice. I functionally classified a new model as causing a previously un-determined dominant disease, and discovered that all CRX diseases are the result of graded differences in gene expression of the same core set of genes. Second, I examined the dependency of all genomic binding sites on CRX activity and identified a core set of regulatory elements that require CRX for activation of the target genes. Third, I interrogated the broader organization of the rod photoreceptor epigenome and demonstrated that the rod packages its DNA according to epigenomic activity state. Perturbations to this state can override this organization, resulting in functional consequences on gene expression beyond the local sequence.
In summary, my in-depth investigation has uncovered new insights into the molecular mechanisms controlling development and maintenance of the rod epigenome and its organization in the nucleus. This new knowledge will provide 1) a new understanding of where and how mutations in a key photoreceptor TF cause gene mis-regulation, and 2) guidance for future human genetic studies to identify new disease-causing mutations affecting photoreceptor integrity, not only in the protein coding sequences but also in specific non-coding regulatory regions.
Chair and Committee
Joseph Corbo, Kristen Kroll, Eugene Oltz, Ting Wang,
Ruzycki, Philip Andrew, "Deciphering mechanisms governing the development of the rod epigenome" (2018). Arts & Sciences Electronic Theses and Dissertations. 1573.