GENE EXPRESSION IN RETINAL DEVELOPMENT, DEGENERATION, AND DIFFERENTIATION

RNA-seq is a next generation sequencing based method for transcriptome wide characterization of gene expression. This dissertation explores the application of RNA-seq, both at the level of the tissue and at the level of the single cell, to the study of cells of the retina in physiologic and pathologic contexts. Fluorescence activated cell sorting and droplet microfluidic based methods for single cell RNA-seq were used to explore differentiating human stem cell derived retinal ganglion cells and murine photoreceptors undergoing normal postnatal development and inherited retinal degeneration. Population level RNA-seq on peripheral blood mononuclear cells derived from patients with age-related macular degeneration (AMD) and control individuals was performed to identify novel blood-based biomarkers for AMD. A draft retinal transcriptome for the Nile rat (Arvicanthis ansorgei), a diurnal organism with a cone-enriched retina, was assembled and annotated. Gene expression perturbations and enriched pathways in dissociation induced epithelial to mesenchymal transition (EMT) in human stem cell derived retinal pigment epithelium (RPE) were characterized. A high throughput biochemical assay to screen for small molecule inhibitors of dual leucine zipper kinase (DLK), a key driver of cell death in retinal ganglion cells, was developed and implemented. This dissertation is unified by the common theme of development and utilization of cutting edge tools at the intersection of molecular biology and bioinformatics to further our collective understanding of retinal development, differentiation, and degeneration