Inherited retinal diseases (IRDs) are a diverse family of disorders which cause vision loss and retinal degeneration. With only 1-2% of the genome being protein-encoding, genetic variation within the expansive noncoding genome is critical to the development of disease phenotypes in the retina. Macular Telangiectasia Type II (MacTel) is an IRD resulting in disruption of central vision and greatly impacting vision-related quality of life. MacTel has an estimated prevalence of 1 in 1000 individuals, affecting approximately two million people globally. Though MacTel etiology largely remains unknown, accumulation of improperly degraded lipids within the retina is a leading hypothesis in its pathogenesis. Additionally, genome-wide association studies have implicated numerous loci in the development of MacTel, including the novel gene locus ceramide synthase 4 (CERS4). As CERS4 plays a critical role in the synthesis of lipid precursors and is highly expressed in the retina, it stands as a promising candidate for influencing MacTel development. We hypothesize that cis-regulatory element (CRE) mutations are central to the genetic frameworks underlying MacTel. We aim to characterize the sufficiency of putative enhancer regions to drive gene expression. We have identified potential CERS4 enhancer regions through a machine learning approach using adult human retina ATAC sequencing datasets. Sufficiency of candidate enhancer regions will be evaluated by insertion to a barcoded reporter library and electroporation into mouse retinas. Following proof of sufficiency, we will perform saturation mutagenesis on identified enhancers to investigate the impact of all possible single nucleotide variants (SNVs) within these regions. The results of our investigation will aid in identifying SNVs of interest within the CERS4 locus, potentially implicating specific mutations towards the development of MacTel. Greater understanding of CRE mutations will improve early clinical diagnosis and inform future therapies for patients with MacTel.

Macular telangiectasia type II is a late-onset retinal degeneration disease which causes loss of central vision and a disruption in cell class proportions in the retina. Genome-wide association studies have identified a point mutation in the 5q14.3 enhancer as associated with MacTel. This enhancer has been shown to regulate the activity of microRNA 9-2. To elucidate the function of this particular enhancer on retinal health and cell class composition, both enhancer knockout and miR9-2 mice models were generated. In adult enhancer knockout mice, there were no significant changes in cell class composition compared with wild-type mice. In the miR9-2 knockout mouse model, it was found that at the 5 week time point, there was a significant increase in müller glial cells. Müller glial cell loss has been observed in Mactel patients, and these cells have been shown to play a crucial role in maintaining proper vascular networks. Future experiments to determine the effects of enhancer and miR9-2 loss on vasculature in the retina would help further identify the role of the 5q14.3 enhancer and its targets on retinal health.