Non-Coding Genetic Vulnerabilities in Human Photoreceptor Function and Disease

PROJECT SUMMARY/ABSTRACT Cis-regulatory elements (CREs) are critical sites within the non-coding genome that orchestrate the expression of genes necessary for normal cellular function. Mutations within CREs contribute to human diseases including disorders of vision. Over the past funding period, our group identified and characterized CREs in the adult and developing retina using single-cell, epigenomic, machine learning, and genetic approaches and used these data to elucidate genetic causes of unsolved visual disorders. Despite this progress, the function of most of retinal CREs and the features that define essential CREs remain unknown. This gap in knowledge is a significant obstacle toward understanding the genetic regulation of normal human vision, identifying disease-causing mutations, and developing corrective strategies. The long-term goal for our research is to decipher how non- coding genetic variation shapes the structure, function, and diseases of the retina and to advance therapeutic approaches. The focused objective of this proposal is to improve high-throughput approaches to study retinal CRE function at scale, to identify CREs that are necessary for retinal photoreceptor development, function, and survival, and to identify predictive features of essential CREs. The central hypothesis driving this work is that higher-throughput techniques will identify essential CREs and will reveal the critical features that impart their function. To test this, we have developed a multiplexed approach to disrupt multiple retinal CREs in parallel and measure the effect on gene expression, cell state, and survival. We complement this approach by using a convolutional neural network-based machine learning approach to predict CRE function. Lastly, we are utilizing powerful new genetic models to identify CREs that control the timing of human and mouse retinal development. The contribution of this research will be to elucidate the features and mechanisms by which CREs regulate genes that are necessary for human photoreceptor function and survival. This will enable the systematic identification and interpretation of disease-associated variants within CREs and improve genetic diagnostics for retinal disease. By opening up the non-coding genome to functional analyses, it will be possible to develop new therapeutic strategies for correcting or preventing inherited retinal diseases.