Abstract
Purpose :
In humans, a large deletion in a non-coding DNA region upstream of Atoh7 has been associated with NCRNA, characterized by optic nerve atrophy, retina detachment, and complete blindness. Atoh7 is a bHLH transcriptional factor required for retinal ganglion cell (RGC) differentiation in mice. Given the phenotypic similarity between NCRNA disorder and Atoh7 inactivation in mice, it was proposed that this non-coding genomic region acts as an enhancer that governs Atoh7 expression. However, previous data have indicated that the deleting of the orthologous mouse enhancer does not phenocopy NCNRA, and RGCs appear to be minimally affected in adult retina. Thus, the functional relevance of the Atoh7 enhancer landscape in mice and humans remains poorly understood, and a mouse model that recapitulates NCNRA disease is yet to be generated. This information is necessary for understanding the pathological manifestations of NCRNA and for improving RGC generation paradigms from stem cells. The purpose of this work is to build a mouse model that recapitulates NCRNA.
Methods :
We used epigenetics profiling to characterize the Atoh7 cis-regulatory landscape dynamics during retinal development in mice in detail. We further utilized CRISPR/CAS9 technology to generate a mouse carrying a homozygous deletion in the enhancer landscape upstream of Atoh7. We carried out histological, electrophysiological and transcriptomic analyses to assess structural and functional defects in adults and embryos from mice carrying the homozygous deletion and matching controls.
Results :
Mouse carrying a homozygous deletion in Atoh7 enhancer landscape and viable and fertile but electrophysiological analysis indicates a strong reduction in A- and B- wave amplitude upon light stimuli. Histological analysis revealed structural defects, including retinal detachment, reduced retina thickness, disruption of the laminar structure, absence of optic stalk and optic nerve, and blood vasculature defects. Further, immunostaining analyses in the adult and embryonic retina of the mutant mice underscore a strong reduction in RGC numbers. Transcriptome analysis of the developing retina revealed significant dysregulation of genes important for RGC genesis.
Conclusions :
Our data reveal that the Atoh7 enhancer landscape is essential for RGC genesis and axonogenesis and indicates a successful generation of a mouse model that recapitulates key features of NCRNA.
This abstract was presented at the 2024 ARVO Annual Meeting, held in Seattle, WA, May 5-9, 2024.