Abstract
Purpose :
CRX is a transcription factor (TF) essential for photoreceptor functional development and maintenance with an important role in the terminal differentiation of photoreceptors. Missense mutations in CRX homeodomain (HD) are linked to variable human retinopathies. We previously reported a murine model carrying the loss-of-function mutation p.R90W that causes recessive Leber Congenital amaurosis (LCA). This study aims to characterize the retinal phenotypes of new murine models for distinct dominant CRX HD mutations, p.E80A and p.K88N.
Methods :
Knockin models were generated by introducing E80A or K88N mutations into the Crx locus. Hematoxylin and eosin (H&E) staining of retinal cross-sections revealed morphological defects at various postnatal (P) ages. Immunostaining of cell-type specific markers reported altered localization and relative abundance of affected cell types. Electroretinography (ERG) detected deficits in visual functions. Quantitative PCR (qPCR) determined differential gene expression of CRX-target genes and photoreceptor-specific TFs.
Results :
New models showed retinal morphologies different from those found in published animal models. Irregular whorls and rosettes were detected within the outer nuclear layer of all E80A and K88N mutants at P14, worse in homozygotes than in heterozygotes. This phenotypic abnormality appeared within the window of photoreceptor differentiation and the severity progressed over the time of photoreceptor development. At P30, homozygotes of both mutations failed to develop rod outer segments and differentiated cones, thus showing null ERG responses. K88N heterozygotes also showed null ERG responses. E80A heterozygotes had shorter rod outer segments and few detectable cones, resulting in significantly decreased A- and B-wave amplitudes in rod ERG responses and undetectable cone responses. Gene expression analysis at P21 found that overall Crx levels in E80A and K88N mutants were comparable to that in wildtype control, but the expression of CRX-dependent phototransduction components were downregulated.
Conclusions :
The murine models of E80A and K88N mutations display dominant Cone-rod dystrophy and LCA-like phenotypes respectively, which match corresponding clinical features in human patients. These models help to understand the underlying cellular and molecular mechanisms of dominant CRX HD mutations, informing treatment windows and strategy designs.
This abstract was presented at the 2022 ARVO Annual Meeting, held in Denver, CO, May 1-4, 2022, and virtually.