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Jerome Roger, Avinash Hiriyanna, Debbie Cheng, Norimoto Gotoh, Rinki Ratna Priya, Matthew Brooks, Harsha Rajasimha, Bo Chang, Anand Swaroop; Long-term preservation of immature cone-like photoreceptors in a mouse model of human LCA caused by dominant CRX frameshift mutation. Invest. Ophthalmol. Vis. Sci. 2013;54(15):3150.
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© ARVO (1962-2015); The Authors (2016-present)
Leber’s congenital amaurosis (LCA) is a severe form of childhood blindness representing 5% of all retinopathies. Mutations in CRX, a key transcription factor for photoreceptor development, are associated with retinal dystrophies including LCA. The pathogenic mechanisms of CRX disease have not been elucidated, and the reported Crx-/- mouse model does not reflect the human phenotype. Here, we describe a spontaneous mutant with autosomal dominant congenital blindness named RIP (Retina with Immature Photoreceptors), caused by a frameshift mutation in Crx leading to the differentiation of retina with immature cone-like photoreceptors.
Linkage analysis and exome capture sequencing were used to identify the mutation. Fundus photography, ERG, immunohistochemistry, RNA-seq, luciferase and gel-shift assays were performed to characterize the RIP mouse and the mutant CRX protein. CRX and frameshift mutants were transfected in mouse retina by in vivo electroporation. Crxp::Nrl mice were used for some of the rescue experiments.
The RIP mutant revealed autosomal dominant pattern of inheritance with congenital blindness and exhibited pigmentation and attenuated vasculature upon fundus examination at one-month of age. We identified a frameshift mutation in Crx resulting in the deletion of the Otx-like domain and an extended C-terminal region. Interestingly, several LCA-causing CRX mutations result in similar protein modifications. In the RIP mutant, the retinal photoreceptors had cone-like nuclei, but no outer segments; however, unlike Crx-/- mice the photoreceptors did not degenerate. RNAseq analysis of P2 and P21 retina revealed a loss of Nrl and Nr2e3 expression and an absence of most rod visual signaling components only at P21. We also showed the lack of transcriptional activity in CrxRIP was due to its inability to bind DNA. By mating RIP mutant with Crxp-Nrl mice in which Nrl is expressed in all photoreceptor precursors, we partially rescued the phenotype due to the existence of only rods with short outer segments. Electroporation in wild type mice of LCA-causing CRX frameshift mutations led to an arrest of photoreceptor maturation.
Our findings show the dominant negative effect of a group of CRX mutations preventing the establishment of rod gene regulatory network and reveal the RIP mouse as a good model for LCA-causing CRX mutations.
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