| Title: |
CRISPR-Cas9 correction of a nonsense mutation in LCA5 rescues lebercilin expression and localization in human retinal organoids |
| Authors: |
Afanasyeva, TAV; Athanasiou, D; Perdigao, PRL; Whiting, KR; Duijkers, L; Astuti, GDN; Bennett, J; Garanto, A; van der Spuy, J; Roepman, R; Cheetham, ME; Collin, RWJ |
| Source: |
Methods & Clinical Development , 29 pp. 522-531. (2023) |
| Publisher Information: |
Elsevier BV |
| Publication Year: |
2023 |
| Collection: |
University College London: UCL Discovery |
| Subject Terms: |
retinal organoid; retinal disease; Leber congenital amaurosis; CRISPR-Cas9; isogenic; lebercilin; whole-genome sequencing; LCA5 |
| Description: |
Mutations in the lebercilin-encoding gene LCA5 cause one of the most severe forms of Leber congenital amaurosis, an early-onset retinal disease that results in severe visual impairment. Here, we report on the generation of a patient-specific cellular model to study LCA5-associated retinal disease. CRISPR-Cas9 technology was used to correct a homozygous nonsense variant in LCA5 (c.835C>T; p.Q279∗) in patient-derived induced pluripotent stem cells (iPSCs). The absence of off-target editing in gene-corrected (isogenic) control iPSCs was demonstrated by whole-genome sequencing. We differentiated the patient, gene-corrected, and unrelated control iPSCs into three-dimensional retina-like cells, so-called retinal organoids. We observed opsin and rhodopsin mislocalization to the outer nuclear layer in patient-derived but not in the gene-corrected or unrelated control organoids. We also confirmed the rescue of lebercilin expression and localization along the ciliary axoneme within the gene-corrected organoids. Here, we show the potential of combining precise single-nucleotide gene editing with the iPSC-derived retinal organoid system for the generation of a cellular model of early-onset retinal disease. |
| Document Type: |
article in journal/newspaper |
| File Description: |
application/pdf |
| Language: |
English |
| Relation: |
https://discovery.ucl.ac.uk/id/eprint/10171401/ |
| Availability: |
https://discovery.ucl.ac.uk/id/eprint/10171401/1/1-s2.0-S2329050123000748-main.pdf; https://discovery.ucl.ac.uk/id/eprint/10171401/ |
| Rights: |
open |
| Accession Number: |
edsbas.EF134731 |
| Database: |
BASE |