Retinal Organoids from an AIPL1 CRISPR/Cas9 Knockout Cell Line Successfully Recapitulate the Molecular Features of LCA4 Disease

Aryl hydrocarbon receptor-interacting protein-like 1 (AIPL1) is expressed in photoreceptors where it facilitates the assembly of phosphodiesterase 6 (PDE6) which hydrolyses cGMP within the phototransduction cascade. Genetic variations in AIPL1 cause type 4 Leber congenital amaurosis (LCA4), which presents as rapid loss of vision in early childhood. Limited in vitro LCA4 models are available, and these rely on patient-derived cells harbouring patient-specific AIPL1 mutations. While valuable, the use and scalability of individual patient-derived LCA4 models may be limited by ethical considerations, access to patient samples and prohibitive costs. To model the functional consequences of patient-independent AIPL1 mutations, CRISPR/Cas9 was implemented to produce an isogenic induced pluripotent stem cell line harbouring a frameshift mutation in the first exon of AIPL1. Retinal organoids were generated using these cells, which retained AIPL1 gene transcription, but AIPL1 protein was undetectable. AIPL1 knockout resulted in a decrease in rod photoreceptor-specific PDE6 alpha and beta, and increased cGMP levels, suggesting downstream dysregulation of the phototransduction cascade. The retinal model described here provides a novel platform to assess functional consequences of AIPL1 silencing and measure the rescue of molecular features by potential therapeutic approaches targeting mutation-independent pathogenesis.

Automated summary

Aryl hydrocarbon receptor-interacting protein-like 1 (AIPL1) is involved in the assembly of phosphodiesterase 6 (PDE6) in photoreceptors, and genetic variations in AIPL1 cause Leber congenital amaurosis (LCA4) which leads to vision loss. Generating patient-derived LCA4 models for studying AIPL1 mutations may have limitations, therefore, a CRISPR/Cas9 approach was used to create an induced pluripotent stem cell line with a frameshift mutation in AIPL1. This model showed dysregulation of the phototransduction cascade, providing a novel platform for studying AIPL1 silencing and potential therapeutic approaches.