Abstract
Purpose :
Retinitis Pigmentosa (RP) describes a group of retinal dystrophies that are primarily characterized by the loss of rod photoreceptors, followed by degeneration of cones in the retina. Genetic variants in RHO, which encodes the visual pigment of rod photoreceptors are the most common cause of autosomal dominant RP (adRP), accounting for 20-30% of cases. Here, we investigated transcriptional alterations in knock-in mice carrying the P23H, M39R, R135W, and P347L pathogenic amino acid substitutions that are associated with adRP, which have distinct cellular and biochemical consequences on the rhodopsin protein.
Methods :
Retinae from either heterozygous knock-in (KI) mice carrying the rhodopsin mutations P23H, M39R, R135W and P347L or control C57BL/6J mice at 21 days of age (P21) were isolated and snap frozen. Total RNA was isolated and processed for bulk RNA sequencing. Library preparation was strand-specific and poly A selection was used as RNA-selection method. Sequencing was done by Illumina® NovaSeq with a depth of 30M read pairs per sample. DESeq2 was used to normalise the gene hit counts. Genes were considered as differentially expressed genes (DEGs) with adjusted P-value (< 0.05) and an absolute log2 fold change >1. Variation or similarity of gene expression was then confirmed by principal component analysis (PCA) and heatmap of sample-to-sample distance. Gene enrichment analysis was performed by investigating KEGG and Reactome pathways. The R-based tool BEAVR was used to generate heatmaps, PCA, and volcano plots.
Results :
The transcriptome of Rho variant retinae at P21 was compared to age-matched controls and differentially expressed genes were identified across the different rhodopsin variants. More specifically, a total number of 338 (P23H), 519 (M39R), 269 (R135W) and 404 (P347L) genes were found to be significantly differentially expressed. Further analyses of upregulated and downregulated genes using KEGG and Reactome pathway analysis revealed common pathways shared by the distinct rhodopsin models. Furthermore, uniquely affected pathways were identified for each of the four rhodopsin models.
Conclusions :
The transcriptome of four mouse models of adRP were analysed and compared to control C57BL/6 mice. Similarities and differences in the gene expression changes between the different rhodopsin mutations were identified, which could provide insights for shared or distinct disease mechanisms.
This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.