Defining the mechanism of photoreceptor cell death in Retinitis Pigmentosa

Retinitis pigmentosa (RP) is a multigenic, untreatable inherited retinal dystrophy (IRD). Mutations in over 100 genes lead to death of the light sensing photoreceptors in the retina, resulting in night blindness, visual field constriction and eventual total visual loss. The IRDs are the leading cause of visual loss in children and working adults and have immense economic and well-being costs associated with them. Although gene therapy has emerged a potential therapeutic, improvements are minimal and slow-moving, as each gene must be targeted one at a time.
It has been postulated that the mechanism through which photoreceptors die in RP is common to all disease-causing alleles. If so, understanding this pathway could lead to a therapeutic target that would be applicable for all forms of disease. It has, however, yet to be fully defined. This project will adopt a 4D cellular medicine approach to determine the transcriptional changes that occur in photoreceptors with single cell resolution as photoreceptors move from health to stress to death in RP. To do so, we will use two different murine models of RP; each with a mutation in a gene involved in a different pathway required for photoreceptor maintenance and function. The Atrd1 mouse harbours a mutation in Pde6b; its protein is crucial for the visual cycle. The Atrd1 mouse develops a fast photoreceptor degeneration over 6 weeks. RPGR appears to be involved in photoreceptor outer segment maintenance / turnover. The Rpgr-mutant mouse develops a slow photoreceptor degeneration over 24 months. By comparing transcriptional changes common to both mice, it is hoped we will identify a pathway to be targeted to rescue photoreceptor loss in RP.
Hypothesis: There is a shared common mechanism of photoreceptor death across multiple models of RP.
This project has 3 aims:
- To fully characterise the retinal degeneration seen in the Atrd1 mouse, a model for Pde6b-
mediated RP.
- To define the underlying mechanism of cell death that leads to photoreceptor loss in the
Atrd1 mouse.
- To functionally integrate Atrd1 datasets with those of the slow progressing RP RPGR model
to identify whether common mechanisms exist.
In this multidisciplinary project, a combination of both laboratory and computational techniques will be applied. The Atrd1 mouse will comprehensively characterised in the MRC HGU's eye phenotyping suite under the supervision of Dr Roly Megaw, to map changes in retinal function (electroretinography, optokinetic drum) and structure (optical coherence tomography) over time. The Atrd1 is a fast model of retinal degeneration, with photoreceptor loss occurring over weeks.
Following this, single cell sequencing experiments will be performed on dissociated retinas at early (P14) and late (P28) timepoints. Datasets will be analysed under the supervision of Dr Catalina Vallejos, with the aim of defining pathway changes in mutant photoreceptors as they die. Further, invading glial and circulating monocytic cells will be interrogated. Results will be compared to scSeq data (12 and 24 month timepoints) of a slow degeneration model of RP-disease (Rpgr). Mechanisms of cell death and retinal invasion will be confirmed with focussed qPCR, spatial transcriptomic methods such as RNAscope, as well as proteomics methods and imaging experiments. Identified pathways will subsequently be targeted to rescue the photoreceptor degeneration seen in our mutant models.
The objective of this PhD project is a better understanding of the mechanisms of photoreceptor cell death to develop novel and much needed treatment options by identifying targets that could rescue or slow down retinal degeneration in all forms of RP.