Selection of candidate genes implicated in cone survival by photoreceptor transcriptome analysis

Abstract

Purpose : Death of rod photoreceptors leads to cone cell degeneration, suggesting an indispensible role of rod cells in cone survival. Surprisingly, in Nrl^-/- rod-free retina, functional cones survive for an extended period of time, indicating that other factors also contribute to the maintenance of cone cells. We hypothesize that direct (cell-cell) or indirect (secreted molecule) interactions between cone photoreceptors and other cell types play an essential role in cone cell survival in Nrl^-/- retina.

Methods : By comparing the gene expression profiles of flow-sorted GFP+ cells in Nrl-GFP:Nrl^-/- (S-cone-like cells) and Nrl-GFP:Nrl^+/+ (rod photoreceptors), we selected genes that are at least 2-fold overexpressed in S-cone-like cells (false discovery rate ≤ 0.01) at postnatal day (P) 6, P10, P14 and P28. These candidate genes were filtered for those encoding membrane and secreted proteins according to their Gene Ontology annotation. To identify novel targets for treatment development, we mainly focused on genes having homologs in human and not well characterized in mouse retina. Expression patterns of the selected genes in Nrl^+/+ and Nrl^-/- mature retina (P28) were then investigated by In situ hybridization to confirm the RNA-seq data.

Results : We identified 2633 genes significantly overexpressed (fold change ≥ 2 and false discovery rate ≤ 0.01) in Nrl-GFP:Nrl^-/- photoreceptors compared to Nrl-GFP:Nrl^+/+. Among them, 901 genes encode for membrane or secreted proteins. Genes with unknown functions and those implicated in cell survival, according to their roles in other organs, were selected. After prioritizing the genes based on their expression level in Nrl-GFP:Nrl^-/- retina and the existence of a human homolog, we finalized a list of 21 candidates to focus on. In situ hybridization confirmed overexpression of 16 genes at P28 in Nrl^-/- compared to Nrl^+/+. All genes were enriched in photoreceptor layer, with 5 mainly absent from other nuclear layers, suggesting a possible role in cone photoreceptor development, function or survival.

Conclusions : Our study provides candidate genes implicated in survival of cone photoreceptors, which may be targets to develop treatments for retinal degenerative diseases. We are currently investigating functions of these genes in the retina by gain- and loss-of-function experiments.

This is an abstract that was submitted for the 2017 ARVO Annual Meeting, held in Baltimore, MD, May 7-11, 2017.