Abstract
Purpose::
To identify genes involved in the pathogenesis of retinal degeneration, we compared the expression profiles of normal and mutant canine retinas. The mutant retinas had a frameshift mutation in RPGR ORF15, with a peak of cell death occurring at 7 weeks of age. Expression changes observed in transcripts classified as tumor suppressor genes (TSG) focused the investigation on their role in photoreceptor loss from inherited retinal degeneration.
Methods::
Normal and mutant dogs retinas at 3, 7, and 16 weeks of age were studied using a canine retinal custom cDNA microarray for gene expression analysis. Gene function was based on GEO website annotation. Northern and western blot analysis was used to confirm expression patterns at the mRNA and protein levels.
Results::
Principal Component Analysis (PCA) showed a significant separation between normal and mutant retinas at each age. Microarray analysis showed that 10 TSG and 25 genes with a putative TSG function are highly expressed in mutant retinas at 7 weeks, and their expression remains elevated at 16 weeks. Northern analysis confirmed the microarray results, and western blots showed increased levels of TSG proteins at 7 weeks of age. In contrast, the normal retina did not have detectable expression levels of TSG at the 3 ages examined.
Conclusions::
In retinas with RPGR ORF15 frameshift mutation, TSG expression increases significantly at 7 weeks of age, a time point when there is a peak in the number of dying cells. The results indicate that TSG might play a role in the regulation of photoreceptor apoptosis and/or survival mechanisms in this inherited retinal degeneration.
Keywords: gene/expression • gene microarray • retina