Comparative Analysis of Global RNA Expression Profiles in the Retinas of Wild Type and Rho–/– Mice

Abstract

Abstract: : Purpose:To obtain an insight into the processes of the degenerating retina by analysing the expression levels of over 30,000 transcripts in mice lacking the rhodopsin gene (Rho–/–), over 8 time–points. Methods:Samples were obtained from Rho–/– 129Sv and wild–type 129sV mice at postnatal days 10, 15, 20, 25, 30, 35, 40 and 60 and analysed using the complete Affymetrix mouse MG–U74 chip set. Each pooled Rho–/– sample was compared to a pooled wild–type sample of the same age using the Affymetrix software MAS 5.0, MDB 3.0 and DMT 3.0. Hierarchical cluster analysis was performed using the EBI EPCLUST software. Real–time PCR and in situ hybridisation studies were used to verify and further investigate array results. Results:Due to the absence of functional rhodopsin in the Rho–/– mouse retinas, the rod cells progressively degenerate over the first 3 months of life. The current study examines genome wide expression in the retina over a critical phase of this degeneration and compares the expression levels to those of wild type mice at the same ages. A large degree of variation was observed in all time points analysed, with genes showing both up and down–regulation in the Rho–/– retina. 3,536 genes, or ESTs, showed changes in 3 or more of the 8 timepoints. Of these, over 200 show significant changes (exclusion limit p<0.05) of greater than 2–fold in at least 3 of the time–points, suggesting involvement of their encoded proteins in disease pathology. A number of these genes have previously been shown to have an involvement in retinal processes, while many have as yet unidentified roles in the retina. Cluster analysis has revealed several functionally related groups that are changed in distinct patterns, including chaperones, immune response and photoreceptor enriched genes. Conclusion:The changes in retinal gene expression identified in this study will help to delineate the fundamental biological processes which occur over the course of the degeneration in the Rho–/– mice, and may point to previously uncharacterised roles in the retina for some of the genes identified in this study.

Keywords: retinal degenerations: cell biology • gene microarray • transcription