Abstract
Abstract: :
Purpose:X–linked juvenile retinoschisis is caused by mutations in the RS1 gene encoding a 24–kDa protein, termed retinoschisin, which is secreted from photoreceptor and bipolar cells as a disulfide–linked oligomeric complex. Although loss of retinoschisin function has been established as the underlying disease mechanism, little is known about the molecular events leading to the loss of cellular architecture in the neural retina. Methods:Mice hemizygous for a disrupted Rs1h gene were used to isolate RNA samples and protein extracts from dissected retinae. Custom made and commercial microarray chips containing reference sets of non–redundant murine transcripts were available. Proteome analysis was done by 2D–gel electrophoresis and protein de novo sequencing of conspicuous spots with ESI/MS/MS. Results:For transcriptome analysis, we have analysed retinal RNA extracts from postnatal day 5, 10, 14 and 21. Over this time period, transcription levels from Rs1h–/– retinae significantly deviate from wildtype for a great number of genes including cone opsin, Rom1, Gnat2, Casp1, Casp8, Fadd and others. This likely reflects a burst of apoptosis observed around P18 within the outer nuclear layer resulting in a major loss of rod and cone photoreceptor cells. Proteom analysis on 2D–gels from retinal protein extracts of 2 months old Rs1h–/– and wildtype retinae revealed several protein spots with differential expression in the diseased versus normal retina. ESI/MS/MS sequencing is in progress to identify the identity of these proteins. Conclusions:Transcriptome and proteome analysis are powerful approaches to investigate the molecular effects on the transcriptional as well as the translational level in an in vivo model of Rs1h deficiency.
Keywords: retinal degenerations: cell biology • gene microarray • proteomics