Abstract
Abstract: :
Purpose: Retinal pigment epithelium (RPE) differentiation involves reciprocal expression of N– and R–cadherin cell adhesion molecules, a pattern that is recapitulated during RPE wounding and subsequent healing. We hypothesize that perturbation of the normal cadherin pattern may cause aberrant RPE phenotypes such as may occur in proliferative vitreoretinopathy (PVR). Utilizing a bioinformatics approach to analyze transcriptional regulatory elements (TREs), the present study was carried out to begin to elucidate the genetic regulatory network by which cadherin subtype switching is regulated in the RPE in the context of broader patterns of coordinated gene regulation during RPE differentiation. Methods: A group of eighty genes, whose expression is altered during RPE wound healing, RPE epithelial–mesenchymal transformation, or PVR, including R– and N–cadherin, were identified in our prior studies or from the published literature. Using the analysis tool PAINT (Promoter Analysis and Interaction Network Tool; www.dbi.tju.edu/dbi/tools/paint/), cognate promoter sequences were retrieved. PAINT contains a database of known or predicted promoter sequences cross–referenced with gene identifiers. Promoters were analyzed to identify TREs for each gene. Candidate regulatory networks were then identified via statistical analysis of TRE occurrence within the gene cluster relative to randomly selected gene lists. Results: The PAINT program exhaustively identified potential TREs, and produced a candidate interaction matrix based on TREs shared between genes within the set. The genes were grouped into fourteen clusters based on TRE profile similarity. Preliminary analysis indicated a trend towards clustering of genes that were coordinately up– or down–regulated. PAINT identified individual TREs that were highly enriched among the promoters analyzed, including Oct1, HNF3alpha, E2F, CDP and SP3, suggesting that these transcription factors may be especially important in coordinating RPE gene expression, with HNF3alpha, SP3 and CDP also specifically found in the cadherin promoters. Conclusions: This computational analysis has identified a specific subset of TREs that may be of importance in regulation of RPE cell differentiation and wound repair in general, and for cadherin regulation in particular. Further studies will be focused on evaluating the role of the transcription factors in cadherin gene regulation in the RPE. These regulatory proteins are potential therapeutic targets for diseases such as PVR. Grant support: NIH RO1EY06658, NIH T32ES07282, and DARPA F30602–01–2–0578.
Keywords: retinal pigment epithelium • transcription factors • gene/expression