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S. J. Binek, D. Choi, J. Stout, J. T. Rosenbaum, B. Appukuttan, J. R. Smith; Comparison of Cis-Regulatory Elements in Genes Differentially Expressed by Retinal and Choroidal Vascular Endothelial Cells. Invest. Ophthalmol. Vis. Sci. 2007;48(13):5180.
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© ARVO (1962-2015); The Authors (2016-present)
Retinal and choroidal vascular beds are involved in distinct ocular diseases. Using gene expression microarray, we previously showed that endothelial cells (ECs) isolated from human retina and choroid show significant differences in levels of expression of approximately 10% of genes. We hypothesized that differential gene expression reflected differences in cellular abundance of key transcription factors (TFs).
We utilized a gene expression data set generated for retinal versus choroidal ECs from 3 cadaver donors using Affymetrix Human Genome Focus Arrays (detecting 8746 transcripts). Transcripts were identified on the basis of significantly different expression by SAM with false discovery rate of 5%, unidirectional change across all donors, and 2-fold difference in level of expression between cell types. Sequences of cis-regulatory motifs within 2000 base pairs upstream of the start codon corresponding to transcripts of interest were identified using TRANSFAC Pro (www.biobase-international.com/pages/). Subsequently PAINT (Promoter Analysis and Interaction Network Toolset, www.dbi.tju.edu/dbi/tools/paint/) was employed to determine abundance of these motifs in genes differentially expressed by the ocular ECs versus the entire genome. In addition, abundance of motifs was compared for genes differentially expressed in retinal and choroidal ECs.
Analysis of the gene expression data set yielded 74 and 67 transcripts that were expressed at increased levels in retinal and choroidal ECs, respectively; 209 cis-regulatory motifs were identified within the corresponding promoters. Differentially expressed genes of ocular ECs contained an over-representation of Nrf2, VBP, MAZ, UF1H3BETA, HSF1 and E2F in comparison to the complete genome. When abundance of the motifs was compared between retinal and choroidal ECs, CP2, MRF-2 and Ik-1 were over-represented in 16 genes that were more highly expressed in retinal ECs, and Lentiviral Poly A was over-represented in 8 genes that were more highly expressed in choroidal ECs.
Our data indicate that certain cis-regulatory elements are over-represented within the promoter of genes that are differentially expressed by retinal versus choroidal ECs. This observation is consistent with the concept of vascular EC diversity and suggests that differences in gene expression may relate to differential cellular abundance of certain TFs. Future studies will focus on identification of cell-specific TFs.
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