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W. Yan, Y. Chen, R.L. Bradford, C. Wang, R. Adler, D. Zack; The COUP–TFs as Potential Modulators of Cone Visual Pigment Gene Expression . Invest. Ophthalmol. Vis. Sci. 2004;45(13):3635.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose:To identify and characterize transcription factors that bind to and regulate cone opsin promoters. Methods:A retinal cDNA one–hybrid library was constructed by directionally inserting retinal cDNA from cone–dominated 13–lined ground squirrel (13–lgs) into the pGADT7 vector. Several potential cis–elements in the proximal promoter of 13–lgs green opsin were used as baits for yeast one hybrid screening, carried out by standard methods. For transcriptional activity assays, HEK293 cells were transiently transfected with expression plasmids for COUP–TF1, COUP–TF2, Crx or Nrl, along with luciferase reporter constructs driven by the proximal promoters (–500, –227, –190 and –89 to +40) of human green, red or blue opsin. Analysis of the expression patterns and cellular localization of COUP–TF2 was done in 13–lgs and chick embryos by RT–PCR and in situ hybridization. Results:Chicken Ovalbumin Upstream Promoter Transcription Factors–1 and 2 (COUP–TF1/2) were among the transcription factors identified in the one–hybrid screen. COUP–TF1 and 2 were found to repress the transcriptional activity of green and red opsin promoters in transient transfection reporter assays, but had only minimal effect on a blue opin promoter. In addition, COUP–TFs inhibited the ability of Crx and Nrl to trans–activate the green opsin promoter. In the cone dominant 13–lgs retina, in situ hybridization showed preferential and strong COUP–TFs expression in cones. In the chick, RT–PCR detected the COUP–TF2 ortholog at all the developmental stages studied [embryonic day (ED) 5, 6, 8, 12, 15, 16, 17 and 18]. Ongoing studies are investigating the cellular expression patterns of COUP–TF2, as well as the effects on its expression of agents that influence visual pigment expression, such as GDNF, staurosporine, CNTF and activin, are being investigated. Conclusions:Those results suggest that COUP–TF1 and 2 may be involved in modulating visual pigment gene expression in cone photoreceptors, and may play a role in regulating cone development.
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