Conserved Transcriptional Activators of the Xenopus Rhodopsin Gene

Abstract

Vertebrate rhodopsin promoters exhibit striking sequence identities proximal to the initiation site, suggesting that conserved transcription factors regulate rhodopsin expression in these animals. We identify and characterize two transcriptional activators of the Xenopus rhodopsin gene: homologs of the mammalian Crx and Nrl transcription factors, XOtx5 and XL-Nrl (originally named XL-maf), respectively. XOtx5 stimulated transcription ϳ10-fold in human 293 cells co-transfected with a plasmid containing the rhodopsin promoter (؊508 to ؉41) upstream of luciferase, similar to the ϳ6-fold stimulation with human Crx. XL-Nrl stimulated transcription ϳ27-fold in mammalian 293 cells co-transfected with the rhodopsin luciferase reporter, slightly more than the ϳ17-fold stimulation with Nrl. Together, the Xenopus transcription factors synergistically activated the rhodopsin promoter (ϳ140-fold), as well as in combination with mammalian homologs. Deletion of the Nrl-response element, TGCTGA, eliminated the synergistic activation by both mammalian and Xenopus transcription factors. Deletion of the conserved ATTA sequences (Ret-1 or BAT-1), binding sites for Crx, did not significantly decrease activation by Crx/XOtx5. However, there was increased activation by Nrl/XL-Nrl and an increased synergy when the Ret-1 site was disrupted. These results illustrate conservation of mechanisms of retinal gene expression among vertebrates. In transgenic tadpoles, XOtx5 and XL-Nrl directed premature and ectopic expression from the Xenopus rhodopsin promoter-GFP transgene. Furthermore, activation of the endogenous rhodopsin gene was also observed in some animals, showing that XOtx5 and XL-Nrl can activate the promoter in native chromatin environment.Photoreceptors are highly specialized cells with complex structures that permit efficient light absorption, high signal transduction amplification, rapid kinetics, and adaptation over a range of light intensities (1). Phototransduction requires the coordinated expression of many genes, including the visual pigments that absorb light, enzymes involved in the cGMP cascade, ion channels plus multiple regulatory and structural proteins. It has been estimated using serial analysis of gene expression that ϳ4% of the genes expressed in the retina encode phototransduction proteins (2). Many of these genes are quite conserved in vertebrates. Moreover, programs of eye development share many conserved features in the animal kingdom (3, 4). Even in such distantly related species as Drosophila and humans, similar transcription factors are involved in development and expression of retina-specific genes (5, 6), although the evolutionary significance is not yet settled (5, 7, 8). Often, proteins involved in growth and differentiation of the eye from one species can substitute for homologs in distantly related species (4, 7). This conservation also extends to the cis-acting elements in proximal promoters of retinal genes. Promoters have exhibited at least partial functionality between mammals and lower vertebrate...

Homeobox-Clock Protein Interaction in Zebrafish

“…This synergism is somewhat reminiscent of the activation of the rhodopsin promoter in retinal photoreceptor cells by CRX and the leucine zipper-containing factor, neural retina leucine zipper (15), which, most interestingly, physically interact with each other (42). Recently, a similar interaction involving OTX5 and neural retina leucine zipper has been demonstrated in the Xenopus retina (43). This study provides new evidence that OTX5 can synergistically interact with the bHLH clock protein, possibly adding another example of protein-protein interaction involving a photoreceptor-specific homeobox.…”

“…It is therefore possible that the scenario of BMAL/CLOCK-E-box OTX5-PCE interaction is important for the expression of many additional genes that genetically define the pineal gland and are essential for pineal function. Moreover, in addition to other identified transcriptional mechanisms discussed above (15,42,43), this mechanism may regulate gene expression in retinal photoreceptors.…”

Retinoic Acid Regulates the Expression of Photoreceptor Transcription Factor NRL

“…NRL 3 is a bZIP transcription factor of the Maf subfamily (22). NRL is conserved in vertebrates and is specifically expressed in photoreceptors and pineal gland (23)(24)(25)(26). Loss of Nrl in mice results in functional S-cones that are derived from post-mitotic precursors normally fated to be rods (25,27).…”

Retinopathy mutations in the bZIP protein NRL alter phosphorylation and transcriptional activity

“…NRL orthologs have been identified in many vertebrates with the exception of chicken [Coolen et al, 2005;Whitaker and Knox, 2004]. To date, 17 mutations and/or variations in the NRL gene have been detected [Bessant et al, 1999;DeAngelis et al, 2002;Martinez-Gimeno et al, 2001;Nishiguchi et al, 2004;Wright et al, 2004;Ziviello et al, 2005]; these include 14 missense and three frameshift mutations (Fig.…”