Opsin expression in the rat retina is developmentally regulated by transcriptional activation.

Treisman, Jessica E.; Morabito, Maria A.; Barnstable, Colin J.

doi:10.1128/mcb.8.4.1570

Cited by 88 publications

(58 citation statements)

References 50 publications

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“…2B) expressed exclusively in rod photoreceptors. The former started to be detected at around P4 or P5, whereas the onset of the latter was slightly earlier (at around P2-P3), consistent with the expression profiles of rhodopsin and Nrl (20)(21)(22). Crx promoterDsRed was detected in bipolar cells at a high level and in photoreceptors at a very low level (Fig.…”

Section: Resultssupporting

confidence: 60%

Controlled expression of transgenes introduced by in vivo electroporation

Matsuda

Cepko

2007

Proc. Natl. Acad. Sci. U.S.A.

615

601

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In vivo electroporation is a powerful technique for the introduction of genes into organisms. Temporal and spatial regulation of expression of introduced genes, or of RNAi, would further enhance the utility of this method. Here we demonstrate conditional regulation of gene expression from electroporated plasmids in the postnatal rat retina and the embryonic mouse brain. For temporal regulation, Cre/loxP-mediated inducible expression vectors were used in combination with a vector expressing a conditionally active form of Cre recombinase, which is activated by 4-hydroxytamoxifen. Onset of gene expression was regulated by the timing of 4-hydroxytamoxifen administration. For spatial regulation, transgenes were expressed by using promoters specific for rod photoreceptors, bipolar cells, amacrine cells, Mü ller glia or progenitor cells. Combinations of these constructs will facilitate a variety of experiments, including cell-typespecific gene misexpression, conditional RNAi, and fate mapping of progenitor and precursor cells.G ain-of-function and loss-of-function studies using transgenic animals have greatly advanced our understanding of the molecular and cellular mechanisms of development and disease. In particular, conditional gene activation and inactivation have been powerful methods for studies of gene function and for the labeling and manipulation of specific cell populations in vivo (1, 2). Site-specific recombination systems (Cre/loxP and Flp/FRT) are widely used to control gene expression in transgenic mice. By crossing two transgenic lines, one expressing Cre (or Flp) under tissue-specific and/or inducible control and the other carrying two loxP (or FRT) sites, DNA recombination can lead to inducible gene expression or loss of gene expression in restricted tissues at specific times (3). Although such strategies are very useful, they are time-consuming and cannot be applied to species that are difficult to manipulate genetically.In vivo electroporation is a convenient technique for the introduction of genes into a variety of animals, including mouse, rat, and chick (4, 5). We previously reported that plasmid DNAs can be easily delivered to developing mouse/rat retinas by in vivo electroporation (6). The plasmid DNAs, electroporated from the scleral side of the postnatal day 0 (P0) retina, are preferentially introduced into retinal progenitor/precursor cells, which give rise to four different cell types; rod photoreceptors in the outer nuclear layer (ONL), bipolar and amacrine cells in the inner nuclear layer (INL), and Müller glial cells, which extend radial processes spanning the entire thickness of the retina [see supporting information (SI) Fig. 7]. The efficiency of electroporation into the developing postnatal retina is quite good, and transgene expression persists at least for a few months. Moreover, compared with other gene transfer methods, such as viral vectors, in vivo electroporation has several advantages. First, various types of DNA constructs, including RNAi vectors, are readily introduced to th...

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Section: Resultssupporting

confidence: 60%

Controlled expression of transgenes introduced by in vivo electroporation

Matsuda

Cepko

2007

Proc. Natl. Acad. Sci. U.S.A.

615

601

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“…Rho expression is rod-specific and the amount of Rhodopsin protein is regulated at the level of transcription. (Treisman, et al, 1988) Transcription factors that regulate the Rho gene also regulate other rod-specific genes that share a similar expression pattern. , Lerner, et al, 2001 Rho expression varies during development, dramatically increasing during maturation of rod outer-segments.…”

Section: Discussionmentioning

confidence: 99%

FIZ1 is expressed during photoreceptor maturation, and synergizes with NRL and CRX at rod-specific promoters in vitro

Mali

Zhang

Hoerauf

et al. 2007

Experimental Eye Research

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FIZ1 (Flt-3 Interacting Zinc-finger) interacts and co-purifies with the rod-specific transcription factor NRL (Neural Retina Leucine zipper). We hypothesize that FIZ1 is part of an interface between cell-specific factors, like NRL, and more ubiquitous regulatory networks that vary the absolute expression levels of some rod-specific genes (i.e. Rhodopsin). As part of an ongoing exploration of FIZ1's role in neural retina, in vivo, we have taken the first look at FIZ1 expression in the developing mouse retina during the retinal maturation period. Using the normal C57/B6 mouse as a model, multiple approaches were used including: immunoblotting, immunohistochemistry, and quantitative real-time PCR. Functional implications of FIZ1/NRL interaction, on NRL-and CRX-mediated activation of the Rhodopsin (Rho) and cGMPphosphodiesterase β-subunit gene (PDE6B) promoters, were examined by co-transfection assays. Immunoblot analysis revealed that FIZ1 protein levels were lowest in immature mouse neural retina (P0). FIZ1 concentration increased at least ten-fold as the neural retina matured to the adult state (P21 and later). Immunohistochemical comparison of immature post-natal and mature adult retina revealed increasing FIZ1 protein in photoreceptors, the inner plexiform layer, and the ganglion cell layer. Total retinal Fiz1 mRNA content increased as the neural retina matured. The expected increase in Rho mRNA level was also monitored as a genetic marker of photoreceptor maturation. In transient co-transfection assays of CV1 cells, FIZ1 synergized with NRL to activate transcription from the Rho and PDE6B gene promoters with some differences. In the case of the Rho promoter, FIZ1 synergized when both NRL and CRX were present. With the PDE6B promoter, FIZ1 synergized with NRL alone, and the inclusion of CRX decreased this synergy.Conclusions-These findings support previous evidence that FIZ1 is present in rodphotoreceptors. (Co-immunoprecipitation from nuclear-protein extracts with rod-specific NRL) FIZ1 expression increases in the neural retina during the retinal maturation period. Additionally, in vitro experiments demonstrate that FIZ1 has the potential to significantly increase the NRLmediated activation of photoreceptor-specific promoters. While CRX is not a strong activator of the PDE6B promoter, alone or with NRL, CRX decreased the synergy of NRL with FIZ1.

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“…Opsin, the protein component of rhodopsin, is specifically expressed in retinal rod photoreceptors and in some cells of the pineal gland (37). In adult mice, 0.06% of the total retinal RNA encodes for opsin (photoreceptors constitute ∼80-90% of all cells in the retina) (38). This translates into the highest expression of any GPCR.…”

Section: Rhodopsin Expressionmentioning

confidence: 99%

G Protein-Coupled Receptor Rhodopsin: A Prospectus

Filipek

Stenkamp

Teller

et al. 2003

Annu. Rev. Physiol.

230

196

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Rhodopsin is a retinal photoreceptor protein of bipartite structure consisting of the transmembrane protein opsin and a light-sensitive chromophore 11-cis-retinal, linked to opsin via a protonated Schiff base. Studies on rhodopsin have unveiled many structural and functional features that are common to a large and pharmacologically important group of proteins from the G protein-coupled receptor (GPCR) superfamily, of which rhodopsin is the best-studied member. In this work, we focus on structural features of rhodopsin as revealed by many biochemical and structural investigations. In particular, the high-resolution structure of bovine rhodopsin provides a template for understanding how GPCRs work. We describe the sensitivity and complexity of rhodopsin that lead to its important role in vision.

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Opsin expression in the rat retina is developmentally regulated by transcriptional activation.

Cited by 88 publications

References 50 publications

Controlled expression of transgenes introduced by in vivo electroporation

Controlled expression of transgenes introduced by in vivo electroporation

FIZ1 is expressed during photoreceptor maturation, and synergizes with NRL and CRX at rod-specific promoters in vitro

G Protein-Coupled Receptor Rhodopsin: A Prospectus

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