Photoreceptor-specific nuclear receptor NR2E3 functions as a transcriptional activator in rod photoreceptors

Abstract

NR2E3, a photoreceptor-specific orphan nuclear receptor, is believed to play a pivotal role in the differentiation of photoreceptors. Mutations in the human NR2E3 gene and its mouse ortholog are associated with enhanced S-cones and retinal degeneration. In order to gain insights into the NR2E3 function, we performed temporal and spatial expression analysis, yeast two-hybrid screening, promoter activity assays and co-immunoprecipitation studies. The Nr2e3 expression was localized preferentially to the rod, and not to the cone, photoreceptor nuclei in rodent retina. The yeast two-hybrid screening of a retinal cDNA library, using NR2E3 as the bait, identified another orphan nuclear receptor NR1D1 (Rev-erbalpha). The interaction of NR2E3 with NR1D1 was confirmed by glutathione S-transferase pulldown and co-immunoprecipitation experiments. In transient transfection studies using HEK 293 cells, both NR2E3 and NR1D1 activated the promoters of rod phototransduction genes synergistically with neural retina leucine zipper (NRL) and cone-rod homeobox (CRX). All four proteins, NR2E3, NR1D1, NRL and CRX, could be co-immunoprecipitated from the bovine retinal nuclear extract, suggesting their existence in a multi-protein transcriptional regulatory complex in vivo. Our results demonstrate that NR2E3 is involved in regulating the expression of rod photoreceptor-specific genes and support its proposed role in transcriptional regulatory network(s) during rod differentiation.

NR2E3mutations in enhanced S-cone sensitivity syndrome (ESCS), Goldmann-Favre syndrome (GFS), clumped pigmentary retinal degeneration (CPRD), and retinitis pigmentosa (RP)

“…A derepression of additional cone-specific genes, e.g., cone transducin (Gnat2), cone phosphodiesterases (Pde6c, Pde6h) cone arrestin (Arr3), but not M-opsin (Opn1mw), probably occurs in these cells from P14 on, in accordance with the reported in vitro repressor function of NR2E3 [Chen et al, 1999Corbo and Cepko, 2005;Haider et al, in press;Kobayashi et al, 1999]. However, a majority of photoreceptors represent a morphologically hybrid cell type expressing both rod and cone genes, providing circumstantial evidence that NR2E3 is essential to suppress cone-specific gene expression in mature rods Cheng et al, 2004;Corbo and Cepko, 2005;Peng et al, 2005]. Cone-specific gene expression is also suppressed in transgenic mice ectopically expressing NR2E3 under the control of the Crx promoter in photoreceptor precursor cells; instead of cones, nonfunctional rod-like photoreceptors are generated [Cheng et al, 2006].…”

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

“…NR2E3 (a nuclear receptor) is required for normal photoreceptor development, and recent studies have demonstrated NR2E3 can interact with CRX and regulate transcription at Opsin promoters. (Cheng, et al, 2004, Haider, et al, 2000 Nuclear receptors, as a family, are noted for both repression and activation effects that are mediated through Znfinger proteins. (Belandia, et al, 2002) Recent studies also indicate that the interaction of NR2E3 with NRL or CRX may not be direct and additional transcriptional regulatory proteins might be necessary for the formation of a stable complex in vivo.…”

“…We have demonstrated the usefulness of this approach to uncover protein interactions that are functionally relevant to photoreceptor-specific gene expression, including: NRL + CRX, and NR2E3 + NR1D1 (RevErb-alpha). (Cheng, et al, 2004, Mitton, et al, 2000 Peng et al, also using yeast two-hybrid traps, have demonstrated that NR2E3 can bind to CRX in vitro, supporting activation of the Rhodopsin (Rho) promoter, and There is evidence that FIZ1 can exist in both the nuclear and cytoplasmic compartments, although its only documented cytoplasmic protein-interaction is with the tyrosine-kinase domain of a receptor essential for myeloid cell development. (Mitton, et al, 2003, Wolf andRohrschneider, 1999) FIZ1 binds to FLT-3 (FMS-like receptor tyrosine kinase, FLK-2) in myeloid progenitors, and their association increases upon stimulation of cells with FLT-3-ligand.…”

In Pursuit of Synthetic Modulators for the Orphan Retina-Specific Nuclear Receptor NR2E3

“…NR2E3, also known as PNR and RNR, is an orphan NR expressed exclusively in photoreceptor cells of the adult and developing retina. [6][7][8][9][10][11] The physiological ligand for NR2E3 remains to be identified. Mutations in NR2E3 have been associated with several forms of retinal degeneration in human patients, including enhanced S-cone syndrome, [12][13][14] autosomal dominant 15,16 and recessive 17 forms of retinitis pigmentosa, Goldmann-Favre syndrome, 13 and clumped pigmentary retinal degeneration.…”