Photoreceptor-specific nuclear receptor NR2E3 functions as a transcriptional activator in rod photoreceptors.
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.
Cheng
Hong
H
Neuroscience Graduate Program, Department of Opthalmology and Visual Sciences, University of Michigan, Ann Arbor 48105, USA.
Khanna
Hemant
H
Oh
Edwin C T
EC
Hicks
David
D
Mitton
Kenneth P
KP
Swaroop
Anand
A
D000818:Animals; D050976:Basic-Leucine Zipper Transcription Factors; D004268:DNA-Binding Proteins / Q000378:metabolism; D005136:Eye Proteins / Q000378:metabolism; D044385:GTP-Binding Protein alpha Subunits; D005982:Glutathione Transferase / Q000378:metabolism; D020962:Heterotrimeric GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism; D018398:Homeodomain Proteins / Q000378:metabolism; D006801:Humans; D051379:Mice; D056953:Nuclear Receptor Subfamily 1, Group D, Member 1; D057093:Orphan Nuclear Receptors; D011233:Precipitin Tests; D011401:Promoter Regions, Genetic; D018160:Receptors, Cytoplasmic and Nuclear / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D017948:Retinal Rod Photoreceptor Cells / Q000502:physiology*; D012243:Rhodopsin / Q000235:genetics / Q000378:metabolism; D015534:Trans-Activators / Q000378:metabolism; D014157:Transcription Factors / Q000235:genetics / Q000502:physiology*; D014158:Transcription, Genetic / Q000502:physiology*; D015293:Transducin; D020798:Two-Hybrid System Techniques
D050976:Basic-Leucine Zipper Transcription Factors; D004268:DNA-Binding Proteins; D005136:Eye Proteins; D044385:GTP-Binding Protein alpha Subunits; C488949:Gnat1 protein, mouse; D018398:Homeodomain Proteins; C064353:NR1D1 protein, human; C403073:NR2E3 protein, human; C490037:NRL protein, human; C498262:Nr1d1 protein, mouse; C485736:Nr2e3 protein, mouse; D056953:Nuclear Receptor Subfamily 1, Group D, Member 1; D057093:Orphan Nuclear Receptors; D018160:Receptors, Cytoplasmic and Nuclear; D015534:Trans-Activators; D014157:Transcription Factors; C109288:cone rod homeobox protein; D012243:Rhodopsin; D005982:Glutathione Transferase; D020962:Heterotrimeric GTP-Binding Proteins; D015293:Transducin